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Biology of Africanized and European honey bees, Apis mellifera, in Venezuela

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Title:
Biology of Africanized and European honey bees, Apis mellifera, in Venezuela
Creator:
Bolten, Alan B., 1945-
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English
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viii, 182 leaves : ; 28 cm.

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Subjects / Keywords:
Africanized honey bees ( jstor )
Bees ( jstor )
Colonies ( jstor )
Drone insects ( jstor )
Eggs ( jstor )
Genotypes ( jstor )
Honey bee colonies ( jstor )
Honey bees ( jstor )
Nurses ( jstor )
Oviposition ( jstor )
Africanized honeybee ( lcsh )
Dissertations, Academic -- Zoology -- UF
Honeybee -- Venezuela ( lcsh )
Zoology thesis Ph. D
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bibliography ( marcgt )
theses ( marcgt )
non-fiction ( marcgt )

Notes

Thesis:
Thesis (Ph. D.)--University of Florida, 1986.
Bibliography:
Includes bibliographical references (leaves 168-181).
Additional Physical Form:
Also available online.
General Note:
Typescript.
General Note:
Vita.
Statement of Responsibility:
by Alan B. Bolten.

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University of Florida
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All applicable rights reserved by the source institution and holding location.
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0029790578 ( ALEPH )
15300408 ( OCLC )

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BIOLOGY OF AFRICANIZED AND EUROPEAN HONEY BEES,
mellifgra, IN VENEZUELA












By

ALAN B. BOLTEN















A DISSERTATION PRESENTED TO THE GRADUATE SCHOOL
OF THE UNIVERSITY OF FLORIDA IN
PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF DOCTOR OF PHILOSOPHY





UNIVERSITY OF FLORIDA 1986















"He must be a dull man who can examine the

exquisite structure of a comb, so beautifully

adapted to Its end, without enthusiastic admiration."

Charles Darwin 1859















ACKNOWLEDGEMENTS


I would I ke to thank the members of my committee: Dr. Thomas C. Emmel, my chairman, for his continued support, guidance and encouragement throughout my graduate education; Dr. Malcolm T. Sanford for stimulating discussions and his thorough editing; and Dr. Jonathan Reiskind for his enthusiasm and helpful suggestions. I appreciate the comments made by Drs. James Nation and Frank Nordlie on the dissertation. I am also grateful to Professor Frank Robinson for introducing me to the excitement and challenges of honey bee research and management. Drs. John Harbo, Anita Collins and Tom Rinderer were excellent field companions, sharing their knowledge of honey bee research techniques, and creating a stimulating research environment, both in Venezuela and during my work in Baton Rouge. I particularly want to thank Dr. John Harbo for the Instrumental inseminations and acknowledge his collaboration on both the bee size and egg laying rate experiments. I would also like to thank Dr. Orley Taylor for giving me the opportunity to study Africanized honey bees.

This research was supported by the U.S. Department of Agriculture Cooperative Agreement No. 58-7B30-8-7 with the University of Kansas (0. R. Taylor, principal investigator). The Mlnlsterio de Agricultura y Cria de Venezuela provided research facilities near Maturin. I would like to thank Med. Vet. Ricardo Gomez Rodriguez for his hospitality and



III









logistic support. Laboratory facilities at the Universidad de Oriente In Jusepin were made available by Professor Dick Pulido.

The research presented In this dissertation and the commitment to complete the writing could not have been accomplished without the collaboration, companionship, encouragement and Insights of my wife, Karen Bjorndal, who shared not only the excitement and successes but also the frustrations and discomforts of Africanized honey bee research.

Finally, I would like to thank my parents, who have always supported and encouraged my work.




































iv















TABLE OF CONTENTS

EAae

ACKNOWLEDGEMENTS . . . . . . . . . . . . . III

ABSTRACT . . . . . . . . . . . . . . . vil

CHAPTERS

I INTRODUCTION . . . . . . . . . . . . 1

Evolutionary Origin and Distribution of Honey Bees . . 1
Importation of African Honey Bees Into Brazil and
Their Dispersal Throughout South and Central America I
Characteristics of Africanized Honey Bees . . . . 3 Purpose of My Research . . . . . . . . . 6
Identification of Honey Bees Used in My Research . . . 7
When and Where Research Was Conducted . . . . . 7

II WORKER BEE DEVELOPMENT TIMES . . . . . . . . 9

Introduction . . . . . . . . . . . . 9
Methods . . . . . . . . . . . . . 14
Results . . . . . . . . . . . . . 18
Discussion . . . . . . . . . . . . 20

III INTERACTION OF MATERNAL GENOTYPE, EGG GENOTYPE AND COMB
CELL SIZE ON HONEY BEE WORKER SIZE AND SIZE VARIATION . 35

Introduction . . . . . . . . . . . . 35
Methods . . . . . . . . . . . . . 38
Results . . . . . . . . . . . . . 41
Discussion . . . . . . . . . . . . 42

IV QUEEN DEVELOPMENT AND MATURATION . . . . . . . 55

Introduction . . . . . . . . . . . . 55
Methods . . . . . . . . . . . . . 57
Results . . . . . . . . . . . . . 63
Discussion . . . . . . . . . . . . 65






v









V QUEEN PUPAL WEIGHTS . . . . . . . . . . 79

Introduction . . . . . . . . . . . . 79
Methods . . . . . . . . . . . . . 80
Results . . . . . . . . . . . . . 83
Discussion . . . . . . . . . . . . 84

VI EGG LAYING AND BROOD PRODUCTION RATES
DURING THE FIRST BROOD CYCLE . . . . . . . . 92

Introduction . . . . . . . . . . . . 92
Methods . . . . . . . . . . . . . 96
Results . . . . . . . . . . . . . 101
Discussion . . . . . . . . . . . . 102

VII SUCCESSFUL HYBRIDIZATION BETWEEN AFRICANIZED
AND EUROPEAN HONEY BEES IN VENEZUELA WITH
IMPLICATIONS FOR NORTH AMERICA . . . . . . . 118

Introduction . . . . . . . . . . . . 118
Methods . . . . . . . . . . . . . 123
Results . . . . . . . . . . . . . 124
Discussion . . . . . . . . . . . . 125

VIII DISCUSSION: FACTORS CONTRIBUTING TO THE SELECTION ADVANTAGE OF AFRICANIZED HONEY BEES IN SOUTH AMERICA-THE RESOURCE UTILIZATION EFFICIENCY HYPOTHESIS . . . 133

Success of Introduced Populations of Honey Bees . . . 133 Factors Affecting Honey Bee Reproductive Rates . . . 135
Factors Contributing to the Selective Advantage
of Africanized Honey Bees in South America . . . 140
Potential Impact of Africanized Honey Bees
In North America . . . . . . . . . . 153

APPENDICES

A WORKER BEE DEVELOPMENT TIMES AND MORTALITY
DURING DEVELOPMENT . . . . . . . . . . 156

B HONEY BEE SIZE, COMB CELL SIZE AND
SIZE VARIATION . . . . . . . . . . . 159

C CHANGES IN QUEEN PUPAL WEIGHT WITH AGE . . . . . 165

D ACCURACY OF TECHNIQUE USED TO ESTIMATE
NUMBER OF BEES IN A COLONY . . . . . . . . 167

LITERATURE CITED . . . . . . . . . . . . . 168

BIOGRAPHICAL SKETCH . . . . . . . . . . . . 182


vi















Abstract of Dissertation Presented to the Graduate School of the University of Florida in Partial Fulfillment of the
Requirements for the Degree of Doctor of Philosophy

BIOLOGY OF AFRICANIZED AND EUROPEAN HONEY BEES,

A mellifera, IN VENEZUELA

By

Alan B. Bolten

August 1986

Chairman: Thomas C. Emmel
Major Department: Zoology

To determine factors responsible for the greater success of

Africanized honey bees, ftL. mellifera, in tropical regions of South

America, demographic parameters affecting colony reproductive rates were evaluated for Africanized and European honey bees under identical conditions in Venezuela. Worker bee development time was evaluated as an interaction between egg genotype, comb cell size and nurse bee genotype. Africanized worker bees developed faster than European bees: 18.9 and 19.8 days, respectively. There was no significant effect of comb cell size or nurse bee genotype. Mortality for different developmental stages was recorded. The relationship of worker bee development time to colony growth rate is discussed.

Africanized queens develop in 14.5 days post-oviposition compared with 15.0 days for European queens. Queen pupal weights were not significantly different. Post-emergence maturation rates for Africanized and European queens were similiar as determined by both the vii









ages when queens attracted drones and the ages when oviposition was initiated.

Daily egg laying rates and brood production during initial colony growth were not significantly different for Africanized and European queens. Africanized and European worker bees did not differentially affect egg laying and brood production rates.

Differences in reproductive rates between Africanized and European honey bees In South America cannot be attributed to differences in intrinsic demographic factors. A hypothesis based on differences in resource utilization efficiency is presented to explain the success of Africanized bees compared with European bees in South America.

Results from reciprocal F, crosses indicate that bee size is a function of egg genotype, comb cell size and maternal genotype. The importance of maternal inheritance for reducing worker bee size variation within a colony Is discussed. Advantages of smaller worker bee size are evaluated for Africanized bees.

There are no effective reproductive isolating mechanisms operating between Africanized and European honey bee populations. Both Africanized and European queens mated with equal success with Africanized drones as measured by the numbers of spermatozoa in the spermatheca. The potential impact of Africanized bees on North America is analyzed with respect to hybridization and genetic introgression, resource competition, and selection advantages for European bees in temperate regions.






viii
















CHAPTER I
I NTRODUCTION


Evolutionary Origin and Distribution of Honey Bees

Honey bees of the genus A.j-j have their greatest diversity in Asia (Michener 1979). Earliest fossil evidence for the genus is from Oligocene deposits in Europe (Zeuner and Manning 1976). The evolutionary relationships of the four generally recognized species of are reviewed by Michener (1974). Three of the species (A. cerana, A. dorsaa and A. flora) are native only to Asia (Michener 1979; Ruttner 1975). The western honey bee (A. mellifera) is native to Africa, western Asia and Europe and may have evolved in tropical or subtropical Africa (Wilson 1971) or the Near East (Ruttner 1975). The widely different climatic conditions and floral resources under which populations of A. mellifera evolved have resulted in a number of geographically recognizable subspecies (Alpatov 1929, 1933; Br. Adam 1966; Dupraw 1965; Ruttner 1968, 1975, 1976a, 1976b; Smith 1961; Wafa,

Rashad and Mazeed 1965).

Importation of African Honey Bees into Brazil and Their Dispersal

Throughout South and Central America

European honey bees (A. mellifera mellifera and A_. a. li.ustica)

had been introduced into Brazil by 1845 (Gerstaker cited in Pellet 1938; Woyke 1969). A. ga. mellifera is native to Europe in the regions west and north of the Alps and extending east into Central Russia; A_. m.

I







2

ligustica is native to the Italian peninsula (Ruttner 1975). Because these European honey bee populations were not very successful in tropical and subtropical habitats of Brazil (Michener 1972), researchers believed that they could Improve Brazil's honey production by breeding a honey bee better adapted to local conditions (Woyke 1969). With this intention, honey bee queens from South Africa (A. m. scutellata, formerly classified as adansonii, see Ruttner 1976a, 1976b, 1981) were imported into southeastern Brazil in 1956 (Kerr 1967). The following year, swarms escaped and hybridized with established European honey bees. The descendents from this hybridization are known as Africanized honey bees (Goncalves 1982). Details of the introduction and subsequent spread throughout South America have been extensively reviewed (Goncalves 1974, 1975, 1982; Kerr 1967; Michener 1972, 1975; Taylor 1977, 1985; Taylor and Williamson 1975; Woyke 1969).

In the 30 years since African honey bees were imported into

southeastern Brazil, their hybridized offspring have rapidly dispersed throughout tropical South and Central America and are now as far north

as Honduras and El Salvador (Rinderer 1986). The dispersion from their original importation site into new areas has been rapid--200-500 km per year (Taylor 1977, 1985; Winston 1979a). As Africanized honey bees have colonized new areas, they have achieved dramatic population densities (Michener 1975). There are now probably more than ten million feral colonies in South and Central America (Winston, Taylor and Otis 1983). Their success in these new habitats, compared with the lack of success of European honey bee populations, may be attributed to their foraging behavior which is more suited to the resource patterns of the tropics (Nunez 1973, 1979a, 1982; Rinderer, Bolten, Collins and Harbo 1984;







3

Rinderer, Collins and Tucker 1985; Winston and Katz 1982). As a result of both foraging success and the length of time throughout the year that resources are available in the tropics, Africanized honey bees have a high annual reproductive rate, which is responsible for both their rate of dispersal into new areas and their high colony densities. Net reproductive rates for Africanized bees are estimated to be 16 colonies per colony per year based on demographic data collected in French Guiana (Otis 1980, 1982a), compared with 0.92-0.96 (Seeley 1978) or, when afterswarms are considered, 3-3.6 (Winston 1980a; Winston, Taylor and Otis 1983) colonies per colony per year for European honey bees in North America.


Characteristics of Africanized Honey Bees

The most well known characteristic that differentiates Africanized honey bees from European honey bees is their stinging behavior (Collins, Rinderer, Harbo and Bolten 1982; Stort 1974, 1975a, 1975b, 1975c, 1976). Because of their stinging behavior, Africanized bees are a health hazard for both humans and domestic animals (Taylor 1986). Collins, Rinderer, Harbo and Bolten (1982) compared the colony defense behavior of Africanized honey bees in Venezuela with European bees under identical conditions in Venezuela and with a population of European bees in Louisiana, U.S.A. Africanized honey bees responded more rapidly and in much greater numbers, resulting in 5.9 times as many stings in a target compared with European honey bees in Venezuela and 8.2 times as many stings compared with European bees in Louisiana. Two additional components of Africanized honey bee defense behavior increase their potential as a health hazard. Compared to European bees, Africanized bees pursue a source of disturbance for a greater distance (160 versus







4

22 meters) and remain disturbed for a greater period of time (28 versus

3 minutes) (Stort 1971 cited in Goncalves 1974). Differences in defense behavior between Africanized and European bees do not appear to be a function of either quantitative differences in pheromone production (Crewe and Hastings 1976) or numbers of olfactory structures on the antennae (Stort and Barelli 1981).

There is a difference in natural comb cell size between Africanized and European populations. The width between opposite sides of the hexagonal cells for the Africanized population in Brazil averaged 5.0 mm compared with 5.4 mm for the European population in Canada (Michener 1972). In a recent study, cells built by Africanized swarms in Venezuela were 4.8-4.9 mm wide and those built by European swarms in Louisiana, U.S.A. were 5.2-5.3 mm wide (Rinderer, Tucker and Collins 1982). Adult Africanized bees are smaller than European bees (62 mg compared with 93 mg, unengorged) (Otis 1982b; Otis, Winston and Taylor 1981). However, as Africanized honey bees disperse into areas with extensive European honey bee populations, size differences between the two populations may become less distinct. Increased hybridization between the two populations could result in bees with an Africanized genome developing in European comb cells, resulting in larger Africanized bees. Therefore, methods used to identify Africanized honey bees based on size parameters, for example, morphometric analysis, may become less reliable. As Daly, Hoelmer, Norman and Allen (1982) point out, there is a "difficulty in using phenotype characters to identify genetically different, but closely related populations" (p. 593). This will be more evident as Africanized honey bees disperse into areas of

Central America and particularly Mexico, where large populations of







5

European honey bees exist. Factors determining honey bee size and potential problems of Africanized honey bee identification based on size are analyzed in Chapter III.

The cuticular hydrocarbon composition of Africanized honey bees is significantly different from that of European honey bees (Carlson and Bolten 1984). The differences are particularly striking for the 35, 37, 39, 41 and 43 carbon alkenes and alkadienes that total over 22% of the hydrocarbons extracted from Africanized bees but only 1-3% of the hydrocarbons extracted from European bees. Because hydrocarbon composition is not affected by honey bee size or diet, using hydrocarbon analysis to distinguish between Africanized and European honey bees has great potential. However, further research to determine heritability patterns for different hydrocarbon components is needed.

Differences between Africanized and European honey bees have also been demonstrated for foraging behavior (Nunez 1973, 1979a, 1982; Rinderer, Bolten, Collins and Harbo 1984; Rinderer, Collins and Tucker 1985; Winston and Katz 1982), egg development times (Harbo, Bolten, Rinderer and Collins 1981), selection preferences for nest cavity sizes (Fletcher 1976; Michener 1972; Rinderer, Collins, Bolten and Harbo 1981; Rinderer, Tucker and Collins 1982), hoarding behavior (Rinderer, Bolten, Harbo and Collins 1982), worker bee longevity (Winston and Katz 1981), morphometric analysis (Daly and Balling 1978), and allozyme patterns (Nunamaker and Wilson 1981; Sylvester 1982). Africanized honey bee populations in South America are reported to have a high colony reproductive (swarming) rate compared with European honey bee populations in North America (Otis 1980, 1982a; Winston 1979b, 1980a; Winston, Dropkin and Taylor 1981; Winston, Taylor and Otis 1983).







6

However, those investigations have not been conducted under similar environmental or experimental conditions. Therefore, comparisons of reproductive rates between Africanized and European honey bees using those data are Inappropriate for either Identifying differences in reproductive rates for tropical and temperate honey bee populations or for identifying factors responsible for the success of Africanized bees In tropical regions.


Purpose of My Research

African and European honey bee populations evolved under very different resource and climatic conditions. The presence of both Africanized and European honey bees In Venezuela provided the opportunity to study both populations under identical conditions in the tropics. Differences between the two honey bee populations that make Africanized bees more successful in tropical regions could then be evaluated. The underlying assumption of my research was that the life history of Africanized honey bee populations in South America (as well as the parental population in Africa) is characterized by a high

reproductive rate. Demographic features expected to be correlated with this high rate of colony reproduction include short worker bee development times, small worker bee size, rapid queen development and maturation, and Increased egg laying and brood production rates. Predictions involving these demographic characteristics led to a series of experiments that are presented and discussed In the following chapters.

In addition, the question of reproductive Isolation versus

hybridization and differential selection between the two populations in tropical conditions was experimentally evaluated. Whether there is







7

hybridization or reproductive isolation between Africanized and European honey bee populations could result in very different scenarios for the potential impact of Africanized honey bees on North America, particularly the U.S.A.


Identification of Honey Bees Used in Mv Research

For the experiments presented here, Africanized honey bee colonies were established from queens removed from feral colonies in an area in eastern Venezuela where there were no known European honey bees. They were identified as Africanized bees primarily by their distinctly smaller comb cell size as compared with European honey bees.

European honey bees used in the experiments were from commercially produced queens from three different queen breeders in the U.S.A. Additional European lines were obtained from the U.S. Department of Agriculture Bee Research Laboratories in Madison, Wisconsin, and Baton Rouge, Louisiana. All of these European queens were either naturally mated or instrumentally inseminated in the U.S.A. and then shipped to Venezuela.


When and Where Research Was Conducted

All field research with Africanized and European honey bees was

conducted from December 1978 through February 1980 at the Ministerio de Agricultura y Cria de Venezuela Africanized Honey Bee Research facilities near Maturin, Monagas. The area originally was a Tropical Dry Forest [sensu Holdridge Life Zone System (Holdridge 1964; Ewel and Madriz 1968)]. The forest had been partially cleared, and the area was grazed by cattle.







8

All Africanized and European honey bee comparisons were made at the same time under identical experimental conditions. Field and experimental methods are described for each of the experiments in the appropriate chapters.

A few experiments with European honey bees only were conducted in the U.S.A. to confirm techniques developed and used while In Venezuela. These experiments were undertaken either at the U.S. Department of Agriculture Bee Breeding and Stock Center Laboratory in Baton Rouge, Louisiana, or at the bee research facilities of the Institute of Food and Agricultural Sciences at the University of Florida, Gainesville.















CHAPTER II
WORKER BEE DEVELOPMENT TIMES




The presence of both Afnicanized and European honey bees,

mellifera, in South America allows for comparisons to be made under Identical conditions between a population that has evolved in the tropics and one that has evolved in temperate regions. Africanized honey bee populations in South America are reported to have a high colony reproductive (swarming) rate compared with European honey bee populations in North America (Otis 1980, 1982a; Seeley 1978; Winston 1979b, 1980a; Winston, Dropkin and Taylor 1981; Winston, Taylor and Otis 1983). However, these Investigations have not been conducted under similar environmental or experimental conditions. Therefore, comparisons of reproductive rates between Africanized and European honey bees using these data are inappropriate either for identifying differences in reproductive rates between tropical and temperate honey bee populations or for identifying factors responsible for the success of Africanized bees in tropical regions.

Reproductive rates in honey bees are a function of colony growth rates which are a result of an interaction of at least three factors:

resource availabil itys resource util izat ion efficiency (foraging success,. brood production efficiency, and bee size), and colony demographic parameters. Worker bee longevity is the only demographic


9









parameter that has been compared between Africanized and European bees under identical conditions. The greater longevity of European honey bees (Winston and Katz 1981) gives European bees a colony growth rate advantage. Other demographic characteristics that affect reproductive rates of Africanized and European honey bees (for example, worker bee development times, brood mortality, queen development and maturation periods, queen fecundity and brood production rates) have not been evaluated for Africanized and European bees under similar conditions. As part of a larger study evaluating these demographic parameters, this study compares worker bee development periods for Africanized and European honey bees in Venezuela.

Smith (1958a) and Tribe and Fletcher (1977) reported that the total development period (from oviposition to adult emergence) for worker bees of mlellifera aansnti (now classified as A. mL. scutellata; Ruttner 1976a, 1976b, 1981) from South Africa was between 18.6-20 days. Similar development times for the Africanized honey bee populations (descendents of A. M. scutellata) in Brazil have been presented (Kerr, Goncalves, Blotta and Maciel 1972; Wiese 1972). Worker bee development times for European populations (primarily A. m. mellifera, ligustica, carnica and caucasica) from Europe and North America range from 20-24 days (Jay 1963).

The differences in development times between African (and

Africanized) and European genotypes, which range from 1.4 to 5.4 days, are difficult to evaluate because they are based on data collected under very different experimental conditions. Jay (1963) summarized a number of factors that affect development times: seasonal variation in temperature; temperature differences in different areas within the brood









nest; colony size, which affects both brood nest temperature and feeding frequency and quality; and nectar and pollen resources,. which also affect feeding quantity and quality. Valid comparison of development times between genotypes or populations can only be made when these factors are controlled under similar experimental conditions.

The Importance of slight temperature differences on development time cannot be overstated. Development times for European worker bees in Wisconsin, U.S.A., averaged 20.5 days but ranged from 20-24 days, depending on differences In temperature In different areas of the brood nest (Milum 1930). Harbo and Bolten (1981) showed that fertilized eggs kept at 34,.80C hatched about 1.4 hours sooner than those kept at 34.30C. This difference in egg hatch time for only a 0.50C difference in temperature can be extrapolated to approximately 10 hours for the entire development period [calculated from Harbo and Bolten (1981)]. However, normal temperatures within a brood nest can vary to a much greater extent (Milum 1930; Jay 1963). For example, when Tribe and Fletcher (1977) determined the development rates of African honey bees in South Africa, they recorded that temperatures in the brood nest varied from 26-340C.

In an incubator with controlled temperature and humidity, eggs from Africanized genotypes hatched significantly sooner than eggs from European genotypes, 69.6 +L 1.06 hours compared with 73.3 +L 1.14 hours (Harbo, Bolten, Rinderer and Collins 1981). Egg development requires that only temperature and humidity be controlled and can therefore be evaluated independently of colony-level parameters. However# differences between Africanized and European genotypes for total worker bee development periods need to be evaluated within a colony In order to







12

allow for normal feeding and growth. Worker bee development rates are a result of an interaction between the egg genotype and the colony. There are three colony-level factors that need to be considered when comparing total development time of Africanized and European worker bees.

First is the effect of comb cell size. There Is a difference in natural comb cell size between Africanized and European populations. The width between opposite sides of the hexagonal cells for the African population In Africa measured 4.77-4.94 mm (Smith 1958a). Cells for the Africanized population In Brazil averaged 5.0 mm (range 4.8-5.4 mm) (Michener 1972), but cells of the Africanized population in Venezuela averaged 4.8 mm (range 4.5-5.0 mm) (Chapter III; Rinderer, Tucker and Collins 1982). Cells from the European population from Ontario, Canada, averaged 5.4 mm (range 5.2-5.7 mm) (Michener 1972), and those from Louisiana,. U.S.A., averaged 5.2-5.3 mm (range 5.2-5.4 mm) (Rinderer, Tucker and Collins 1982). Adult bee size is a function of comb cell size (Grout 1937); adult Africanized bees are smaller than European bees (62 mg compared with 93 mg, unengorged) (Otis 1982b; Otis, Winston and Taylor 1981).

Abdellatif (1965) suggested that larvae in smaller comb cells received less food which caused them to elongate and become sealed

earlier. Also,. Tribe and Fletcher (1977) suggested that the difference In development time for African and European genotypes may be a function of the small African bee size. Therefore, the effect of comb cell size needs to be considered when comparing development times of Africanized and European honey bees.

The second colony-level factor is the effect of nurse bee genotype. There may be behavioral differences and/or physiological differences in







13

the way in which nurse bees from the two populations interact with the developing larvae. Mellnichenko (1962) suggested that differences between nurse bee genotypes might affect developmental rates as well as size of developing larvae. For European honey bees, Lindauer (1953) calculated that each developing larva requires over 2785 adult bee visits taking a total of 10.3 hours. This appears to provide sufficient opportunity for possible genotype differences, either quantitative or qualitative, to affect development rates. In addition to potential qualitative or quantitative differences In feeding of larvae, nurse bees of different genotypes may also maintain different brood nest temperatures. Therefore, development times for Africanized and European

worker bees were evaluated in both Africanized and European colonies.

The third colony-level factor affecting worker development is

colony size (number of worker bees in a colony). Colony size affects 'both brood nest temperature and larval feeding rates, which, as already discussed, are two major factors affecting development times.

In addition to these colony-level parameters, resource conditions also affect development time. Nelson and Sturtevant (1924) reported that development of European bees was more rapid with increased larval feeding associated with a nectar flow. Ribbands (1953) and Jay (1963) both summarized evidence of the effect of food on larval development rates. Therefore, all comparisons of worker development times were conducted simultaneously to avoid any differences due to resource conditions.

This paper reports the results from a comparison of the development times of Africanized and European honey bees under identical conditions in Venezuela. The experimental design allowed for the discrimination







14

between the effects of egg genotype and the colony-level parameters of comb cell size and nurse bee genotype on worker bee development time. These experiments were conducted during July-October 1979.


Methods5

Table 2-1 summarizes the experimental design. Four experimental colony treatments were established as follows:

I. Africanized comb cell size, Africanized nurse bees (A55)

ii. Africanized comb cell size, European nurse bees (H2)

iii. European comb cell size, Africanized nurse bees (A41)

iv. European comb cell size, European nurse bees (IBR877).

Each experimental colony was a five-frame hive (22 liters) with

four empty combs and one comb with honey and pollen and approximately 2 kg of young adult bees (Africanized or European, depending upon treatment). Because natural nectar and pollen resources were available Irregularly throughout the experimental period (16 weeks), the colonies were supplemented with honey and pollen as necessary.

European comb was built from commercially-produced beeswax foundation that had been fastened into standard wooden frames. Africanized comb was naturally built (not from foundation) by Africanized bees in empty standard wooden frames to facilitate manipulation and colony inspection.

The queens in colonies with Africanized nurse bees (A55 and A41) were Africanized queens produced by standard queen rearing methods (Laidlaw 1979) and then naturally mated to Africanized drones. Mating occurred In an area of eastern Venezuela that had a large feral population of Africanized honey bees with no known European honey bees present (near San Jose de Buja, Monagas, Venezuela). The feral colonies







15

from which the Africanized queen mothers were extracted were also from this area. The colonies were identified as Africanized honey bees by both their behavior and their small comb cell size characteristic of the Africanized population (4.5-5.0 mm, see Chapter III).

The queens in the colonies with European nurse bees (H2 and IBR877) were European queens that had been mated to European drones In the U.S.A. and transported to Venezuela. Line H2 was from a commerical queen producer in the southeastern U.S.A.; IBR877 was an outbreed line from the U.S. Department of Agriculture Bee Breeding and Stock Center Laboratory in Baton Rouge, Louisiana, U.S.A.

The source for the Africanized egg genotype (A26) was a queen

removed from a feral colony of Africanized honey bees in the San Jose de Buja area. The colony was identified as Africanized by its behavior and characteristic comb cell size. The source of the European egg genotype

(Y5) was a queen commercially produced in the southeastern U.S.A. and

shipped to Venezuela.

Because adult longevity Is from 2 to 5 weeks for European honey

bees (Woyke 1984) and 2 to 3 weeks (or less) for Africanized honey bees (Winston 1979b; Winston and Katz 1981)o experimental colonies were established 10 weeks prior to the start of the experiment. This was sufficient time to Insure that at the beginning of the developmental trial all the adult bees present within the experimental colonies had developed in those colonies, and, therefore, were offspring of a known genetic line having developed within a known comb cell size. Before development times were measured, the worker bee populations in the experimental colonies were equalized as much as possible by removing random samples of bees from the most populous colonies.







16

The experimental colonies were placed In an apiary under a roof

with completely open sides. The roof served two purposes. Firsts. the colonies were in complete shade, which reduced any effects from differences In ambient temperature and sunlight. Second, colonies could be opened and inspected In order to monitor development with a minimum of disturbance, especially during rain.

Eggs were collected from queens of the two designated egg source lines (A26 and Y5)p that were established in modified colonies similar to those used by queen producers in the U.S.A. (Harp 1973). These colonies consisted of five standard frames with the middle frame isolated from the four others by queen excluder side and top panels. The excluder panels have a mesh size that restricts the queen from passing through because of her wider thorax., but allows worker bees to pass through to feed and communicate with the queen. Thus, the queen was isolated on a specific comb so that eggs could be collected that would then be placed into one of the four experimental colonies to evaluate development time.

The comb used had the appropriate comb cell size for the

experimental colony into which it would be placed. Comb cell size was measured in each experimental nurse bee colony and for each Africanized and European egg comb put into each experimental colony.

The queens were caged on each comb for 24 hours so that a large uniform egg sample could be collected. Eggs were monitored only from the center of each frame, which insured a more uniform temperature during development as well as uniform brood nest position. The large egg sample also insured that the monitored eggs were in a normal environment surrounded by similarly-aged developing bees. The thirty







17

or forty eggs selected to be monitored for development formed either a 3x10 or a 4x10 cell area. A reference point which facilitated locating the designated development sample was indicated by colored pins inserted five cells to the left of each original egg row.

The two combs (one comb from each egg source) were placed In the

center of each experimental colony at the same time. The cells with the eggs to be monitored faced each other in order to reduce any effects of brood nest position. Two frames of brood were removed from each experimental colony to make room for the experimental frames. This also reduced the amount of brood being reared in each experimental colony insuring that the monitored eggs would be optimally fed.

The pairs of frames were put into the four experimental colonies on four successive days because of the 24 hours needed to collect each set of eggs. Once the eggs were put into the experimental colonies they were inspected every day at 0800 hours. The survivorship and developmental status of each original test group egg was recorded.

The sample size of eggs monitored was selected to minimize the time each colony would need to be opened for observation in order to minimize disturbance. When a colony was disturbed.. bees flew from the comb and ran on the bottom of the hives resulting in temperature fluctuations and interrupted feeding of larvae. Colonies were carefully opened, using minimal amounts of smoke. Adult bees were not shaken off the combs but rather gently pushed aside to observe the development stage within the cells. Inspections more frequent than every 24 hours also increased the level of disturbances especially In the Africanized colonies (A55 and A41). The advantage of more frequent monitoring to more accurately







18

record time of developmental changes was outweighed by the negative effects of disturbance on development rates.

Temperature of the brood nest in the space between the two

monitored combs was recorded periodically 4~y placing a thermometer into that area through a hole in the hive cover. This was only an approximate measure of temperature because development occurs within the cells where temperature is less affected by the ventilating air currents within the hive.




Table 2-2 presents the development times for unsealed brood, sealed brood, and total development times from oviposition to adult emergence for Africanized and European egg genotypes in each of the four experimental treatments. Table 2-3 summarizes the argumentation for evaluating the interactions of egg genotypes with the colony-level parameters of comb cell size and nurse bee genotype on worker bee development times. The best tests to use to compare differences between the Africanized and European egg genotypes are evaluating pairs in each of the four experimental nurse bee colonies (A55P H2, A41, and 1BR877)s i.e., AxB,. CxD, ExF, and GxH. In these comparisons,, the colony-level parameters (colony population, position within the brood nest, temperature,. comb cell size, and nurse bee genotype) are identical and allow for only differences in egg genotype to be compared. Tables 2-4, 2-5 and 2-6 summarize the results from the statistical analyses for the unsealed brood, sealed brood,. and total development times, respectively. Data were analyzed with the Kolmogorov-Smirnov one-tailed test using the chi square distribution, df =2 (Siegel 1956).









Africanized worker bees developed faster than European bees (ACEG x BDFH). The unsealed larval period was 4.3 + 0.4 days compared with 4.9 + 0.4 days, P<0.001; the sealed larval and pupal period was 11.6 + 0.5 days compared with 11.9 + 0.4 days, P<0.01; and the total development time was 18.9 +t 0.3 days compared with 19.8 +L 0.4 days, P<0.001. There was no significant effect of comb cell size or nurse bee genotype on development times. These differences in total development time are similar to the differences found between three different lines of Africanized and three different lines of European honey bees compared in another study (19.2 days compared with 20.0 days, Table A-i).

Comb cell sizes for the experimental colonies and egg sample frames are presented in Table 2-7. Temperatures recorded for all experimental colonies varied from 35-36oC.

When differences in development times between Africanized and

European egg genotypes were compared for each stage of development, the greatest difference was observed in the unsealed larval stage (Table 28). However, this was not a result of a differential acceleration of development during the unsealed larval period for the Africanized honey bees. The proportion of unsealed larval development time to total development time and the proportion of sealed brood development time to total development time were compared for the Africanized and European honey bee populations following angular transformations of the proportions (Sokal and Rohlf 1969). These proportions were not significantly different between the Africanized and European honey bees.

The differences recorded for the unsealed brood stage between the Africanized and European honey bees may be an artifact of the experiment for two reasons. First, the 24-hour observation interval may obscure







20

exact timing of developmental changes. Second, the process of sealing is not a precise developmental stage and may take from six hours (Lindauer 1953) to 24 hours (Jay 1963). When the unsealed and sealed brood stages are combined,. the proportional differences between the two populations are the same as for egg development times and total development times (Table 2-8).

Mortality for different developmental stages for each colony

treatment Is presented in Table 2-9 (see also Table A-2). Mortality was high (26-37%) for larvae in the experimental colony (H2) with European nurse bees on Africanized comb cell size. The high mortality during the larval stage may be a result of the reduced ability of larger European nurse bees to feed the developing larvae In smaller Africanized comb cells. There was also a high egg mortality recorded for European eggs in A41 and 1BR877--34 and 70%, respectively. Woyke (1977) reports normal mortality may be as high as 10-50% depending on the season. Garofalo (1977) also reports varying mortalities depending on both the size of the colony and the time of year: eggs 10-25%p larvae 11-37%p pupae 5-7%, and all developmental stages combined 25-53%.




This study is the first to evaluate worker bee development times between Africanized and European honey bees as an interaction between egg genotype and colony-level parameters. Differences In worker bee development times were independent of the colony-level parameters of comb cell size and nurse bee genotype but were dependent on egg genotype differences between the Africanized and European populations. The difference In development times between these two populations was not as







21

large as expected from previous reports, which underscores the importance of making comparisons under identical conditions.

The proportional difference (5.7%) In egg development times between Africanized (A26) and European (Y5) honey bees reported by Harbo, Bolten, Rinderer and Collins (1981) is identical to the proportional difference in total development time reported In the present study (5.7%, see Table 2-8). Egg development time is a function of the Inherent rate characteristic of the particular genotype because colonylevel parameters (e.g., feeding) are not involved (Harbo, Bolten, Rinderer and Collins 1981). Using egg development to evaluate differences in total development between genotypes (or populations) is advantageous because egg development times are easier to evaluate, take less time, have fewer variables to control (temperature and humidity only), and can be evaluated in an incubator rather than in a colony, avoiding problems associated with disturbing the colony during observations. It must be noted, however, that by using egg development times one can only extrapolate proportional differences between genotypes for total development time but cannot extrapolate the absolute

total development time.

A prerequisite for high reproductive rates would be a rapid colony growth rate. However, the importance of worker development time to the rate of colony growth (increase in numbers of bees in a colony) has apparently been misunderstood, e.g., see Fletcher (1977as 1978), Fletcher and Tribe (1977a), Tribe and Fletcher (1977), Winston (1979b), Winston, Dropkin and Taylor (1981), Winston and Katz (1982), Winston,. Taylor and Otis (1983). The difference in worker development times observed for African ized and European honey bees is not a factor







22

contributing to either differences In rate of colony population Increase or to differences in reproductive rates between the two honey bee populations.

The importance attributed to worker development time on the rate of colony growth may be a result of confusing colony population increase (increase in the number of bees in the colony) with general population growth models designed for other species in which all Individuals are potential reproductive. For honey bees, individual (or worker bee) development time is not equal to generation time. Organism growth models must be used to evaluate colony growth even though the number of Individual worker bees within the hive increases. The hive is the organism. Worker bee development time does not affect the rate of colony growth. Worker development time affects only the length of time between a given change in egg laying rate and its resulting change in population Increase or decrease. Africanized bees develop in 19 days and begin their population increase (=growth) on the 19th day of the colony cycle, compared with the 20th day for European bees. This difference is trivial compared to potential differences from other demographic factors that do affect rates of colony growth. Egg laying and brood production rates, worker bee longevity, brood mortality,, and resource availability are factors that do affect the rate of colony population increase ando therefore affect the reproductive rates.

Tribe and Fletcher (1977) have suggested that African worker bees have a shorter unsealed development stage because they do not grow as large as European honey bees. They compare their data for African bees with data for European bees in the literature and conclude that African bees have a 20-30% shorter unsealed larval stage. There are four







23

problems with their analysis. First, as already pointed out,. using the duration of the unsealed stage has inherent problems because It is not a precise development stage. Second, comparisons based on data collected under different experimental conditions are not valid. Third, their logic is perhaps circular with respect to the question of larval size and larval development times. In the present study, development time was not size-related for either Africanized or European honey bees. Africanized honey bees that developed In European comb had the same development times as those that developed in Africanized comb even though Africanized bees reared in European comb were significantly larger (16%; Chapter III). The same relationship was true for European honey bees with a 17% increase In size of bees from European comb compared with bees from Africanized comb. And fourth, their comparison is In itself incorrect. Rather than compare the differences In unsealed development times between African and European populations to determine If the African population has a relative shorter duration as unsealed larvae, they should have used the proportion of unsealed development period to total development period in order to compare African and European populations. In the present study, the relative times spent as an unsealed larvae to the total development time for both the Africanized and European genotypes were not significantly different. The differences in development time between Africanized and European populations appear constant throughout development without any

developmental acceleration during the larval stage for either Africanized or European honey bees.







24

TABLE 2-1. Experimental matrix for evaluating Interaction of egg
genotype, comb cell size, and nurse bee genotype on worker
development times. A H represent each treatment.


AFRICANIZED EUROPEAN
EGG GENOTYPE (A26) EGG GENOTYPE (Y5) AFRICANIZED COMB CELLS

AFRICANIZED NURSE
BEES (A55) A B

EUROPEAN NURSE
BEES (H2) C D


EUROPEAN COMB CELLS

AFRICANIZED NURSE
BEES (A41) E F

EUROPEAN NURSE
BEES (IBR877) G H







25

TABLE 2-2. Interaction of egg genotype, comb cell size, and nurse bee genotype on worker bee development time (days): median, (range), mean +SD, (n = sample size).


AFRICANIZED EGG GENOTYPE (A26)


USa SBb TDTc


AFRICANIZED COMB CELLS

4.0 12.0 19.0
AFRICANIZED NURSE (4-5) (11-12) (18-20)
BEES (A55) 4.2 + 0.4 11.6 + 0.5 18.8 + 0.5
(n = 30) (n = 30) (n = 30)


4.0 12.0 19.0
EUROPEAN NURSE (4-5) (11-12) (19-20)
BEES (H2) 4.4 + 0.5 11.6 + 0.5 19.1 + 0.2
(n = 29) (n = 29) (n = 29)


EUROPEAN COMB CELLS

4.0 12.0 19.0
AFRICANIZED NURSE (4-5) (11-12) (18-19)
BEES (A41) 4.2 + 0.4 11.7 + 0.5 18.9 +0.2
(n = 30) (n = 30) (n = 30)



4.0 12.0 19.0
EUROPEAN NURSE (4-5) (11-12) (19)
BEES (IBR877) 4.3 + 0.5 11.7 + 0.5 19.0 + 0
(n = 26) (n = 26) (n = 26)


4.0 12.0 19.0
TOTALS (4-5) (11-12) (18-20)
4.3 + 0.4 11.6 + 0.5 18.9 + 0.3 (n = 115) (n = 115) (n = 115)


aUS = unsealed brood (unsealed larval development period only).
bSB = sealed brood (pre-pupae and pupae).
CTDT = total development time (oviposition to adult emergence).







26

TABLE 2-2--extended.




EUROPEAN EGG GENOTYPE (Y5)


us SB TDT




5.0 12.0 20.0
(4-5) (11-12) (19-20)
4.9 +L 0.3 11.8 + 0.4 19.6 + 0.5
(n =37) (n =37) (n =37)


5.0 12.0 20.0
(4-5) (12) (19-20)
4.9 .0.4 12. 0 +t 0 19.9 + 0.4
(n =22) (n =22) (n =22)




5.0 12.0 20.0
(4-6) (12-13) (19-21)
5.0 +t 0.4 12.0 + 0.2 20.0 + 0.3
(n =19) (n =19) (n =19)


5.0 12.0 20.0
(4-6) (12-13) (19-21)
4.7 +0.8 12.1 .0.4 19.8 + 0.7
(n 7) (n =7) (n =7)


5.0 12.0 20.0
(4-6) (11-13) (19-21)
4.9 + 0.4 11.9 .0.4 19.8 + 0.4
(n =85) (n =85) (n =85)







27

TABLE 2-3. Summary of hypotheses and tests for evaluating development
times; letters represent treatments (see Table 2-1).


Hi: Worker bee development is faster for Africanized genotypes than
for European genotypes.

A x B African~Ized comb cell size; Africanized nurse bees
C x D Africanized comb cell size; European nurse bees
E x F European comb cell size; Africanized nurse bees
G x H European comb cell size; European nurse bees
A x H Africanized comb cell size and nurse bees compared with
European comb cell size and nurse bees
AC x BD Africanized comb cell size; both nurse bee genotypes
combined
EG x FH European comb cell size; both nurse bee genotypes combined
AE x BF Africanized nurse bees; both comb cell sizes combined
CG x DH European nurse bees; both comb cell sizes combined
ACEG x BDFH Both comb cell size and both nurse bee genotype variables
comb ined

H2: Worker bee development is more rapid in Africanized comb cells

than in European comb cells.

A x E Africanized egg genotype; Africanized nurse bees
C x G Africanized egg genotype; European nurse bees
B x F European egg genotype; Africanized nurse bees
D x H European egg genotype; European nurse bees
AC x EG Africanized egg genotype; both nurse bee genotypes
comb ined
BD x FH European egg genotype; both nurse bee genotypes combined

H3: Worker bee development is more rapid with Africanized nurse bees

than with European nurse bees.

A x C Africanized egg genotype; Africanized comb cell size
E x G Africanized egg genotype; European comb cell size
B x D European egg genotype; Africanized comb cell size
F x H European egg genotype; European comb cell size
AE x CG Africanized egg genotype; both comb cell sizes combined
BF x DH European egg genotype; both comb cell sizes combined







28

TABLE 2-3--continued.


H4: Worker bee development is more rapid with Africanized comb cells
and Africanized nurse bees than with European comb cells and
European nurse bees.

A x G Africanized egg genotype
B x H European egg genotype







29
TABLE 2-4. Unsealed brood development times. Hypotheses were tested
using Kolmogorov-Smirnov one-tailed test, chi-square
distribution, df = 2, alpha = 0.05 (Siegel 1956).


Hi: Worker bee development is faster for Africanized genotypes than for
European genotypes.

A X B ***a
CXD **
E X F**
G XH NS
AXH NS
AC X BD EG X FH
AE X BF CG X OH
ACEG X BDFH
H2: Worker bee development is more rapid in Africanized comb cells than
in European comb cells.

AXE NS
CXG NS
BXF NS
D XH NS
AC X EG NS
BD X FH NS

H3: Worker bee development is more rapid with Africanized nurse bees
than with European nurse bees.

AXC NS
EXG NS
BXD NS
F XH NS
AE X CG NS
BF X DH NS

H4: Worker bee development is more rapid with Africanized comb cells
and Africanized nurse bees than with European comb cells and
European nurse bees.

AXG NS
BXH NS



a ** =p<.
b = P bAnalysis may be NS because test used is conservative for small sample sizes using chi-square distribution.







30

TABLE 2-5. Sealed brood development times. Hypotheses were tested
using Kolmogorov-Smirnov one-tailed test, chi-square
distribution, df = 2, alpha = 0.05 (Siegel 1956).


Hi: Worker bee development is faster for Africanized genotypes than for
European genotypes.

AXB NS
CXD *a
EXF NS
GXH NS
A XH NS
AC X BD *
EG X FH *
AE X BF *
CG X DH **
ACEG X BDF **

H2: Worker bee development is more rapid in Africanized comb cells than
in European comb cells.

AXE NS
C XG NS
B XF NS
DXH NS
AC X EG NS
BD X FH NS

H3: Worker bee development is more rapid with Africanized nurse bees
than with European nurse bees

AXC NS
E XG NS
BXD NS
FXH NS
AE X CG NS
BF X DH NS

H4: Worker bee development is more rapid with Africanized comb cells
and Africanized nurse bees than with European comb cells and
European nurse bees.

AXG NS
BXH NS



a = P<0.05
** = P<0.01.







31

TABLE 2-6. Total worker bee development times. Hypotheses were tested
using Kolmogorov-Smirnov one-tailed test, chi-square
distribution, df = 2, alpha = 0.05 (Siegel 1956).


HI: Worker bee development is faster for Africanized genotypes than for
European genotypes.

A X B ***a
C X D E X F G X H
AXH **
AC X BD
EG X FH AE X BF CG X DH
ACEG X BDFH

H2: Worker bee development is more rapid in Africanized comb cells than
In European comb cells.

AXE NS
C XG NS
BXF NS
D XH NS
AC X EG NS
BD X FH NS

H3: Worker bee development is more rapid with Africanized nurse bees
than with European nurse bees

A XC NS
EXG NS
BXD NS
F XH NS
AE X CG NS
BF X DH NS

H4: Worker bee development is more rapid with Africanized comb cells
and Africanized nurse bees than with European comb cells and
European nurse bees.

AXG NS
BXH NS



a ** =p<
= P<0.O01.







32

TABLE 2-7. Comb cell size for worker development time experiment: comb
measurements = mm for 10 consecutive, horizontal cells, mean
+ SD, (sample size).


COMB CELL SIZE


NURSE BEE AFRICANIZED EGG EUROPEAN EGG
COLONY GENOTYPE GENOTYPE


AFRICANIZED COMB CELL SIZEa

AFRICANIZED NURSE 47.5 + 0.58 49.8 + 0.50 45.8 + 0.50
BEES (A55) (4) (4) (4)

EUROPEAN NURSE 48.2 + 0.96 48.5 + 0.58 48.5 + 0.58
BEES (H2) (4) (4) (4)

EUROPEAN COMB CELL SIZEb

AFRICANIZED NURSE 54.0 + 0.0 54.0 +0.0 54.0 + 0.0
BEES (A41) (3) (3) (3)

EUROPEAN NURSE 53.3 + 0.58 53.3 + 0.58 53.7 + 0.58
BEES (IBR877) (3) (3) (3)



aNatural comb built without foundation. bBuilt from foundation.







33

TABLE 2-8. Comparison of differences in development times (in days)
for Africanized and European honey bees for different
developmental stages.


DEVELOPMENTAL STAGES


EGG HRS UNSEALED SEALED UNSEALED TOTAL
(DAYS)a BROOD BROODc & SEALED DEVELOPMENT


AFRICANIZED EGG 69.6 4.3 11.6 15.9 18.80
GENOTYPE (A26) (2.90)


EUROPEAN EGG 73.6 4.9 11.9 16.8 19.87
GENOTYPE (Y5) (3.07)


% DIFFERENCEd 5.7 14.0 2.6 5.7 5.7



aFrom Harbo, Bolten, Rinderer and Collins (1981); data used are their Africanized #3 = A26 and their European #5 = Y5. bUnsealed larval period only. cPre-pupae and pupae.
d% Difference = [(Y5)-(A26)/(A26)] x 100.







34

TABLE 2-9. Mortality during different developmental stages.


AFRICANIZED EGG GENOTYPE (A26) EUROPEAN EGG GENOTYPE (Y5)


Ela E2b Llc L2d SBe Nf El E2 LI L2 SB N


AFRICANIZED COMB CELL SIZE
AFRICANIZED NURSE BEES

A55 1 5 0 3 1 40 1 0 0 2 0 40

EUROPEAN
NURSE BEES

H2 1 0 3 7 0 40 1 0 0 13 0 36

EUROPEAN COMB CELL SIZE

AFRICANIZED NURSE BEES

A41 0 0 0 0 0 30 8 2 0 0 0 29

EUROPEAN
NURSE BEES

IBR877 0 0 2 2 0 30 13 6 1 0 0 27



aMortality during first 24 hours in test colony (acceptance). bMortality between 24-72 hours (before hatching). CMortality between 72-96 hours (at time of hatching). dMortality during older larval stages, before sealing. eMortality during the pupal stage. N = total eggs monitored.















CHAPTER III
INTERACTION OF MATERNAL GENOTYPE, EGG GENOTYPE AND COMB CELL SIZE ON HONEY BEE WORKER SIZE AND SIZE VARIATION


Introduction

In the evolution of eusociality in bees (Apoidea), there is a

considerable decrease in size variation of the workers within a colony. Worker size variation within a colony of primitively eusocial sweat bees (Halictidae) or bumble bees (Apidae) is much greater than the size variation of workers within colonies of highly eusocial stingless bees (Meliponinae: MeliDona and Trigona) or honey bees (Apinae: AjU) (Brian 1952; Kerr and Hebling 1964; Medler 1965; Michener 1974). For example, the coefficient of variation (CV) for worker weights in a bumble bee colony may be as high as 31-37% (calculated from Brian 1952 for Bombus agrorum, Table B-i) whereas the CV for worker weights within a honey bee (A" mellifera) colony is only 4-7% (Table B-2).

An effect of the reduction of size variation is that the mechanism for the division of labor of workers within a colony shifts from being size dependent to primarily age dependent (Michener 1974). In the primitively eusocial bumble bees, division of labor is size related (Brian 1952); large workers may be twice the size (linear measurements) of small workers within the same colony (Medler 1965). In highly

eusocial stingless bees and honey bees, division of labor is primarily age dependent (Free 1965; Gary 1975; Kerr and Hebling 1964; Lindauer 1953; Seeley 1982). In honey bees, the workers proceed through a series

35







36
of age-related tasks. However, the sequence and duration of the different stages are flexible and depend on the needs of the colony.

An advantage of worker size variation within bumble bee colonies

may be efficient utilization of diverse nectar and pollen resources that may be size dependent. Different sized workers within a colony specialize on those resources that they can most efficiently exploit (Heinrich 1979a). However,. highly eusocial bees are not at a disadvantage with respect to resource utilization because they have evolved complex communication systems that allow foragers to monitor changing nectar conditions and to recruit workers from the colony to a particular resource. Therefore, both the species characterized by workers of highly variable sizes and those species characterized by uniformly-sized workers have evolved behaviors that enhance the efficiency of nectar and pollen exploitation.

The difference In intra-colony worker size variation between primitively eusocial and highly eusocial species of bees is so significant that Kerr and Hebling postulated that "some controlling mechanism leads to reduced variances among mature workers [Meliponinae and APIS], which are therefore of relatively uniform size" (1964,. p. 267). Waddington (1981) hypothesized that the evolution and maintenance of the complex communication systems in AgLj, Trigona and Meipn depend upon uniformity of worker bee size within a colony. Differences in bee size may result in miscommunication because resource profitabilityt" may be size dependent. For example, a high quality resource for a small bee may not be a high quality resource for a larger bee. However, a regulatory mechanism for reduced size variation has not been Identified.







37

Honey bee size and size variation is a result of both genetic and environmental factors. Research has focused primarily on the extrinsic factors that affect bee size (e.g., comb cell *size, nutrition and temperature). Honey bee worker sizes and honey bee comb cell sizes have been shown to be inter-related: because of the manner by which comb cells are constructed (Darwin 1859/1958), worker body size affects the diameter of cells they construct (Baudoux 1933; Glushkov 1958)y and worker bee size is correlated to the size of cells in which they are reared (Baudoux 1933; Buchner 1955; Glushkov 1958; Grout 1937; Michallov 1927-28 cited in Alpatov 1929; Tuenin 1927).

This Interaction between comb cell size and egg genotype may at first appear to provide a mechanism for both regulating bee size and reducing size variation among bees within a colony. However.. the comb cell itself can become a source of variation in bee size. Although comb cell size appears quite uniform, especially when first constructed, the cells become variable in size as the number of generations reared in them increases, because pupal cocoons adhere to the cell walls, reducing cell diameter (Abdellatif 1965; Alpatov 1929; Buchner 1955; Grout 1937). For example, there Is a 25% reduction in cell volume between cells from new and old combs (Table B-3). Comb cell volume has a greater variance than cell diameter and is not correlated with diameter (Table B-3).

In addition to comb cell size, there are other extrinsic factors that affect development and resultant bee size, e.g., quantity and quality of larval food, and temperature and humidity at which the larvae and pupae are reared (Buchner 1955; Fyg 1959; Jay 1963; Kulzhinskaya 1956; Michallov 1927-28 cited in Alpatov 1929). These same factors not only affect absolute size but are sources of size variation.







38

The importance of the genetic component to bee size can be Inferred from the fact that different geographic populations of honey bees differ with respect to worker bee size (Alpatov 1929; Ruttner 1968, 1975, 1976a, 1976b; Wafa, Rashad and Mazeed 1965). Because honey bee queens mate with many different drones (Adams, Rothmans Kerr and Paulino 1977; Peer 1956; Roberts 1944; Taber 1954; Taber and Wendel 1958), the genetic component becomes an additional factor affecting size variation.

Africanized honey bees in Venezuela (descendents of A. a.

ss.1j~J..lat.) were smaller and had a smaller comb cell diameter (mean 4.8 mm between opposite sides of the hexagonal cells in the comb) compared with European bees in Venezuela (mean 5.4 mm) (Tables B-3 and B-4; Rinderer, Tucker and Collins 1982). An opportunity therefore, existed to experimentally evaluate the interaction of both genotype and comb cell size on resultant worker bee size and size variation by studying both the Africanized and European honey bee populations under identical experimental conditions. The results from this study provide information not only on the proximal question involving the factors affecting bee size but also provide a mechanism by which size variation may be reduced within a honey bee colony.




Nine genotypes were evaluated: three Africanized, three European, and three F1 reciprocal hybrids. The Africanized genotypes (A26# A57, and B39) were established from queens removed from feral colonies located in an area in eastern Venezuela with no known European honey bees. They were Identifiled as Afrlcanized honey bees by their comb cell sizes which were significantly smaller than European comb cell sizes (Tables B-3 and B-4; Michener 1972, 1975; Rinderer, Tucker and Collins







39

1982). The European genotypes (YD28 and WEI) were imported into Venezuela from the U.S. Department of Agriculture Bee Breeding and Stock Center Laboratory in Baton Rouge, Louisiana, U.S.A., and from a commercial queen producer from southeastern U.S.A., respectively. Queen YD28 was artificially inseminated with the spermatozoa from one drone; queen WEI was naturally mated. The third European genotype (SDY1) was a daughter from line YK produced by another commercial queen producer from southeastern U.S.A. and artificially inseminated in Venezuela with a single drone from the same commercial line.

Two reciprocal hybrid lines were established from artificially

reared queens CLaidlaw 1979) that were instrumentally inseminated with spermatozoa from single drones: Africanized queen x European drone (SDA12) and European queen x Africanized drone (SOYlO and SDY11). The Africanized queen and drone source was A26. The European queen and drone source was line YK. The hybrid lines were therefore genetically similar, but were the reciprocal of each other with respect to their queen and drone sources.

Queens were produced by the standard method of transferring young larvae from the desired queen line into artificial queen cells which were then Introduced into cell-producing colonies (Laidlaw 1979). Mature queen cells were put into an incubator (35 t 10C) 72 hours prior to adult emergence. Newly emerged virgins were marked for individual identification and then put Into individual cages and maintained in a strong, queenless colony for approximately one week until they were artificially inseminated.

Drones for Instrumental inseminations were produced by caging drone comb containing sealed drone pupae from the desired drone source lines.







40

As drones emerged, they were placed into special holding cages and maintained in a colony so that worker bees could feed them until they matured. This manipulation insured that the drones used for inseminations were from the desired queen lines.

To collect eggs for the bee size experiments, queens from the nine genotypes were confined for five hours in their own colonies to a section of Africanized comb (mean cell size = 4.8 mm), using 8 x 8 cm push-in cages. These cages were made from 3 mm mesh hardware cloth and had queen excluder material soldered to the top to enable worker bees to pass through to tend the queen (Harbo, Bolten, Rinderer and Collins 1981). After five hours, the queens were removed from the combs. The 8 x 8 cm sections of comb with eggs from each queen were cut out and fitted Into special frames. The nine sections were then placed In a strong Africanized colony (Africanized nurse bees and Africanized comb cell size) for development. The following day, eggs were collected In European combs (mean cell size = 5.4 mm) using the same procedure with the same nine queens except that the nine sections were put Into a European colony (European nurse bees and European comb cell size) for development. Having all nine egg sources for each comb cell size treatment (Africanized or European) develop in the same colony controlled for additional variables affecting development and bee size: temperature and humidity, nurse bee genotypes and colony size (see Chapter ID).

Fresh pupal weights were compared for each of the nine genotypes

reared in both Africanized and European comb cell sizes. Pupal weights were measured on the 16th day after oviposition. This age corresponds to the period during pupal development of least weight change (Melampy







41

and Willis 1939). This was confirmed for fresh pupal weights by weighing a sample of pupae every 24 hours from day 11.5 post oviposition to 17.5 days post oviposition (Table B-5). Although Africanized bees develop one day faster than European bees (Chapter ID), pupal weights can be compared because there Is no significant difference in weights between adjacent days during this period of pupal development (Table B5).

Pupal weights were used instead of adult weights in order to reduce variation resulting from differences in food engorgement and/or feces accumulation. Pupae were carefully removed from their comb cells by first removing the cappings and then spreading the cell walls with a forceps in order that the pupae could easily be removed without rupturing. Weights (to 1.0 mg) were recorded using Mettler Type H4 and H6 balances. Comb cell diameters were determined by measuring ten adjacent cells; three sets of measurements were made from each comb.




Table 3-1 presents the experimental design matrix. The interaction of egg genotype and comb cell size on worker bee pupal weights for each of the nine genotypes is summarized In Table 3-2. Table 3-3 presents the results of the statistical analyses. When Africanized and European genotypes are reared simultaneously in the same colony (same comb cell size, nurse bee genotype, temperature and humidity.. and colony size), the weights of the worker bees produced are different. African ized bee pupae (111.1 +t 7.6 mg) that developed in African ized comb cells were smaller than European bee pupae (123.3 + 6.3 mg) that also developed In Africanized comb cells (ACE x MOQ, P<0.001). When worker bees of European genotypes are reared in AfricanIzed comb cells, the cells are







42

sealed with strongly convex cappings similar to the way cells containing drones are sealed In order to accomodate their larger size. Africanized bee pupae (123.8 + 6.2 mg) that developed in European comb cells were smaller than European bee pupae (139.5 + 5,4 mg) that also developed in European comb cells (BOF x NPR, P<0.001). For each of the nine genotypes investigated, worker bee pupae that developed In Africanized comb cells were smaller than pupae that developed In European comb cells, P<0.001. There is a 43% increase In comb cell volume between Africanized and European combs (Table B-3), but the Africanized and European genotypes only increased in pupal weight by 11.4% and 13.1%0 respectively (Table 3-2). These results show that both genotype and comb cell size affect worker bee size.

Table 3-4 presents the results for the pupal weights of the

reciprocal F1 hybrids and their respective maternal lines. Data from only European comb cells were used in order to observe genotype effects without the constraint of the small Africanized comb cells on European genotypes. Table 3-5 summarizes the results of the statistical analyses. The pupal weights of the hybrids from this reciprocal F1 cross were significantly different from each other (H x J; H x L; P<0.001), but were the same as their respective maternal line (B x H; J x R; L x R).




Reduction of Bee Size Variation

Bee size is a result of not only the Interaction of egg genotype and comb cell size but also the maternal genotype. This can be seen by evaluating the reciprocal hybrid crosses. The genotype component for bee size Is not a result of "simple" inheritance because pupal weights







43

of genetically similar, reciprocal F1 hybrids are not the same. Because reciprocal F, hybrids are phenotypically different from each other.. but phenotypically similar to their maternal lines.. maternal genotype must Interact with cell size and egg genotype to determine pupal weight. This is the first character in honey bees that has been shown to be influenced by maternal inheritance. Other genetic mechanisms cannot explain these results. The mechanism for maternal inheritance in worker size may be through egg size, which has been shown to be inherited (Roberts and Taber 1965; Taber and Roberts 1963).

Alles (1961) and Mellnichenko (1962) suggested that differences between nurse bee genotypes might affect size of developing larvae. However, McGregor (1938) found that bee size was not affected by nurse bee genotype. In the experiments presented in this chapter, pupae from each of the genotypes were reared simultaneously in the same colony for each comb size treatment. Therefore differences between the pupal weights of Africanized and European genotypes cannot be attributed to either nurse bee differences, cell size, or temperature but must be a result of both egg and maternal genotype differences.

The importance of maternal inheritance on bee size is that it reduces worker bee size variation within a colony. If maternal inheritance were not operating* worker bees of different sizes would be produced within a colony because of cell size differences and genotype differences. The effectiveness of maternal inheritance for reducing bee size variation can be demonstrated by comparing the degree of variation for the two parameters of bee size (comb cell volume and genotype) with the degree of worker bee size variation. Abdellatif (1965) showed that







44

when comb cell size variation increased 300%, bee size variation increased only 50%.

The genetic variation of worker bees within a colony Is great because queens mate on the average with as many as 17 drones (Adams, Rothman, Kerr and Paulimo 1977). There is some degree of mixing of spermatozoa in the spermatheca resulting In spermatozoa from at least 5 to 6 drones being used during one time interval (Page and Metcalf 1982). Evidence that maternal inheritance reduces size variation in genetically diverse worker offspring can be demonstrated by evaluating the size variation of offspring from single-drone and multiple-drone inseminated queens. The progeny of queens that were Inseminated by spermatozoa from single drones (SDA12, SDY1O, SDY11, SDY1 and YD28) were expected to be less variable than multiply-inseminated queens (A26, A57, B39 and WE1) because all eggs from the former queens would have been fertilized by a genetically identical male gamete. (Drones are haploid; all spermatozoa are produced by mitosis and are therefore genetically Identical.) Evaluating the coefficient of variation (CV) for each treatment of genotype and comb cell size, there Is no difference between the variation of progeny from single-drone inseminations versus those from multiple inseminations, as shown in Table 3-6 (Mann-Whitney U test, onetailed, alpha = 0.05).

Additional evidence of maternal Inheritance reducing size variation in genetically heterogeneous offspring comes from analyzing the results of the reciprocal F, cross. Because of the Influence of maternal inheritance, subspecific differences In size between Africanized and European populations were not reflected In Increased size variation of the hybrids compared with the parental types (Table 3-4).







45

Further evidence of the effectiveness of maternal inheritance reducing size variation can be seen by comparing the size variation within a colony to the size variation within a population. Alpatov (1929) found that within honey bee colonies# worker size variation (e.g., for tongue length) was less than the variation for the local population of a managed apiary. For seven different apiaries in Russia$

each apiary had an average 22.3% (range 5-423) increased variation over the mean colony variation within the apiary. Although the genetic homogeneity of the apiaries is artificially high as a result of management practices of the beekeepers compared with the variation of natural populations of animals (Alpatov 1929), within-colony variation was still noticeably reduced.

Evolution of complex communication systems in highly eusocial species may be responsible for selection for reduced size variation (Waddington 1981; Waddington., Herbst and Roubik 1986). Foragers within honey bee colonies have the ability to communicate Information to nest mates about the directions distance and "profitability" of new resources (von Frisch 1967) which may be interpreted correctly only if worker bees within the colony are the same size (Waddington 1981). Profitability of the resource may be size-dependent as Waddington (1981) suggested. That is, a high quality resource for a small bee may not be a high quality resource for a larger bee.

In addition to the profitability component, correct interpretation of the distance component of the honey bee waggle dance (von Frisch 1967; Wenner 1962) may also be size-dependent. Different distance dialects occur not only between subspecies (Boch 1957; Gould 1982) but also between colonies (Esch 1978 cited In Gould 1982). There is greater







46

variation in individual dialects in colonies that are genetically heterogeneous compared with colonies that are genetically homogeneous (Gould 1982). Variation in bee size within a colony may accentuate differences in distance dialects and Increase the possibility of miscommunication. Therefore, worker size variation within a colony of honey bees needs to be reduced in order for a communication system that recruits foragers to a particular floral resource to function correctly and efficiently with respect to either the profitability (Waddington 1981; Waddington.. Herbst and Roubik 1986) or distance component. Maternal effects operate to reduce bee size variation within a colony of honey beesp thereby allowing their communication system to function

ef f ectivel y.

Africanized and European Honey Bee Size Difference

Several hypotheses have been suggested to explain the smaller

worker bee size of the Africanized population. One advantage suggested for smaller size Is more rapid development times# permitting more rapid colony growth resulting In increased reproductive swarming (Fletcher 1977a; Fletcher and Tribe 1977a; Tribe and Fletcher 1977). However# cell size and bee size do not affect development times ands in addition, worker development times do not affect colony growth rates (Chapter ID.

Fletcher and Tribe (1977a) and Tribe and Fletcher (1977) suggested that smaller bee size would permit greater numbers of worker bees to be reared on the same amount of food compared with larger bees. Advantages of increased worker numbers Include frequency of reproductive swarming, colony defense and foraging success (Wilson 1971). Thus, smaller$. individual bee size maximizes the use of the limited food that







47

characterizes the unreliable nectar availability in Africa (Tribe and Fletcher 1977). Smaller bee size increases the resource utilization efficiency of Africanized honey bees and may be a factor In the success and high reproductive rates of Africanized honey bees compared with European honey bees in tropical areas of South America (see Chapter VIII).

I suggest two other hypotheses to explain the advantages of smaller size in the Africanized population. First, smaller size Is more efficient with respect to dissipating heat loads in tropical habitats (see also Heinrich 1979b). Fletcher (1978) reports that foraging may stop during the hottest part of the day, which would avoid the disadvantages of smaller size with respect to gaining a heat load. The sizes of two other subspecies of honey bees in Africa support this hypothesis. One of the smallest subspecies in Africa, A. aw. litoreas is found In a very hot and dry area along the coast of Kenya and Tanzania. One of the largest subspecies, &. m_. monticola, is found at higher elevations and colder temperatures on Mount Kenya.

Secondly, the advantage of smaller bee size may actually lie with the advantages of smaller cell size. For a given nest cavity volume, a

larger number of worker bees can be produced if cell sizes are smaller. There Is approximately a 25% increase in the number cells for a given comb area with smaller Africanized comb cells compared with larger European comb cells. Considering the advantages of Increased worker numbers In a colony (Wilson 1971), the increase In worker numbers as a result of smaller cell size may be important, particularly if nest cavity volumes are limited.







48

Because maternal inheritance affects bee size, methods that use components of size to identify Africanized bees, e.g., morphometric analysis, may be invalid. Offspring from the cross of a European queen x Africanized drone (SDY1O and SDY11) are the same weight as offspring from European queen x European drone (SDY1) (Table 3-4). More importantly, the offspring from the cross of a European queen x Africanized drone are significantly different from offspring from Africanized queen x Africanized drone and Africanized queen x European drone matings. The European queen x Africanized drone mating represents the most probable scenario for initial hybridization in North America (see Chapter VII). That is, a virgin queen from a managed or a feral European colony mates with Africanized drones and produces offspring with a 50% Africanized genome. Analyzing the offspring using size as a component for identification may result in a false negative identification of Africanized bees. The extent of the problem would depend upon the degree to which particular linear measurements are either affected by maternal inheritance and/or are correlated with bee weight. As Daly, Hoelmer, Norman and Allen (1982) point out, there is a "difficulty in using phenotype characters to identify genetically different, but closely related populations" (p. 593).







49

TABLE 3-1. Effect of comb cell size and egg genotype
on bee pupal weights. Experimental design
matrix (code letters A-R used in tables of
statistical analyses).


COMB CELL SIZE


EGG GENOTYPES AFRICANIZED EUROPEAN


AFRICANIZED QUEEN X AFRICANIZED DRONE

A26a A B
A57a C D
B39a E F

AFRICANIZED QUEEN X
EUROPEAN DRONE

SDA12b G H

EUROPEAN QUEEN X
AFRICANIZED DRONE

SDY1Ob I
SDY11b K L

EUROPEAN QUEEN X
EUROPEAN DRONE

YD28b M N
WEla 0 P
SDY1b Q R

aNatural matings, multiple inseminations. bSingle drone insemination.







50

TABLE 3-2. Effect of comb cell size and egg genotype on bee pupal
weights (mg). Means + SD, (sample size).


COMB CELL SIZE


EGG GENOTYPES AFRICANIZEDa EUROPEANb % INCREASE


AFRICANIZED QUEEN X
AFRICANIZED DRONE

A26c 105.4 + 4.6 121.7 + 5.5 15.5
(60) (80)
A57c 117.2 + 6.8 128.6 + 4.4 9.7
(30) (40)
B39c 116.5 + 3.6 123.1 + 6.3 5.7
(30) (40)
COMBINED 111.1 + 7.6 123.8 4+ 6.2 11.4
(120) (160)
AFRICANIZED QUEEN X EUROPEAN DRONE

SDA12d 112.9 + 4.4 122.4 + 4.4 8.4
(30) (40)
EUROPEAN QUEEN X AFRICANIZED DRONE

SDYlOd 114.6 + 3.7 133.7 + 3.3 16.7
(10) (23)
SDY11 115.5 + 3.2 138.9 + 4.9 20.2
(30) (30)
COMBINED 115.3 + 3.3 136.7 + 5.0 18.6
(40) (53)
EUROPEAN QUEEN X
EUROPEAN DRONE

YD28d 126.8 + 4.2 138.7 + 3.5 9.4
(30) (30)
WE1c 125.1 + 4.3 143.3 + 2.9 14.5
(30) (30)
SDYld 115.4 + 4.2 135.0 + 6.9 17.0
(20) (19)
COMBINED 123.3 + 6.3 139.5 + 5.4 13.1
(80) (79)

aWidth between opposite sides of the hexagonal cell is 4.8 mm. bWidth between opposite sides of the hexagonal cell is 5.4 mm. cNatural matings, multiple inseminations. dSingle drone insemination.









TABLE 3-3. Worker bee size: hypotheses and analyses (MannWhitney U test, one-tailed, alpha = 0.05).
Letters refer to experimental treatments, see
Table 3-1.


Hl: Africanized bee pupae are smaller than European bee
pupae independent of comb cell size.

ACE x MOQ ***a
BDF x NPR ACE x NPR
BDF x MOQ NS


H2: For a given egg genotype, pupae that develop in
Africanized comb cell size are smaller than pupae
that develop in European comb cell size.

A x B C x D E x F
G x H I x J K x L
M x N 0 x P
Q x R
ACE x BDF MOQ x NPR



a*** p






52

TABLE 3-4. Reciprocal F cross. Pupal weights (mg), mean + SD,
(sample size). Data are from European comb cell size only.

AFRa QUEEN AFR QUEEN EURa QUEEN EUR QUEEN
X X X X
AFR DRONEb EUR DRONEc AFR DRONEc EUR DRONEc

(A26) (SDA12) (SDY10) (SDY11) (SDY1)


121.7 + 5.5 122.4 + 4.4 133.7 + 3.3 138.9 + 4.9 135.0 + 6.9
(80) (40) (23) (30) (19)

B H J L R



aAFR = Africanized; EUR = European. bNaturally mated.
cSingle-drone, artificial insemination.







53

TABLE 3-5. Maternal effect: hypotheses and analyses (MannWhitney U test, one-tailed, alpha = 0.05).
Letters refer to experimental treatments.. see
Table 3-4. The analyses of the following
hypotheses (a postiori) demonstrate that the pupal weights of hybrids from a reciprocal F, cross are different from each other (H4) but are the same as
their respective queen mothers (H2 and H6).


Hl: B < R ***a

H2; B < H NS

H3: B < J
B < L

H4: H < J
H < L

HS: H < R

H6: J < R NS
L < R NS



a*** p<0.001.







54
TABLE 3-6. Coefficients of variation for pupal weights from
artificial, single drone inseminations and
natural, multiple matings.


COMB CELL SIZE


EGG GENOTYPEa AFRICANIZED EUROPEAN


SINGLE DRONE INSEMINATIONS

SDA12 3.9 3.6
SDY1O 3.2 2.5
SDY11 2.8 3.5
SDYl 3.6 5.1
YD28 3.3 2.5

MULTIPLE INSEMINATIONS

A26 4.4 4.5
A57 5.8 3.4
B39 3.1 5.1
WEl 3.4 2.0


ANALYSESb NS NS

aSee Table 3-1 for explanation of genotypes. bMann-Whitney U test, one-tailed, alpha = 0.05.















CHAPTER IV
QUEEN DEVELOPMENT AND MATURATION


Introduction

African honey bees, mellifera scutellata (formerly classified as adansoni; Ruttner 1976a, 1976b, 1981), were introduced into southeastern Brazil in 1956 (Kerr 1967; Michener 1975; Woyke 1969). The

following year, swarms escaped and hybridized with the established European honey bees (primarily &. a_. lioustica and mellifera) that had been introduced by 1845 (Gerstaker cited in Pellet 1938; Woyke 1969). The descendents from this hybridization are known as Africanized honey bees (Goncalves 1982).

Africanized honey bees in South America have a very high annual reproductive rate compared with European honey bees in temperate regions. Based on demographic data collected in French Guiana, the net reproductive rate for Africanized bees is estimated to be 16 colonies per colony per year (Otis 1980, 1982a). In comparison, the annual rate determined for European honey bees in North America was 0.92-0.96 (Seeley 1978) or 3-3.6 when afterswarms are considered (Winston 1980a; Winston, Taylor and Otis 1983). This dramatic difference in reproduction between these two honey bee populations may be a result of length of time throughout the year that resources are available in the tropics compared with temperate regions (see Chapter VIII) and/or




55







56

demographic characteristics of Africanized honey bees that account for high reproductive rates.

The reproductive rate of Africanized honey bees results in a swarmto-swarm interval of approximately 90 days (Winston 1979b). During that period, a virgin queen emerges, develops pheromones necessary to attract drones, and mates; ovarian follicles mature; oviposition Is initiated; and the colony population growth period begins prior to the next swarming. One expected demographic feature for a population with a high reproductive rate would be a short queen maturation interval (Fletcher 1977a). For the Africanized queens in French Guiana, the maturation interval from pupal eclosion to initiation of oviposition was 9.7 days (Otis 1980), over 10% of their swarm-to-swarm Interval (calculated from Winston 1979b). Fletcher and Tribe (1977b) report that in the parental African population, oviposition begins on the 8th to 9th day after queen emergence. European queens begin ovipositing between the 6th and 17th day after emergence (Laidlaw and Eckert 1962; Oertel 1940; Root 1947). Otis (1980) calculated that the mean interval from pupal eclosion to oviposition for European queens (10.7 days) was not significantly different from that of Africanized queens (9.7 days). However, comparisons between reported values for both Africanized and European honey bees are Inappropriate because the data were collected under very different experimental conditions. Therefore, this study was undertaken to determine If the queen maturation interval for Africanized honey bees is significantly different than that for European honey bees under identical conditions. Three aspects of queen maturation were evaluated: 1) larval, pupal and total development time from egg to adult emergence;







57

2) post-emergence development of queen attractiveness to drones; and 3) time from adult emergence to initiation of oviposition.

In the studies reported here, queen development and maturation were evaluated under controlled conditions. Total development time is defined as the time from oviposition to adult emergence. These experimental conditions avoid the problems of previous studies that evaluated queen development and maturation in colonies that were swarming (e.g., Otis 1980). Under natural swarming conditions, queens are very often confined within their cells by worker bees and prevented from emerging for 1-10 days after pupal eclosion (Otis 1980). Confinement makes calculations of development times difficult, and, because maturation proceeds during confinement, maturation time calculated from emergence to beginning of oviposition would be under estimated.

Methods

Queen Development Times

The Africanized egg source (A26) and the Africanized cell-producing colonies (A37 and A43) were established from queens removed from feral colonies found In an area of eastern Venezuela where there were no known European honey bees. They were identified as Africanized honey bees by their behavior and characteristic comb cell size (4.5-5.0 mm wide between opposite sides of the hexagon, see Chapter III). The European egg source (Y5) and the European cell-producing colonies (19, 27 28, F, H and H1) came from European queens commercially produced in the southeastern U.S.A. and shipped to Venezuela. European colony IBR was a stock supplied by the U.S. Department of Agriculture Bee Breeding and

Stock Center Laboratory, Baton Rouge, Louisiana, USA.







58

Eggs of known ages were collected from the Africanized (A26) and European (Y5) egg sources by the standard commercial queen-producing technique of caging the queen on an empty comb within a colony (Harp 1973; Laidlaw 1979; see also Chapter II). After 6 hours, the combs with the egg samples were moved to strong incubator-colonies for the eggs to develop and larvae to hatch and be fed. Very young larvae, 12-18 hours old, were transferred (grafted) into beeswax queen-cell cups primed with royal jelly and then introduced into queen-cell-producing colonies (=nurse bee colonies) (Laidlaw 1979). All cell-producing colonies had large worker bee populations and were intentionally crowded into two standard Langstroth hive bodies. Queens In the cell-producing colonies were removed 48 hours before introducing the grafted cells. All young, unsealed brood was also removed 2-4 hours before introducing the grafted cells. Twenty grafted AfrIcanized and twenty grafted European cells were introduced into each cell-producing colony. There were twenty cell cups to a frame, ten on the top bar and ten on the middle bar. Both

Africanized and European larvae were grafted into the same frame, five each on the top bar and five each on the middle bar. All cell cups were equally spaced about 8 mm apart, centered on the bars.

In one experimental trial, the effect of nurse bee genotypes on

queen development was evaluated by comparing queen development times for both Africanized and European egg genotypes in both Africanized and European cell-producing colonies. In another trial, development times for Africanized queens In Africanized and European cell-producing colonies were compared. In both trials, the Africanized cell-producing colonies had comb cell sizes characteristic of Africanized honey bees (4.8 mm wide; Chapter III).







59

The queen-cell-producing colonies were Inspected only after the queen cells had been sealed In order to avoid disturbance which could affect development times. Once the cells are sealed, cell-producing colonies only maintain the appropriate temperature for the pupae to

develop normally. On the sixth day after grafting, each sealed cell was protected by placing a 3 mm wire mesh tube around it to avoid any problems associated with queens being confined to their cells by worker bees. In addition, this also prevented any emerged virgin queens from destroying sealed cells that had not yet emerged. Beginning 24 hours before any expected queen emergence, the cell-producing colonies were

inspected daily at 0630,- 1200 and 1730 hours to record queen emergence. In two trials, cell-producing colonies were inspected daily at 0630, 1200 and 1730 hours, beginning 24 hours prior to estimated sealing time, in order to determine unsealed development times. Development of Attractiveness of VIrain AfrIcanized and-European Honey
Bee Queens to Drones
Two Africanized queen mothers (A26 and A57) were removed from feral colonies in eastern Venezuela. The two European queen mothers (We and

Yk) were shipped to Venezuela from different commercial queen breeders In southeastern USA.

Queens from the four queen mothers CA26, A57, We and Yk) were

produced as described above. Sealed queen cells were removed from the cell-producing colony and placed in an Incubator (35 t 10C) 48 hours prior to emergence. After emergence, the queens were marked for individual Identification and maintained in separate cages in a queen storage colony (Laidlaw 1979).

In order to test for the degree of attractiveness to drones, each queen was tethered in a clean, plastic screen bag. The mesh size was 1







60

x 1 mm, and each bag was approximately 5 x 10 cm. Bags were individually suspended on monofilament line about 6.5 meters above the ground, centered between two poles 20 meters apart. Queens could be rapidly raised and lowered by a pulley system. Queens were put into the mesh bags just prior to testing in order to avoid any pheromone accumulation.

The testing location was In an open field in a drone congregation area (Zmarlicki and Morse 1963)p which was located by walking with a helium-filled weather balloon with mature queens suspended 10-20 meters above the ground. The drone congregation area was Identified when hundreds of drones oriented to the tethered queens. Boundaries appeared to be quite distinct and stable through time. Both Africanized and European drones were probably present, but the identity of each drone responding to specific queens during the experiment was not known because there are no reliable techniques to identify individual Africanized and European honey bees. How drone congregation areas become established is not understood, but these areas are probably where most mating occurs.

Individual queens were tested for drone response on consecutive days, beginning on the day of emergence. Only one queen at a time was tested so that the relative attraction of each queen would not be influenced by other queens being tested simultaneously. Testing lasted for a maximum of 3 minutes for each queen, even If no drone response was observed. Periodically, empty bags (blanks) were tested to insure that drones were responding only to the queens and not orienting to the experimental set-up and responding to the mesh bags. At no time did drones respond to the blanks. A random sequence for testing individual







61

queens was established on each day of the experiment. Each queen was tested more than once on each day and always in a new mesh bags to avoid any pheromone accumulation or contamination. Each testing session was begun by suspending an older queen that had previously been determined to be maximally attractive, in order to insure that a responding drone population was available. This process was also repeated if the testing session was interrupted by rain, extreme cloudiness., or high winds-conditions that normally reduce drone flight activity. The testing took place between 1400 and 1600 hours.

Drone response was evaluated by assigning one of the following

ranks to the test queen:

Rank 0 = no response

Rank 1 = drones oriented to the test subject but only flew past;

no circling of the test subject

Rank 2 = drones oriented to the test subject and persisted in a

wide circling formation more than 2 m from the subject

Rank 3 = drones oriented to the test subject and formed a loose

comet-like formation down wind more than 0.5 m to the

test subject; formation was volatile# continually

fragmenting and reforming; drones did not land on the

mesh bag

Rank 4 = drones oriented to the test subject and formed a tight

comet-like formation down wind less than 0.5 m from the

test subject; formation was persistent and did not

fragment even as the test subject was lowered; drones

landed on and walked over the mesh bag.







62

These ranking categories were easily discriminated and were not affected by the absolute numbers of drones flying. No estimates of the drone population were made.

Time Pgst-Emercence to the Initiation of Oviposition

Queens were produced as described above from one Africanized egg

source (A26) and one European egg source (We). Twenty-seven Africanized and twenty-five European mature queen cells (two days prior to emergence) were each Introduced into a four-frame queenless mating colony. Any natural queen cells in the mating colonies were destroyed before introducing the experimental queen cells. This insured that the only queen in the mating colony would be the experimental queen. When only one queen cell is present.. worker bees usually do not confine her to her cell and the problem of calculating maturation time Is avoided.

Because Africanized and European queens did not develop at the same rate, the day of queen emergence was determined by the mean time of emergence for a sample of sister queens from the same graft that were left to emerge in an Incubator at 35 + 10C. On the eleventh day after the queens emerged, the colonies were Inspected and the age of the brood was evaluated to determine the age post-emergence when the queens had begun ovipositing. Those colonies in which there were no larvae were inspected three and five days later.

This experiment took place during the dry season. Clear weather prevailed so that mating flights were not affected by weather conditions. Both Africanized and European drones were in the area.







63

Results

Queen Development Times

Table 4-1 presents the experimental matrix for evaluating the

interaction of egg genotype and nurse bee genotype on queen development times for Africanized and European queens. Table 4-2 presents the total development times from oviposition to adult emergence for Africanized queens and European queens in Africanized and European cell-producing colonies. Table 4-3 presents the analyses for the paired comparisons in each cell-producing colony. These paired comparisons avoid any differences between cell-producing colonies because colony size (nurse bee population), brood area temperature, and quantity and quality of larval food are factors that affect queen development (Beetsma 1979; Laidlaw 1979; Johansson and Johansson 1973). Africanized queens develop in 14.5 days post-oviposition compared with 15.0 days for European queens (P<0.001, Kolmogorov-Smirnov one-tailed test, chi-square distribution, df = 2; Siegel 1956). There was no significant effect of the cell-producing colony on queen development times (Kolmogorov-Smirnov two-tailed test, chi-square distribution, df = 2, alpha = 0.05) (Tables 4-4 and 4-5).

Table 4-6 presents the development times for the Africanized queens In Africanized and European cell-producing colonies. There was no difference in Africanized queen development time between Africanized and European cell-producing colonies (Kolmogorov-Smirnov one-tailed test, chi-square distribution, df = 2, alpha = 0.05).

The median unsealed development times from oviposition to sealing for both the Africanized and European queens was 7.5 days (Table 4-7). However, the Africanized and European genotypes were significantly







64

different as a result of the distribution around the median (P
The response of drones to tethered, virgin queens is summarized in Table 4-8. There were no differences between Africanized and European queens with respect to either the earliest age at which a positive drone response (Rank 1) was observed or the earliest age at which a maximum drone response (Rank 4) was observed. Both Africanized and European virgin queens were able to attract drones (Rank 1) on the day they emerged. Africanized virgin queens can maximally attract drones (Rank 4) by the fourth day post-emergence; European virgin queens can elicit a Rank 4 response by the fifth day post-emergence. This difference was not significant (Kolmogorov-Smirnov one-tailed test, chi-square distribution, df = 2, alpha = 0.05). The data in Table 4-8 have been combined for the two Africanized and two European queen lines. However, the queens within a population (Africanized or European) or within a line within a population were not uniform with respect to drone response or the rate of maturation. There were differences between the two Africanized lines and between individuals within the same line for the earliest age for a Rank 4 response. This same variation between lines and within lines existed for the European population. Time Post-Emergence to Initiation of Oviposition

Table 4-9 presents the data for time post-emergence to the

initiation of oviposition for both Africanized and European queens. Africanized queens began oviposition at 8.5 days post-emergence whereas European queens begin at 7.5 days (P






65

Discussion

Queen Development

Development time from oviposition to emergence for Africanized

queens in this study was 14.5 days, which is the same as the development period reported for both Africanized bees in French Guiana (Winston 1979c) and their parental population, A. m. adansonij scutellatata) in South Africa (Anderson, Buys and Johannsmeier no date; Fletcher 1978; Fletcher and Tribe 1977c). The European queens in the present study developed in 15.0 days, which is about one day shorter than expected from previous reports (Jay 1963; Laidlaw 1979). Therefore, the difference in development times for Africanized and European queens was not as great as expected and underscores the importance of making comparisons under the same experimental conditions.

Queens have approximately a 25% shorter development period than

worker bees. Differences in total development times between queens and worker bees are primarily due to a much shorter sealed development stage, i.e., 7.5 days compared with 12 days for European bees and 7.0 days compared with 12 days for Africanized bees. The sealed development stage in worker bees is approximately 60% of the total development time, whereas in queens it is approximately 50%.

European queens took 3.4-5.6% longer than Africanized queens to develop. This difference is similar to the 5.3% difference in development times between European and Africanized worker bees from the same two egg sources (A26 and Y5) (see Chapter II).

There was no effect of nurse bee genotype on queen development times. However, queens were produced more successfully in European colonies. Africanized nurse bees were easily disturbed when the grafted







66

cells were Introduced into the colonies, resulting in poor survival or

acceptance of the grafted larvae (5-50% for Africanized colonies, compared with 35-95% for European colonies). In addition, Africanized colonies were difficult to manage because of excessive stinging that occurred when manipulating the strong colonies that were necessary for proper queen production.

Page and Erickson (1984) found evidence that nurse bee colonies

preferentially raised queens from more closely related larvae. However, in the present study, no evidence for kin recognition was observed. Africanized and European nurse bee colonies reared Africanized and European queens with equal frequency (Table 4-2). Rate of Ma1turation

Attractiveness of queens to drones is a function of the amount of pheromone (9-oxodec-trans-2-enoic acid) produced in the mandibular glands of the queens (Butler 1971; Boch, Shearer and Young 1975). In England, using European genotypes, Butler (1971) tethered virgin queens of various ages 6 meters above the ground in areas where drones were flying. He determined that queens younger than 5 to 6 days old seldom elicited a positive drone response. Maximum positive responses from drones were observed In queens 8 or more days old. Bule' results differ from those presented in this study and may be attributed to either differences In experimental conditions or genetic differences between the queen lines studied rather than to differences due to any tropical or temperate conditions. The response of drones to queens reported in this chapter was evaluated in a drone congregation area which may account for the differences between the studies.







67

In addition, Africanized drones (not present In Butler's study) may have a lower response threshold to queen pheromone and therefore would respond to queens with less pheromone present than would European drones. This hypothesis is suggested by the observation that there are

differences in the sensory receptors on the antennae of Africanized drones compared with European drones (Dietz 1978). Further comparison between African ized and European drones is needed to determine any differences In the threshold of response and whether or not this would give the African ized drones a mating advantage.

Another factor that needs to be considered when using this

behavioral bioassay (drone response) to compare rate of maturation of queens Is that the pheromone is not continually produced but rather Is pulsed In Its production (R. Boch, pers. comm.). This factor may help to explain some of the variation of responses produced by queens within the same line. For examples in a few trials# a queen elicited a decreased response compared with the previous response she had elicited.

In the present study, the time post-emergence to the initiation of oviposition for Africanized queens was 8.5 days and for European queens was 7.5 days. There are no other data available that allow for valid comparisons. For example, Otis (1980) reports that the mean interval from emergence to initiation of oviposition was 7.8 days for Africanized queens in French Guiana. However, these data were collected by observing queen maturation In colonies that had swarmed and, therefore, the time from emergence to oviposition would be shortened because of a variable period of queen confinement El-10 days (Otis 1980)] within the cells. In another set of data, Otis (1980) reports the mean maturation Interval from eclosion to oviposition was 9.7 days. However, he does







68

not indicate how he determined when eclosion occurred, or if he was using the terms eclosion and emergence Interchangeably. The normal time from eclosion to emergence for queens is approximately 12 hours (Jay 1963).

Because of their high reproductive rate and resultant short swarmto-swarm interval, Africanized honey bees were expected to have a rapid queen maturation interval compared with European honey bees. Fletcher and Tribe suggest "that in the adansonil scutellatata] race, natural selection has worked strongly in favour of minimizing the period between the loss of a queen [from swarming] and the re-establishment of oviposition by a new queen" (1977b, p. 167). The surprising result from this study was that both Africanized and European queens matured at approximately the same rate, determined both by their attractiveness to drones and the time from adult emergence to initiation of oviposition.

As Fletcher and Tribe (1977b) suggested, one would expect natural selection to be operating to minimize the maturation interval for queens, in order to maximize brood production between swarming periods. However, Africanized queens may be under a second and possibly more important selection pressure which may affect their maturation interval. Africanized swarms may travel great distances (Fletcher 1978; Michener 1975). Otis (1980) confirmed that at least some queens issuing with afterswarms had already mated. If new queens issuing with these swarms have mated prior to swarming or mate while enroute, then delayed maturation, particularly with respect to development of ovarian follicles, would be advantageous. Follicular development would increase the queen's weight and make it more difficult for her to fly. Prior to issuing with the prime swarm, older queens usually stop egg laying







69

several days before the swam departs allowing time for their ovaries to recess. Therefore, maturation for Africanized queens may be delayed in order for the swarms with new queens to be able to migrate long distances. Rather than selection operating to shorten the maturation intervals selection may be operating to delay maturation to enable long swarm migration distances.

The variation In queen maturation rates (see Tables 4-8 and 4-9)

observed both within a population and within a queen line suggests that the physiological parameters involved In the process of maturation may be genetically determined. The rate of maturation is an important

economic characteristic for commercial queen producers to consider in their selection programs. Reducing the time from emergence to initiation of opposition can significantly increase the number of

queens produced in each mating colony during the queen-producing season.







70

TABLE 4-1. Experimental matrix for the comparison of total
development times (oviposition to adult emergence)
for both Africanized and European honey bee queens.


EGG GENOTYPES


NURSE BEE GENOTYPEa AFRICANIZED (A26) EUROPEAN (Y5)


AFRICANIZED
A43 A B
A37 C D

COMBINED E F

EUROPEAN

19 G H
27 I J
28 K L
F M N
H 0 P
IBR Q R

COMBINED S T

COMBINED AFRICANIZED
AND EUROPEAN U V

aQueen-cell-producing colony.







71

TABLE 4-2. Total development times (in days from
oviposition to adult emergence) for
Africanized and European honey bee queens:
median, (sample size).


EGG GENOTYPES


NURSE BEE GENOTYPEa AFRICANIZED (A26) EUROPEAN (YS)


AFRICANIZED

A43 14.0 (1) 15.0 (4)
A37 14.5 (7) 15.0 (4)

COMBINED 14.5 (8) 15.0 (8)

EUROPEAN

19 14.5 (15) 15.0 (19)
27 14.5 (17) 15.0 (13)
28 14.0 (13) 14.5 (12)
F 14.0 (10) 14.5 (9)
H 14.0 (7) 14.5 (8)
IBR 14.0 (8) 14.8 (8)

COMBINED 14.2 (70) 15.0 (69)

COMBINED AFRICANIZED
AND EUROPEAN 14.5 (78) 15.0 (77)

aQueen-cel l-producing colony.







72

TABLE 4-3. Analyses for the comparison of queen development
times for both Africanized and European honey bee
genotypes. Letters A V represent different
treatments; see Table 4-1 for explanation.
Kolmogorov-Smirnov one-tailed test, chi-square
distribution, df = 2, alpha 0.05 (Siegel 1956).



A x B NSa
C x D NSa
ExF ,b
G x H
IxJ **
KxL **
MxN **
OxP *
QxR *
S x T U x V



aSmall sample size, chi-square distribution is conservative.
b P<0.05 * = P<0.01 P<0.001.







73

TABLE 4-4. Analyses of queen development times for the
Africanized egg genotype in the different cellproducing colonies. Letters represent different
cell-producing colonies; see Table 4-1 for
explanation. Kolmogorov-Smirnov two-tailed test,
chi-square distribution, df = 2, alpha = 0.05
(Siegel 1956).



A xC NS
AxG NS
Ax I NS
AxK NS
A xM NS
AxO NS
AxQ NS
C xG NS
Cx I NS
C xK NS
C xM NS
C xO NS
CxQ NS
GxI NS
G xK NS
G xM NS
G xO NS
GxQ NS
I xK NS
I xM NS
I xO NS
IxQ NS
K xM NS
K xO NS
K xQ NS
M xO NS
M xQ NS
0 xQ NS







74

TABLE 4-5. Analyses of queen development times for the
European egg genotype in the different cellproducing colonies. Letters represent different
cell-producing colonies; see Table 4-1 for
explanation. Kolmogorov-Smirnov two-tailed test#
chi-square distribution, df =2, alpha =0.05
(Siegel 1956).



B xD NS
B xH NS
B xJ NS
B xL NS
B xN NS
B xP NS
B xR NS
D xH NS
D xJ NS
D xL NS
D xN NS
D xP NS
D xR NS
H xJ NS
H xL NS
H xN NS
H xP NS
H xR NS
J xL NS
J x N NS
j xP NS
J xR NS
L xN NS
L xP NS
L xR NS
N xP NS
N xR NS
P xR NS







75

TABLE 4-6. Total development time (in days from oviposition
to adult emergence) of Africanized queens in
Africanized and European cell-producing colonies:
median, (sample size).


NURSE BEE GENOTYPEa AFRICANIZED EGG GENOTYPE (A26)


AFRICANIZEDb

A43 14.4 (10) A

A37 14.6 (6) B

A43 & A37 14.4 (16) C

EUROPEAN

H1 14.4 (16) D

IBR 14.2 (14) E

H1 & IBR 14.4 (30) F


ANALYSESc
Ax D NS
AxE NS
BxD NS
B xE NS
CxF NS

a(lueen-cell-producing colonies. bAfricanized comb cell size.
cKolmogorov-Smirnov one-tailed test, chi-square distribution, df = 2, alpha = 0.05.







76

TABLE 4-7. Unsealed (egg and larval periods combined) development times
(in days) for Africanized and European queens: median,
(sample size).


EGG GENOTYPES

EFFECT OF tGG
NURSE BEE GENOTYPEa AFRICANIZED (A26) EUROPEAN (Y5) GENOTYPE


19 7.5 (17) 7.5 (19) *

28 7.2 (14) 7.5 (12) *

19 & 28 7.5 (31) 7.5 (31) *


EFFECT OF NURSE BEE
GENOTYPEc NS NS

aCell-producing colonies, European nurse bees, European comb cell size. bKolmogorov-Smirnov one-tailed test, chi-square distribution, df = 2,
* = P CKolmogorov-Smirnov two-tailed test, chi-square distribution, df = 2, alpha = 0.05.







77

TABLE 4-8. Drone response to tethered virgin queens: median
day of response post-emergence, range, (sample
size).


DAY OF RESPONSE LEVEL


RANK 1 RANK 2 RANK 3 RANK 4


AFRICANIZED QUEEN 0a 3.5 3.5 4.0
GENOTYPES -- 1-5 1-5 1-5
(2) (6) (6) (5)


EUROPEAN QUEEN 0a 1.5 4.0 4.5
GENOTYPES -- 1-5 2-5 4-5
(1) (4) (3) (2)


ANALYSESb -- NS NS NS

aDay 0 = day of adult emergence. bKolmogorov-Smirnov one-tailed test, chi-square distribution, df = 2, alpha = 0.05.







78

TABLE 4-9. Time post-emergence to initiation of oviposition:
median, range, (sample size).


DAYS POST-EMERGENCE


AFRICANIZED GENOTYPE (A26) 8.5
7.5-12
(10)



EUROPEAN GENOTYPE (We) 7.5
6-10
(16)


ANALYSISa P<0.05

aKolmogorov-Smirnov two-tailed test, chi-square distribution, df = 2.















CHAPTER V
QUEEN PUPAL WEIGHTS


Introduction

Africanized honey bees in South America are hybridized descendents of African honey bees (__ mellifera scutellata) and European honey bees (primarily 8. M. ligustica and A. m-. mellifera) (Goncalves 1982; Woyke 1969). The annual net reproductive rate of Africanized honey bees in South America is four to five times greater than that of European honey bees in temperate regions: 16 colonies per colony per year compared with 3-3.6 (Otis 1980, 1982a; Winston 1980a; Winston, Taylor and Otis 1983). Differences in reproductive rates between these two honey bee populations may be a result of: 1) colony demography; 2) temperate vs. tropical climate and floral resources; 3) resource utilization behaviors; or 4) a combination of factors. Because Africanized and European honey bees have not been compared under identical experimental conditions, it is not possible to determine to what extent reproductive differences are a result of genetic or environmental parameters.

One demographic parameter associated with rapid colony growth and a high rate of colony reproduction would be a high oviposition rate (Brian 1965; Moeller 1961; Wilson 1971). In the evolution of social insects, queen oviposition rates have increased primarily due to one of the following: increased number of ovarioles, increased length of the


79







80

ovarioles, more rapid egg maturation, and reduction in egg size (Hagan 1954 and Iwata and Sakagami 1966 cited in Wilson 1971; Wilson 1971). Honey bee queens have a very large number of ovarioles (>300) and, for European queens, the number of ovarioles has been shown to be an inherited character (Eckert 1934) which is positively correlated with queen pupal weight (Hoopingarner and Farrar 1959). Queen weight was also found to be correlated with brood production (Boch and Jamieson 1960). If it is assumed that both Africanized and European honey bees have the same relationship between queen weight and brood production, then weights of queens from the two populations can be compared to determine potential differences in fecundity.

In honey bees, differentiation between worker and queen castes is not genetically determined, but rather is regulated by the quantity and quality of food fed to developing larvae during the first 3 days (Beetsma 1979). Therefore, a number of factors other than genotype affect queen size, e.g., age of larvae used to produce queens, population of the cell-producing colony, quantity and quality of food fed to developing larvae, and temperature (Beetsma 1979; Johansson and Johansson 1973; Laidlaw 1979; Weiss 1974; Woyke 1971). Because of differences in queen rearing methods and experimental conditions, previous comparisons of size between Africanized and European queens may be inappropriate. This study was undertaken to compare queen pupal weights for Africanized and European honey bees under identical experimental conditions in Venezuela.

Methods

Four Africanized honey bee lines (A26, A57, A61 and A62) were established from queens removed from feral colonies In an area in








81
eastern Venezuela that had no known European honey bees. They were Identified as Africanized honey bees by their comb cell size, which was significantly smaller than European comb cell size (Chapter III). Two European lines (YK and WE) were established from queens shipped to Venezuela by commericial queen producers in the southeastern U.S.A. Three additional European lines (YD, N and GK) were established from queens shipped to Venezuela from the U.S. Department of Agriculture Bee Breeding and Stock Center Laboratory, Baton Rouge, Louisiana, U.S.A.

Queens were produced from these nine lines by standard queen

rearing methods (Laidlaw 1979). Egg samples from the nine queen mothers were collected by confining the queens to an empty comb within their own colonies using an 8 x 8 cm push-in cage made from 3 mm mesh hardware cloth. -CQueen excluder material was soldered to the tops of the push-in cages, allowing worker bees to move in and out in order to feed and tend the queen (Harbo, Bolten, Rinderer and Collins 1981). Both Africanized and European eggs were collected in European size comb. After approximately 4-6 hours, the queens were released, and combs containing the eggs were put Into a strong colony In order for the eggs to be incubated and for the larvae to be fed. Africanized and European eggs were both put into the same incubator-colony in order to control for any differences in early larval feeding and temperature.

Young larvae approximately 12-15 hours old were transferred

(grafted) into artificial, beeswax, queen-cell cups and then introduced into the cell-producing colonies. Twenty larvae from one of the

Africanized lines and twenty larvae from one of the European lines were grafted into each cell-producing colony. To control for extrinsic factors affecting queen size, analyses of Africanized and European







82

queens were limited to paired comparisons (one Africanized and one European line) that were each simultaneously introduced into the same incubator-colony and then grafted into the same queen-cell-producing colony. Possible effects from different cell-producing colonies on queen pupal weight were evaluated by grafting the same queen lines into different cell-producing colonies.

Only European cell-producing colonies were used because of the difficulty in producing queens in Africanized colonies. Africanized cell-producing colonies remained disturbed for a long period of time after the grafted larvae were introduced, which resulted in poor acceptance (survival) of the larvae (see Chapter IV).

Queen pupal weights are used for comparison because adult weights vary with respect to engorgment of food, dehydration, feces accumulation, and differential ovariole development. Although queen pupal weights vary with age of the pupae, there is a period from the 10th through the 13th day post-oviposition when queen pupal weight is constant (Table C-1). Queen pupal weight comparisons can therefore be made during this period (Hoopingarner and Farrar 1959). Although there is a 0.5 day difference in development time between Africanized and European queens (Chapter IV), the 3-4 day pupal period during which there is no significant weight change is of sufficient duration to allow Africanized and European queens to be accurately and consistently compared. Africanized and European queen pupae were weighed on the 11th day post-oviposition. Queen cells from each of the lines were randomly selected to avoid any position effect from location on the grafting frame. Weights were measured to the nearest 1.0 mg using either a Mettler Type H4 or H6 balance.







83

Queen cell lengths were measured at the time the queen pupal

weights were determined. A calipers was used to determine the external length from the base to the apex of the queen cell.

Res ul1ts

Queen pupal weights for four Africanized and five European lines are presented in Table 5-1. European queen pupal weights were significantly larger than Africanized queen pupal weights for two different pairwise comparisons, (YK vs. A26 and YK vs. A57; P<0.05 to

P<0.001; Mann-Whitney U test, one-tailed). Africanized queen pupal weights were significantly larger in one pairwise comparison (A62 vs. N; P<0.02; Mann-Whitney U test, two-tailed). For three pairwise comparisons, there was no statistical difference (A26 vs. WE, A57 vs. YD, and A61 vs. GK; Mann-Whitney U test, one-tailed, alpha = 0.05). Because different cell-producing colonies had no significant effect on queen pupal weights (see below), the means for the nine queen lines can be ranked and analyzed (Table 5-2). There was no significant difference between the Africanized and European honey bee populations for queen pupal size (Mann-Whitney U test, one-tailed, alpha = 0.05).

Queen cell lengths for Africanized and European lines are presented in Table 5-3. In six out of eight pairwise comparisons, there was no significant difference in queen cell lengths between Africanized and European queens (Mann-Whitney U test, one-tailed, alpha = 0.05). For the pair in cell-producing colony 2, the European line was significantly larger than the Africanized line (P







84
Spearman's rank correlation coefficient was determined for queen

pupal weights and queen cell lengths (Table 5-4). In general, there was no significant correlation between queen pupal weight and queen cell length (alpha =0.05). However, one Africanized line (A26) In cellproducing colony 2 had a significant correlation (P<0.05) and one European line (YK) in cell-producing colony 4 had a significant correlation (P<0.01).

The effect of cell-producing colonies on queen pupal weights and

queen cell lengths is presented in Table 5-5. There was no significant difference for Africanized queen line A26 in four different cellproducing colonies (one-way analysis of variance, alpha = 0.05); nor was there a significant difference for Africanized queen line A57 in two different cell-producing colonies (Mann-Whitney U test, two-tailed,. alpha = 0.05). There was no significant difference in pupal weights for the European queen line YK in four different cell-producing colonies (one-way analysis of variance, alpha = 0.05), but there was a significant effect of cell-producing colonies on queen cell length (P<0.001). When cell-producing colony 2 was removed from the analysis, there was no significant difference in queen cell length.


Uiscussion
If we assume for both Africanized and European honey bees that queen weight is correlated with egg production or fecundity (Boch and Jamieson 1960), we would then expect that egg laying rates would follow the same ranking as presented In Table 5-2 for queen pupal weights. Based on these pupal weights# we would predict that there would be no difference in egg laying rates for the Africanized and European honey bee populations. In fact, when egg laying rates for Africanized and







85

European honey bee queens were compared, there was no significant difference between queens from the two populations (Chapter VI). There were, however, significant differences in pupal weights between individual queen lines both between and within each population (Table 51). There were also significant differences in egg laying rates between individual queen lines both between and within the two populations (Chapter VI).

In this study, Africanized queens were reared in European colonies because of low acceptance (survival) of grafted cells in Africanized colonies (Chapter IV). Because queen-worker caste differentiation in honey bees is regulated by larval feeding (Beetsma 1979), rearing Africanized queens in European colonies may have obscured differences in pupal weights between Africanized and European queens. Possibly, European worker bees may rear larger Africanized queens than would

Africanized worker bees because European worker bees are themselves larger (Chapter III), and may feed the developing queen larvae differently. Although virgin European queens have been reported to weigh more than virgin Africanized queens--208 vs. 199 mg (Goncalves, Kerr and Nocoes 1972 cited in Michener 1975)--there was no indication of conditions under which the queens were reared.

Further analysis of queen weights between Africanized and European honey bees is needed, preferably in a 2 x 2 experimental design: Africanized and European queens reared in both Africanized and European cell-producing colonies. In addition, the relationship between queen pupal weights and brood production needs to be evaluated for both Africanized and European honey bee lines to determine if the same relationship exists for both populations.







86

The European queen lines evaluated were a diverse representation of the European population from North America, whereas the Africanized queen lines may only reflect a small portion of the Africanized population. The location for the sources of the Africanized lines was limited to feral colonies found In one area of eastern Venezuela. A greater diversity of Africanized lines needs to be evaluated in order to be able to generalize about queen pupal weights and oviposition rates for the population as a whole.







87

TABLE 5-1. Comparison of Africanized and European queen pupal weights
(mg): mean + SD, (sample size), (genotype).

AFRICA NIZED EUROPEAN

CELL BUILDERa GENOTYPES GENOTYPES ANALYSESb


1 257.1 + 7.6 286.4 + 12.0
(9) (A26) (9) (YK)

2 255.8 + 9.2 284.4 + 20.6 **
(9) (A26) (9) (YK)

3 262.3 + 6.4 266.7 + 9.7 NS
(3) (A26) (9) (WE)

4 260.0 + 9.0 272.5 + 16.5
(11) (A57) (13) (YK)

5 243.0 + 13.4 282.6 + 12.0
(3) (A26) (3) (YK)

6 248.6 + 14.3 264.0 + 15.4 NS
(4) (A57) (5) (YD)

7 291.8 + 7.9 257.3 + 18.2 _c
(5) (A62) (4) (N)

8 237.2 + 21.7 232.9 + 0.4 NS
(5) (A61) (2) (GK)

aCell-produclng colonies; European nurse bees and European comb
bcell size.
Mann-Whitney U test, one-tailed, alpha = 0.05;
* = P<0.05, ** = P<0.01, *** = P<0.O01.
CDifference in wrong direction for one-tailed test; two-tailed test results in a P






88

TABLE 5-2. Queen pupal weights for the nine lines
analyzed.

POPULATION QUEEN LINE MEAN PUPAL WEIGHT(MG)


European GK 233
Africanized A61 237
Africanized A26 256
European N 257
Africanized A57 257
European YD 264
European WE 267
European YK 280
Africanized A62 292



ANALYSISa NS


aMann-Whitney U test, one-tailed, alpha = 0.05.







89
TABLE 5-3. Comparison of Africanized and European queen cell
lengths (mm): mean +SD, (sample size),
(genotypes).


AFRICANIZED EUROPEAN
CELL BUILDERa GENOTYPES GENOTYPES ANALYSESb


1 2.58 + 0.1 2.50 + 0.1 NS
(9) (A26) (9) (YK)

2 2.62 + 0.2 2.70 + 0.1
(9) (A26) (9) (YK)

3 2.54 + 0.1 2.48 + 0.1 NS
(3) (A26) (9) (WE)

4 2.58 + 0.1 2.44 + 0.1 _c
(7) (A57) (11) (YK)

5 2.67 + 0.1 2.57 + 0.1 NS
(3) (A26) (3) (YK)

6 2.50 + 0.02 2.57 + 0.1 NS
(4) (A57) (5) (YD)
7 2.78 + 0.04 2.81 + 0.1 NS
(5) (A62) (3) (N)

8 2.79 + 0.1 2.76 + 0.1 NS
(5) (A61) (2) (GK)

aCell-producing colonies; European nurse bees and European comb
b cell size.
Mann-Whitney U test, one-tailed, alpha = 0.05; P<0.05. CDifference In wrong direction for one-tailed test; two-tailed test results in a P<0.02.







90

TABLE 5-4. Correlation of queen cell length (mm) and queen pupal
weight (mg): mean + SD, (sample size). Measurement
made on day 11.25 post-oviposition.


QUEEN CELL QUEEN PUPAL
QUEEN GENOTYPE LENGTH WEIGHT CORRELATIONSa


AFRICANIZED

A26 (CB1)b 2.58 + 0.1 257.1 + 7.6 NS
(9) (9)
A26 (CB2) 2.62 + 0.2 255.8 + 9.2 *
(9) (9)
A26 (CB3) 2.54 + 0.1 262.3 + 6.4 -(3) (3)
A26 (CB5) 2.67 + 0.1 243.0 + 13.4 -(3) (3)
A57 (CB4) 2.58 + 0.1 259.1 + 10.8 NS
(7) (7)
A57 (CB6) 2.50 + 0.02 248.6 + 14.3 NS
(4) (4)
A62 (CB7) 2.78 + 0.04 291.8 + 7.8 NS
(5) (5)
A61 (CB8) 2.79 + 0.1 237.2 + 21.7 NS
(5) (5)
EUROPEAN

YK (CB1) 2.50 + 0.1 286.4 + 12.0 NS
(9) (9)
YK (CB2) 2.70 + 0.1 284.4 + 20.6 NS
(9) (9)
YK (CB4) 2.44 + 0.1 272.2 + 17.8 **
(11) (11)
YK (CB5) 2.57 + 0.1 282.6 + 12.0 -(3) (3)
WE (CB3) 2.48 + 0.1 268.4 + 8.8 NS
(8) (8)
YD (CB6) 2.57 + 0.1 264.0 + 15.4 NS
(5) (5)
N (CB7) 2.81 + 0.1 264.3 + 14.0 -(3) (3)
GK (CB8) 2.76 + 0.1 232.9 + 0.4 -(2) (2)

aSpearman's rank correlation coefficients, alpha = 0.05;
* = P<0.05; ** = P<0.01. bCB = cell-producing colony number; refer to Table 5-1 for explanation.









TABLE 5-5. Effect of cell-producing colony on queen cell length and
queen pupal weight: mean + SD, (sample size).


AFRICANIZED QUEEN GENOTYPE (A26)

CB1a CB2 CB3 CB5 ANALYSESb


QUEEN CELL 2.58 2.62 2.54 2.67 NS
LENGTH + 0.1 + 0.2 + 0.1 + 0.1
(9) (9) (3) (3)

QUEEN PUPAL 257.1 255.8 262.3 243.0 NS
WEIGHT + 7.6 + 9.2 + 6.4 + 13.4
(9) (9) (3) (3)


AFRICANIZED QUEEN GENOTYPE (A57)

CB4 CB6 ANALYSESc


QUEEN CELL 2.58 2.50 NS
LENGTH + 0.1 + 0.02
(7) (4)

QUEEN PUPAL 260.0 248.6 NS
WEIGHT + 9.0 + 14.3
(11) (4)


EUROPEAN QUEEN GENOTYPE (YK)

CB1 CB2 CB4 CB5 ANALYSESb


QUEEN CELL 2.50 2.70 2.44 2.57 ***d
LENGTH + 0.1 + 0.1 + 0.1 + 0.1
(9) (9) (11) (3)

QUEEN PUPAL 286.4 284.4 272.5 282.6 NS
WEIGHT + 12.0 + 20.6 + 16.5 + 12.0
(9) (9) (13) (3)



aCB = cell-producing colony number; refer to Table 5-1 for explanation. bOne-way analysis of variance, alpha = 0.05; *** = P<0.001. cMann-Whitney U test, two-tailed, alpha = 0.05. dwith CB2 removed, ANOVA Is NS.















CHAPTER VI
EGG-LAYING AND BROOD PRODUCTION RATES DURING THE FIRST BROOD CYCLE


Introduction

Africanized honey bees in South America are descendents from the hybridization of African honey bees (&La mellifera scutellata) and European honey bees (primarily A.. M. ligustica and A. m. mellifera) (Goncalves 1982; Woyke 1969). In tropical and sub-tropical regions of South America, Africanized bees have been more successful than European bees as determined by their rapid rates of dispersal and high population densities (Mlchener 1975; Taylor 1977, 1985). It is not surprising that Africanized bees are more successful in these regions because they are descendents of honey bees that evolved under similar tropical conditions in Africa.

Rates of dispersal and population densities achieved by Africanized honey bees require a high colony reproductive rate. Africanized honey bees in South America have a reproductive rate that is four to five times greater than the reproductive rate of European honey bees In North America (Otis 1980, 1982a; Winston 1980a; Winston, Taylor and Otis 1983). However, based on this comparison, one cannot identify the

factors that account for differences in reproductive rates nor identify the factors leading to the success of Africanized bees in South America. Because the comparison of reproductive rates was not based on data collected under similar environmental or experimental conditions, it is


92




Full Text
40
As drones emerged, they were placed into special holding cages and
maintained in a colony so that worker bees could feed them until they
matured. This manipulation insured that the drones used for
inseminations were from the desired queen lines.
To collect eggs for the bee size experiments, queens from the nine
genotypes were confined for five hours in their own colonies to a
section of Africanized comb (mean cell size = 4.8 mm), using 8 x 8 cm
push-in cages. These cages were made from 3 mm mesh hardware cloth and
had queen excluder material soldered to the top to enable worker bees to
pass through to tend the queen (Harbo, Bolten, Rinderer and Collins
1981). After five hours, the queens were removed from the combs. The 8
x 8 cm sections of comb with eggs from each queen were cut out and
fitted into special frames. The nine sections were then placed in a
strong Africanized colony (Africanized nurse bees and Africanized comb
cell size) for development. The following day, eggs were collected in
European combs (mean cell size = 5.4 mm) using the same procedure with
the same nine queens except that the nine sections were put into a
European colony (European nurse bees and European comb cell size) for
development. Having all nine egg sources for each comb cell size
treatment (Africanized or European) develop in the same colony
controlled for additional variables affecting development and bee size:
temperature and humidity, nurse bee genotypes and colony size (see
Chapter II).
Fresh pupal weights were compared for each of the nine genotypes
reared in both Africanized and European comb cell sizes. Pupal weights
were measured on the 16th day after oviposition. This age corresponds
to the period during pupal development of least weight change (Melampy


47
characterizes the unreliable nectar availability in Africa (Tribe and
Fletcher 1977). Smaller bee size increases the resource utilization
efficiency of Africanized honey bees and may be a factor in the success
and high reproductive rates of Africanized honey bees compared with
European honey bees in tropical areas of South America (see Chapter
VIII).
I suggest two other hypotheses to explain the advantages of smaller
size in the Africanized population. First smaller size is more
efficient with respect to dissipating heat loads in tropical habitats
(see also Heinrich 1979b). Fletcher (1978) reports that foraging may
stop during the hottest part of the day, which would avoid the
disadvantages of smaller size with respect to gaining a heat load. The
sizes of two other subspecies of honey bees in Africa support this
hypothesis. One of the smallest subspecies in Africa A. oi. 1 itorea, is
found in a very hot and dry area along the coast of Kenya and Tanzania.
One of the largest subspecies, A. m. monticola, is found at higher
elevations and colder temperatures on Mount Kenya.
Secondly, the advantage of smaller bee size may actually lie with
the advantages of smaller cell size. For a given nest cavity volume, a
larger number of worker bees can be produced if cell sizes are smaller.
There is approximately a 25% increase in the number cells for a given
comb area with smaller Africanized comb cells compared with larger
European comb cells. Considering the advantages of increased worker
numbers in a colony (Wilson 1971), the increase in worker numbers as a
result of smaller cell size may be important, particularly if nest
cavity volumes are limited.


38
The importance of the genetic component to bee size can be inferred
from the fact that different geographic populations of honey bees differ
with respect to worker bee size (Alpatov 1929; Ruttner 1968, 1975,
1976a, 1976b; Wafa, Rashad and Mazeed 1965). Because honey bee queens
mate with many different drones (Adams, Rothman, Kerr and Paulino 1977;
Peer 1956; Roberts 1944; Taber 1954; Taber and Wendel 1958), the genetic
component becomes an additional factor affecting size variation.
Africanized honey bees in Venezuela (descendents of A. ott..
scutellata) were smaller and had a smaller comb cell diameter (mean 4.8
mm between opposite sides of the hexagonal cells in the comb) compared
with European bees in Venezuela (mean 5.4 mm) (Tables B-3 and B-4;
Rinderer, Tucker and Collins 1982). An opportunity, therefore, existed
to experimentally evaluate the interaction of both genotype and comb
cell size on resultant worker bee size and size variation by studying
both the Africanized and European honey bee populations under identical
experimental conditions. The results from this study provide
information not only on the proximal question involving the factors
affecting bee size but also provide a mechanism by which size variation
may be reduced within a honey bee colony.
Methods
Nine genotypes were evaluated: three Africanized, three European,
and three reciprocal hybrids. The Africanized genotypes (A26, A57,
and B39) were established from queens removed from feral colonies
located in an area in eastern Venezuela with no known European honey
bees. They were identified as Africanized honey bees by their comb cell
sizes which were significantly smaller than European comb cell sizes
(Tables B-3 and B-4; Michener 1972, 1975; Rinderer, Tucker and Collins


TABLE OF CONTENTS
Page
ACKNOWLEDGEMENTS 111
ABSTRACT v11
CHAPTERS
I INTRODUCTION 1
Evolutionary Origin and Distribution of Honey Bees 1
Importation of African Honey Bees into Brazil and
Their Dispersal Throughout South and Central America . 1
Characteristics of Africanized Honey Bees 3
Purpose of My Research 6
Identification of Honey Bees Used in My Research 7
When and Where Research Was Conducted 7
II WORKER BEE DEVELOPMENT TIMES .... 9
Introduction .... 9
Methods 14
Results 18
Discussion 20
IIIINTERACTION OF MATERNAL GENOTYPE, EGG GENOTYPE AND COMB
CELL SIZE ON HONEY BEE WORKER SIZE AND SIZE VARIATION ... 35
Introduction ............ 35
Methods ..... 38
Results 41
Discussion 42
IVQUEEN DEVELOPMENT AND MATURATION .... 55
Introduction 55
Methods 57
Results ................. 63
Discussion 65
v


25
TABLE 2-2
Interaction of egg genotype, comb cell size, and nurse
bee genotype on worker bee development time (days):
median, (range), mean + SD, (n = sample size).
AFRICANIZED EGG GENOTYPE (A26)
USa
SBb
TDTC
AFRICANIZED COMB CELLS
AFRICANIZED NURSE
BEES (A55)
4.0
(4-5)
4.2 + 0.4
(n = 30)
12.0
(11-12)
11.6 + 0.5
(n = 30)
19.0
(18-20)
18.8 + 0.5
(n = 30)
EUROPEAN NURSE
BEES (H2)
4.0
(4-5)
4.4 + 0.5
(n = 29)
12.0
(11-12)
11.6 + 0.5
(n = 29)
19.0
(19-20)
19.1 + 0.2
(n = 29)
EUROPEAN COMB CELLS
AFRICANIZED NURSE
BEES (A41)
4.0
(4-5)
4.2 + 0.4
(n = 30)
12.0
(11-12)
11.7 + 0.5
(n = 30)
19.0
(18-19)
18.9 + 0.2
(n = 30)
EUROPEAN NURSE
BEES (IBR877)
4.0
(4-5)
4.3 + 0.5
(n = 26)
12.0
(11-12)
11.7 + 0.5
(n = 26)
19.0
(19)
19.0 + 0
(n = 26)
TOTALS
4.0
(4-5)
4.3 + 0.4
(n = 115)
12.0
(11-12)
11.6 + 0.5
(n = 115)
19.0
(18-20)
18.9 + 0.3
(n = 115)
aUS = unsealed brood (unsealed larval development period only).
bSB = sealed brood (pre-pupae and pupae).
CTDT = total development time (oviposition to adult emergence).


48
Because maternal inheritance affects bee size, methods that use
components of size to identify Africanized bees, e.g., morphometric
analysis, may be invalid. Offspring from the cross of a European queen
x Africanized drone (SDY10 and SDY11) are the same weight as offspring
from European queen x European drone (SDY1) (Table 3-4). More
importantly, the offspring from the cross of a European queen x
Africanized drone are significantly different from offspring from
Africanized queen x Africanized drone and Africanized queen x European
drone matings. The European queen x Africanized drone mating represents
the most probable scenario for initial hybridization in North America
(see Chapter VII). That is, a virgin queen from a managed or a feral
European colony mates with Africanized drones and produces offspring
with a 50% Africanized genome. Analyzing the offspring using size as a
component for identification may result in a false negative
identification of Africanized bees. The extent of the problem would
depend upon the degree to which particular linear measurements are
either affected by maternal inheritance and/or are correlated with bee
weight. As Daly, Hoelmer, Norman and Allen (1982) point out, there is a
'difficulty in using phenotype characters to identify genetically
different, but closely related populations" (p. 593).


106
TABLE 6-1. Daily egg laying rates of Africanized and European queens
with European nurse bees trial 1.
ONE-DAY HIGH
MEAN + SD (n)
AFRICANIZED QUEENS
A57 (W42)
550
473.2 + 68.5 (6)
A62 (W81)
857
825.0 + 47.0 (3)
EUROPEAN QUEENS
YK (Y4)
951
922.0 + 41.0 (2)
YD5 (Y42)
763
689.6 + 66.4 (5)
GK30 (Y63)
710
636.7 + 80.8 (3)
GK30 (Y64)
705
575.6 + 101.3 (5)
ANALYSIS3
NS
^ann-Wh itney U test, two-tailed, alpha = 0.05; evaluated for all
samples (n = 24).


20
exact timing of developmental changes. Second, the process of sealing
is not a precise developmental stage and may take from six hours
(Lindauer 1953) to 24 hours (Jay 1963). When the unsealed and sealed
brood stages are combined, the proportional differences between the two
populations are the same as for egg development times and total
development times (Table 2-8).
Mortality for different developmental stages for each colony
treatment is presented in Table 2-9 (see also Table A-2). Mortality was
high (26-37%) for larvae in the experimental colony (H2) with European
nurse bees on Africanized comb cell size. The high mortality during the
larval stage may be a result of the reduced ability of larger European
nurse bees to feed the developing larvae in smaller Africanized comb
cells. There was also a high egg mortality recorded for European eggs
in A41 and IBR87734 and 70%, respectively. Woyke (1977) reports
normal mortality may be as high as 10-50% depending on the season.
Garofalo (1977) also reports varying mortalities depending on both the
size of the colony and the time of year: eggs 10-25%, larvae 11-37%,
pupae 5-7%, and all developmental stages combined 25-53%.
Discussion
This study is the first to evaluate worker bee development times
between Africanized and European honey bees as an interaction between
egg genotype and colony-level parameters. Differences in worker bee
development times were independent of the colony-level parameters of
comb cell size and nurse bee genotype but were dependent on egg genotype
differences between the Africanized and European populations. The
difference in development times between these two populations was not as


TABLE 4-9. Time post-emergence to initiation of oviposition
median, range, (sample size).
AFRICANIZED GENOTYPE (A26)
DAYS POST-EMERGENCE
8.5
7.5-12
(10)
EUROPEAN GENOTYPE (We)
7.5
6-10
(16)
ANALYSIS3
P<0.05
Kolmogorov-Smirnov two-tailed test, chi-square distribution,
df = 2.


TABLE D-l. Accuracy of technique used to estimate number of bees in
colony.
SAMPLE NO.
COUNTED
ESTIMATED3
DIFFERENCE
% DIFFERENCE
1
2284
2324
40
1.8
2
2906
2918
12
0.4
3
3085
3080
5
0.2
4
3079
3124
45
1.5
5
2657
2717
60
2.2
6
2046
2102
56
2.7
aThe number of
adult bees was estimated by determining the
mean
individual bee weight from three, 150-200
bee samples.
The total
weight of bees in each sample was then divided by the mear
i individual
bee weight to get an estimate of the total
number of bees
in each
sample.
167


114
TABLE 6-9. Colony adult population changes during first brood cycle.
ESTIMATED DAILY
DAY 0a DAY 12b DAY 17c MORTALITY RATEd
AFRICANIZED NURSE BEES
AFRICANIZED QUEENS
A62 (W81)
A57 (W42)
EUROPEAN QUEEN
GK30 (Y63)
EUROPEAN NURSE BEES
AFRICANIZED QUEENS
A62 (W85)
AS7 (W41)
EUROPEAN QUEEN
GK30 (Y61)
10,169
10,169
4,850
4,408
10,169
6,511
10,435
10,435
5,375
6,102
10,435
6,321
2,634
2,007
443
480
4,987
305
MEAN = 409
3,267
4,297
422
361
4,607
343
MEAN = 375
ANALYSIS
NS
^Population at Day 0 was estimated as described in methods.
Population at Day 12 estimated by: Day 0 [(Day 0 Day 17)(12/17)].
^Population at Day 17 was estimated as described in methods.
(Day 0 Day 17)/17.
^iann-Whitney U test, one-tailed, alpha = 0.05.


101
Results
Daily Egg Laving Rates
There were no significant differences in daily egg laying rates
between Africanized and European queens (Tables 6-1 to 6-5). There
were, however, significant differences between individual queen lines
both between the two populations and within each population (Table 6-6).
There was no significant effect of worker bee type (Africanized vs.
European) on egg laying rates for either Africanized or European queens
(Table 6-5). Worker bee type also had no effect on daily egg laying
rates of sister queens (Table 6-7).
Brood Production Rates during First Brood Cycle
The amounts of unsealed, sealed and total brood at day 12 and at
day 17 for each of the experimental colonies are presented in Table 6-8.
There was no significant effect of worker bee population type on brood
production at either day 12 or day 17. When all colony treatments were
combined, total brood produced at day 12 was significantly correlated
with total brood produced at day 17 (P<0.05).
Changes in worker bee population for each colony are shown in Table
6-9. The estimated daily mortality rates for Africanized worker bees
were not significantly different than those for European worker bees.
The numbers of unsealed brood, sealed brood and total brood at day
12 and day 17 expressed as percentages of the adult population are
presented in Table 6-10. There was no significant difference between
Africanized and European worker bees.
Sister queens are compared in Table 6-11. The performance of the
European queen pair was similar with either Africanized and European
worker bees. One sister of Africanized queen pair (A62) performed


107
TABLE 6-2. Daily egg laying rates of Africanized and European queens
with European nurse bees, trial 2.
ONE-DAY HIGH MEAN + SD (n)
AFRICANIZED QUEENS
A57 (W42)
A62 (W81)
EUROPEAN QUEENS
GK30 (Y63)
740 699.8 +39.5 (4)
953 915.8 + 30.7 (4)
779 736.8 + 32.2 (4)
ANALYSIS3
NS
^iann-Whitney U test, two-tailed, alpha = 0.05; evaluated for all
samples (n = 12).


31
TABLE 2-6. Total worker bee development times. Hypotheses were tested
using Kolmogorov-Smirnov one-tailed test, chi-square
distribution, df = 2, alpha = 0.05 (Siegel 1956).
HI: Worker bee development is faster for Africanized genotypes than for
European genotypes.
A X B
***a
C X D
***
E X F
***
G X H
***
A X H
**
AC X BD
***
EG X FH
***
AE X BF
***
CG X DH
***
ACEG X BDFH
***
H2: Worker bee development is more rapid in Africanized comb cells than
in European comb cells.
AXE NS
C X G NS
B X F NS
D X H NS
AC X EG NS
BD X FH NS
H3: Worker bee development is more rapid with Africanized nurse bees
than with European nurse bees
A X C NS
E X G NS
B X D NS
F X H NS
AE X CG NS
BF X DH NS
H4: Worker bee development is more rapid with Africanized comb cells
and Africanized nurse bees than with European comb cells and
European nurse bees.
A X G NS
B X H NS
a ** = P<0.01
*** = P<0.001.


TABLE B-4. Comparison of Africanized and European comb cell diameter
and comb cell depth: mean + SD, range (sample size).
DIAMETER3
DEPTH
(mm)
(mm)
CORRELATIONS
AFRICANIZED COMB CELLS
4.8 + 0.1
11.8 + 0.2
NS
4.7 4.9
11.4 12.3
(17)
(17)
EUROPEAN COMB CELLS
5.4 + 0.05
12.2 + 0.4
NS
5.3 5.4
11.5 12.8
(26)
(26)
ANALYSES0
P<0.001 PcO.OOl
determined by measuring 10 horizontal, consecutive cells; cell-wall
thickness not considered.
bSpearman,s rank correlation coefficient, alpha = 0.05.
t-test, one-tailed.


178
Seeley, T.D., RH. Seeley and P. Akratanakul. 1982. Colony defense
strategies of the honeybees in Thailand. Ecol. Monogr. 52:43-63.
Shearer, D.A., R. Boch, R.A. Morse and F.M. Laigo. 1970. Occurrence of
9-oxodec-trans-2-enoic acid in queens of Apis dorsata, Apis cerana
and Apis mellifera. J. Insect Physiol. 16:1437-1441.
Siegel, S. 1956. Nonparametric statistics for the behavioral sciences.
McGraw-Hill Book Co., New York.
Simpson, J. 1966. Congestion of adult honeybees with and without
adequate hive space. J. Apic. Res. 5:59-61.
Simpson, J. 1973. Influence of hive-space restriction on the tendency
of honeybee colonies to rear queens. J. Apic. Res. 12:183-186.
Simpson, J., and I.B.M. Riedel. 1963. The factor that causes swarming
by honeybee colonies in small hives. J. Apic. Res. 2:50-54.
Smith, F.G. 1958a. Beekeeping observations in Tanganyika 1949-1957.
Bee World 39:29-36.
Smith, F.G. 1958b. Communication and foraging range of African bees
compared with that of European and Asian bees. Bee World 39:
249-252.
Smith, F.G. 1961. The races of honeybees in Africa. Bee World 42:
255-260.
Smith, M.V. 1972. Marking bees and queens. Bee World 53:9-13.
Sokol, R.R., and F.J. Rohlf. 1969. Biometry. W.H. Freeman and Co.,
San Francisco.
Stort, A.C. 1974. Genetic study of aggressiveness of two subspecies of
Apis mellifera in Brazil, 1. Some tests to measure aggressiveness.
J. Apic. Res. 13:33-38.
Stort, A.C. 1975a. Genetic study of aggressiveness of two subspecies
of Apis mellifera in Brazil, 2. Time at which the first sting
reached a leather ball. J. Apic. Res. 14:171-175.
Stort, A.C. 1975b. Genetic study of the aggressiveness of two
subspecies of Apis mellifera in Brazil, IV. Number of stings in the
gloves of the observer. Behavior Genetics 5:269-274.
Stort, A.C. 1975c. Gentica! study of the aggressiveness of two
subspecies of Apis mel1ifera in Brasil, V. Number of stings in the
leather ball. J. Kans. Ent. Soc. 48:381-387.
Stort, A.C. 1976. Genetic study of the aggressiveness of two
subspecies of Apis mellifera in Brazil, III. Time taken for the
colony to become aggressive. Ciencia e Cultura 28:1182-1185.


61
queens was established on each day of the experiment. Each queen was
tested more than once on each day and always in a new mesh bag, to avoid
any pheromone accumulation or contamination. Each testing session was
begun by suspending an older queen that had previously been determined
to be maximally attractive, in order to insure that a responding drone
population was available. This process was also repeated if the testing
session was interrupted by rain, extreme cloudiness, or high winds
conditions that normally reduce drone flight activity. The testing took
place between 1400 and 1600 hours.
Drone response was evaluated by assigning one of the following
ranks to the test queen:
Rank 0 = no response
Rank 1 = drones oriented to the test subject but only flew past;
no circling of the test subject
Rank 2 = drones oriented to the test subject and persisted in a
wide circling formation more than 2 m from the subject
Rank 3 = drones oriented to the test subject and formed a loose
comet-like formation down wind more than 0.5 m to the
test subject; formation was volatile, continually
fragmenting and reforming; drones did not land on the
mesh bag
Rank 4 = drones oriented to the test subject and formed a tight
comet-like formation down wind less than 0.5 m from the
test subject; formation was persistent and did not
fragment even as the test subject was lowered; drones
landed on and walked over the mesh bag.


CHAPTER VI
egg'laying Arc BROOD production rates during the first brood cycle
Introduction
Africanized honey bees in South America are descendents from the
hybridization of African honey bees (Ad is me!1 ifera scutellata) and
European honey bees (primarily A. [n. 1 iaustica and A. m. mel 1 ifera)
(Goncalves 1982; Woyke 1969). In tropical and sub-tropical regions of
South America, Africanized bees have been more successful than European
bees as determined by their rapid rates of dispersal and high population
densities (Michener 1975; Taylor 1977, 1985). It is not surprising that
Africanized bees are more successful in these regions because they are
descendents of honey bees that evolved under similar tropical conditions
in Africa.
Rates of dispersal and population densities achieved by Africanized
honey bees require a high colony reproductive rate. Africanized honey
bees in South America have a reproductive rate that is four to five
times greater than the reproductive rate of European honey bees in North
America (Otis 1980, 1982a; Winston 1980a; Winston, Taylor and Otis
1983). However, based on this comparison, one cannot identify the
factors that account for differences in reproductive rates nor identify
the factors leading to the success of Africanized bees in South America.
Because the comparison of reproductive rates was not based on data
collected under similar environmental or experimental conditions, it is
92


26
TABLE 2-2~extended.
EUROPEAN EGG GENOTYPE (Y5)
US
SB
TDT
5.0
(4-5)
4.9 + 0.3
(n = 37)
12.0
(11-12)
11.8 + 0.4
(n = 37)
20.0
(19-20)
19.6 + 0.5
(n = 37)
5.0
(4-5)
4.9 + 0.4
(n = 22)
12.0
(12)
12.0 + 0
(n = 22)
20.0
(19-20)
19.9 + 0.4
(n = 22)
5.0
(4-6)
5.0 + 0.4
(n = 19)
12.0
(12-13)
12.0 + 0.2
(n = 19)
20.0
(19-21)
20.0 + 0.3
(n = 19)
5.0
(4-6)
4.7 + 0.8
(n = 7)
12.0
(12-13)
12.1 + 0.4
(n = 7)
20.0
(19-21)
19.8 + 0.7
(n = 7)
5.0
(4-6)
4.9 + 0.4
(n = 85)
12.0
(11-13)
11.9 + 0.4
(n = 85)
20.0
(19-21)
19.8 + 0.4
(n = 85)


68
not indicate how he determined when eclosin occurred or if he was
using the terms eclosin and emergence interchangeably. The normal time
from eclosin to emergence for queens is approximately 12 hours (Jay
1963).
Because of their high reproductive rate and resultant short swarm-
to-swarm interval Africanized honey bees were expected to have a rapid
queen maturation interval compared with European honey bees. Fletcher
and Tribe suggest "that in the adansonii f=scute11ata] race natural
selection has worked strongly in favour of minimizing the period between
the loss of a queen [from swarming] and the re-establishment of
oviposition by a new queen" (1977b, p. 167). The surprising result from
this study was that both Africanized and European queens matured at
approximately the same rate, determined both by their attractiveness to
drones and the time from adult emergence to initiation of oviposition.
As Fletcher and Tribe (1977b) suggested, one would expect natural
selection to be operating to minimize the maturation interval for
queens, in order to maximize brood production between swarming periods.
However, Africanized queens may be under a second and possibly more
important selection pressure which may affect their maturation interval.
Africanized swarms may travel great distances (Fletcher 1978; Michener
1975). Otis (1980) confirmed that at least some queens issuing with
afterswarms had already mated. If new queens issuing with these swarms
have mated prior to swarming or mate while enroute, then delayed
maturation, particularly with respect to development of ovarian
follicles, would be advantageous. Follicular development would increase
the queen's weight and make it more difficult for her to fly. Prior to
issuing with the prime swarm, older queens usually stop egg laying


TABLE 2-1. Experimental matrix for evaluating interaction of egg
genotype, comb cell size, and nurse bee genotype on worker
development times. A H represent each treatment.
AFRICANIZED EUROPEAN
EGG GENOTYPE (A26) EGG GENOTYPE (Y5)
AFRICANIZED COMB CELLS
AFRICANIZED NURSE
BEES (A55) A B
EUROPEAN NURSE
BEES (H2) C D
EUROPEAN COMB CELLS
AFRICANIZED NURSE
BEES (A41) E F
EUROPEAN NURSE
BEES (IBR877) G H


91
TABLE 5-5 Effect of cell-producing colony on queen cell length and
queen
pupal weight: mean
+ SD, (sample size).
AFRICANIZED QUEEN
GENOTYPE
(A26)
CBla
CB2
CB3
CB5
ANALYSES13
QUEEN CELL
LENGTH
2.58
+ 0.1
(9)
2.62
+ 0.2
(9)
2.54
+ 0.1
(3)
2.67
+ 0.1
(3)
NS
QUEEN PUPAL
WEIGHT
257.1
+ 7.6
(9)
255.8
+ 9.2
(9)
262.3
+ 6.4
(3)
243.0
+ 13.4
(3)
NS
AFRICANIZED QUEEN
GENOTYPE
(A57)
CB4
CB6
ANALYSES
c
QUEEN CELL
LENGTH
2.58
+ 0.1
(7)
2.50
+ 0.02
(4)
NS
QUEEN PUPAL
WEIGHT
260.0
+ 9.0
(11)
248.6
+ 14.3
(4)
NS
EUROPEAN QUEEN GENOTYPE (YK)
CB1
CB2
CB4
CB5
ANALYSES5
QUEEN CELL
LENGTH
2.50
+ 0.1
(9)
2.70
+ 0.1
(9)
2.44
+ 0.1
(11)
2.57
+ 0.1
(3)
***d
QUEEN PUPAL
WEIGHT
286.4
+ 12.0
(9)
284.4
+ 20.6
(9)
272.5
+ 16.5
(13)
282.6
+ 12.0
(3)
NS
aCB = cell-producing colony number; refer to Table 5-1 for explanation.
b0ne-way analysis of variance, alpha = 0.05; *** = P<0.001.
^Mann-Whitney U test, two-tailed, alpha = 0.05.
dWith CB2 removed, ANOVA is NS.


151
density-dependent factors that are responsible for regulating queen
rearing in colonies preparing to reproduce. In European honey bees,
initiation of queen rearing prior to reproduction is not a result of a
decrease in queen pheromone production (Seeley and Fell 1981). Two
other possibilities are suggested by Seeley and Fell (1981). First,
there may be failure to adequately disperse queen pheromone in crowded
colonies prior to swarming. And, second, worker bee response to queen
pheromone may change prior to swarming.
Threshold levels for queen pheromone that inhibit queen rearing in
worker bees may be different for Africanized and European honey bees.
Also, dispersal of queen pheromone by "messenger bees (Seeley 1979) may
be different for Africanized bees compared with European bees. Baird
and Seeley (1983) developed an equilibrium model to explain the
regulation of queen rearing in colonies preparing to reproduce. Their
model postulated that "there is a balance between nurse bees becoming
inhibited from queen rearing and nurses losing their inhibition, and
that whether a colony does or does not rear queens reflects the
equilibrium percentage of inhibited nurses" (Baird and Seeley 1983, p.
221). Therefore, differences between Africanized and European honey
bees with respect to density-dependent factors regulating queen rearing
may result in differences in reproductive rates by affecting: 1) adult
population size when colonies reproduce; 2) prime swarm size; and 3)
number of afterswarms. Some of these density-dependent factors have
been compared for Africanized bees in South America with European bees
in North America under different environmental and experimental
conditions (Winston, Dropkin and Taylor 1981). Unfortunately, there are
no data collected under identical conditions that allow valid


176
Ribbands C.R. 1953. The behaviour and social life of honeybees. Bee
Research Assoc., Ltd. England.
Rinderer T.E. 1986. Africanized bees: An overview. Am. Bee J.
126:98-100, 128-129.
Rinderer T.E. A.B. Bolten^A.M. Collins and J.R. Harbo. 1984.
Nectar-foraging characteristics of Africanized and European
honeybees in the neotropics. J. Apic. Res. 23:70-79.
Rinderer T.E., A.B. Bolten, J.R. Harbo and A.M. Collins. 1982.
Hoarding behavior of European and Africanized honey bees
(Hymenoptera: Apidae). J. Econ. Ent. 75:714-715.
Rinderer, T.E. A.M. Collins, A.B. Bolten and J.R. Harbo. 1981. Size
of nest cavities selected by swarms of Africanized honeybees in
Venezuela. J. Apic. Res. 20:160-164.
Rinderer T.E. A.M. Collins and K.W. Tucker. 1985. Honey production
and underlying nectar harvesting activities of Africanized and
European honeybees. J. Apic. Res. 24:161-167.
Rinderer T.E., R.L. Hellmich II R.G. Danka and A.M Collins. 1985.
Male reproductive parasitism: A factor in the Africanization of
European honey-bee populations. Science 228:1119-1121.
Rinderer, T.E., and H.A. Sylvester. 1981. Identification of
Africanized bees. Am. Bee J. 121:512-516.
Rinderer, T.E., K.W. Tucker and A.M. Collins. 1982. Nest cavity
selection by swarms of European and Africanized honeybees. J.
Apic. Res. 21:98-103.
Roberts, W.C. 1944. Multiple mating of queen bees proved by progeny
and flight tests. Glean. Bee Cult. 72:255-259, 303.
Roberts, W.C. and S. Taber. 1965. Egg-weight variance in honey bees.
Ann. Ent. Soc. Am. 58:303-306.
Root, A.I. 1947. ABC and XYZ of bee culture. Root Co. Medina, Ohio.
Roubik, D.W. 1978. Competitive interactions between neotropical
pollinators and Africanized honey bees. Science 201:1030-1032.
Roubik, D.W. 1979. Africanized honeybees, stingless bees and the
structure of tropical plant-pollinator communities. Pages 403-417
in D. Caron (ed.), Proc. IVth Inti. Symp. on Pollination. Mise.
Publ. no. 1, Maryland Agrie. Exp. Sta. Univ. of Maryland, College
Park.
Roubik, D.W. 1980. Foraging behavior of competing Africanized
honeybees and stingless bees. Ecology 61:836-845.


139
the extent to which brood rearing ceases during resource dearths.
European honey bee populations in temperate areas have a distinct
seasonal decline in brood production and may stop brood rearing
altogether for a variable period during winter (Bodenheimer 1937;
Bodenheimer and Ben-Nerya 1937; Jeffree 1955; McLellan 1978; Nolan 1925,
1928). On the other hand, Winston reports that many Africanized
colonies in French Guiana
persist during the relative dearth season (March to June)
without the cessation of brood rearing characteristic for
temperate conditions, and are strong enough (i.e., have a
relatively high worker population and sufficient young
workers) to grow rapidly to swarming strength when resources
improve. (Winston 1980b, p. 164).
This difference between tropical and temperate conditions allows
tropical honey bee colonies to grow rapidly when resources become
available and thereby increase their potential reproductive rates
compared with temperate honey bee colonies.
More recently, investigations of both Africanized and European
honey bee populations under identical experimental conditions in
Venezuela have been undertaken. These studies have evaluated both
demographic parameters as well as resource utilization behaviors. These
investigations include the research presented in Chapters II-VII;
studies by Winston and Katz (1981, 1982); and the research by the U.S.
Department of Agriculture Bee Breeding and Stock Center Laboratory
(Collins, Rinderer, Harbo and Bolten 1982; Harbo, Bolten, Rinderer and
Collins 1981; Rinderer, Bolten, Collins and Harbo 1984; Rinderer,
Bolten, Harbo and Collins 1982; Rinderer, Collins, Bolten and Harbo
1981; Rinderer, Collins and Tucker 1985; Rinderer, Tucker and Collins
1982). Results of these investigations present quite a different


APPENDIX D
ACCURACY OF TECHNIQUE USED TO ESTIMATE NUMBERS OF BEES IN A COLONY


However those investigations have not been conducted under similar
environmental or experimental conditions. Therefore, comparisons of
reproductive rates between Africanized and European honey bees using
those data are inappropriate for either identifying differences in
reproductive rates for tropical and temperate honey bee populations or
for identifying factors responsible for the success of Africanized bees
in tropical regions.
Purpose of Mv Research
African and European honey bee populations evolved under very
different resource and climatic conditions. The presence of both
Africanized and European honey bees in Venezuela provided the
opportunity to study both populations under identical conditions in the
tropics. Differences between the two honey bee populations that make
Africanized bees more successful in tropical regions could then be
evaluated. The underlying assumption of my research was that the life
history of Africanized honey bee populations in South America (as well
as the parental population in Africa) is characterized by a high
reproductive rate. Demographic features expected to be correlated with
this high rate of colony reproduction include short worker bee
development times, small worker bee size, rapid queen development and
maturation, and increased egg laying and brood production rates.
Predictions involving these demographic characteristics led to a series
of experiments that are presented and discussed in the following
chapters.
In addition, the question of reproductive isolation versus
hybridization and differential selection between the two populations in
tropical conditions was experimentally evaluated. Whether there is


APPENDIX A
WORKER BEE DEVELOPMENT TIMES AND MORTALITY
DURING DEVELOPMENT


12
allow for normal feeding and growth. Worker bee development rates are a
result of an interaction between the egg genotype and the colony. There
are three colony-level factors that need to be considered when comparing
total development time of Africanized and European worker bees.
First is the effect of comb cell size. There is a difference in
natural comb cell size between Africanized and European populations.
The width between opposite sides of the hexagonal cells for the African
population in Africa measured 4.77-4.94 mm (Smith 1958a). Cells for the
Africanized population in Brazil averaged 5.0 mm (range 4.8-5.4 mm)
(Michener 1972), but cells of the Africanized population in Venezuela
averaged 4.8 mm (range 4.5-5.0 mm) (Chapter III; Rinderer, Tucker and
Collins 1982). Cells from the European population from Ontario, Canada,
averaged 5.4 mm (range 5.2-5.7 mm) (Michener 1972), and those from
Louisiana, U.S.A., averaged 5.2-5.3 mm (range 5.2-5.4 mm) (Rinderer,
Tucker and Collins 1982). Adult bee size is a function of comb cell
size (Grout 1937); adult Africanized bees are smaller than European bees
(62 mg compared with 93 mg, unengorged) (Otis 1982b; Otis, Winston and
Taylor 1981).
Abdellatif (1965) suggested that larvae in smaller comb cells
received less food which caused them to elongate and become sealed
earlier. Also, Tribe and Fletcher (1977) suggested that the difference
in development time for African and European genotypes may be a function
of the small African bee size. Therefore, the effect of comb cell size
needs to be considered when comparing development times of Africanized
and European honey bees.
The second colony-level factor is the effect of nurse bee genotype.
There may be behavioral differences and/or physiological differences in


82
queens were limited to paired comparisons (one Africanized and one
European line) that were each simultaneously introduced into the same
incubator-colony and then grafted into the same queen-cell-producing
colony. Possible effects from different cell-producing colonies on
queen pupal weight were evaluated by grafting the same queen lines into
different cell-producing colonies.
Only European cell-producing colonies were used because of the
difficulty in producing queens in Africanized colonies. Africanized
cell-producing colonies remained disturbed for a long period of time
after the grafted larvae were introduced which resulted in poor
acceptance (survival) of the larvae (see Chapter IV).
Queen pupal weights are used for comparison because adult weights
vary with respect to engorgment of food, dehydration, feces
accumulation, and differential ovariole development. Although queen
pupal weights vary with age of the pupae, there is a period from the
10th through the 13th day post-oviposition when queen pupal weight is
constant (Table C-l). Queen pupal weight comparisons can therefore be
made during this period (Hoopingarner and Farrar 1959). Although there
is a 0.5 day difference in development time between Africanized and
European queens (Chapter IV), the 3-4 day pupal period during which
there is no significant weight change is of sufficient duration to allow
Africanized and European queens to be accurately and consistently
compared. Africanized and European queen pupae were weighed on the 11th
day post-oviposition. Queen cells from each of the lines were randomly
selected to avoid any position effect from location on the grafting
frame. Weights were measured to the nearest 1.0 mg using either a
Mettler Type H4 or H6 balance.


179
Stort, A.C. and N. Bareli 1. 1981. Genetic study of olfactory
structures in the antennae of two Apis me!1ifera subspecies. J.
Kans. Ent. Soc. 54:352-358.
Sylvester? H.A. 1982. Electrophoretic identification of Africanized
honeybees. J. Apic. Res. 21:93-97.
Taber, S. III. 1954. The frequency of multiple mating of queen honey
bees. J. Econ. Ent. 47:995-998.
Taber, S. III. 1961. Successful shipments of honeybee semen. Bee
World 42:173-176.
Taber, S., Ill, and W.C. Roberts. 1963. Egg weight variability and its
inheritance in the honey bee. Ann. Ent. Soc. Am. 56:473-476.
Taber, S., Ill, and J. Wendel. 1958. Concerning the number of times
queen bees mate. J. Econ. Ent. 51:786-789.
Taylor, O.R. 1977. The past and possible future spread of Africanized
honeybees in the Americas. Bee World 58:19-30.
Taylor, O.R. 1985. African bees: Potential impact in the United
States. Bull. Ent. Soc. Am. 31:14-24.
Taylor, O.R. 1986. Health problems associated with African bees. Ann.
Int. Med. 104:267-268.
Taylor, O.R., R.W. Kingsolver and G.W. Otis. In press. A neutral
mating model for honey bees (Apis me!1ifera L.). J. Apic. Res.
Taylor, O.R., and M. Spivak. 1984. Climatic limits of tropical African
honeybees in the Americas. Bee World 65:38-47.
Taylor, O.R., and G.B. Williamson. 1975. Current status of the
Africanized honey bee in northern South America. Am. Bee J.
115:92-93, 98-99.
Tribe, G.D., and D.J.C. Fletcher. 1977. Rate of development of the
workers of Apis mellifera adansonii L. Pages 115-119 in D.J.C.
Fletcher (ed.), African bees: Taxonomy, biology and economic use.
Apimondia, Pretoria.
Tuenin, T.A. 1927. Variation of bees and their organs. Am. Bee J.
67:19,
Visscher, P.K., and T.D. Seeley. 1982. Foraging strategy of honeybee
colonies in a temperate deciduous forest. Ecology 63:1790-1801.
von Frisch, K. 1967. The dance language and orientation of bees.
Belknap Press, Cambridge, Mass.
Waddington, K.D. 1981. Patterns of size variation in bees and
evolution of communication systems. Evolution 35:813-814.


15
from which the Africanized queen mothers were extracted were also from
this area. The colonies were identified as Africanized honey bees by
both their behavior and their small comb cell size characteristic of the
Africanized population (4.5-5.0 mm, see Chapter III).
The queens in the colonies with European nurse bees (H2 and IBR877)
were European queens that had been mated to European drones in the
U.S.A. and transported to Venezuela. Line H2 was from a commerical
queen producer in the southeastern U.S.A.; IBR877 was an outbreed line
from the U.S. Department of Agriculture Bee Breeding and Stock Center
Laboratory in Baton Rouge, Louisiana, U.S.A.
The source for the Africanized egg genotype (A26) was a queen
removed from a feral colony of Africanized honey bees in the San Jose de
Buja area. The colony was identified as Africanized by its behavior and
characteristic comb cell size. The source of the European egg genotype
(Y5) was a queen commercially produced in the southeastern U.S.A. and
shipped to Venezuela.
Because adult longevity is from 2 to 5 weeks for European honey
bees (Woyke 1984) and 2 to 3 weeks (or less) for Africanized honey bees
(Winston 1979bj Winston and Katz 1981), experimental colonies were
established 10 weeks prior to the start of the experiment. This was
sufficient time to insure that at the beginning of the developmental
trial all the adult bees present within the experimental colonies had
developed in those colonies, and, therefore, were offspring of a known
genetic line having developed within a known comb cell size. Before
development times were measured, the worker bee populations in the
experimental colonies were equalized as much as possible by removing
random samples of bees from the most populous colonies.


TABLE 3-6. Coefficients of variation for pupal weights from
artificial, single drone inseminations and
natural, multiple matings.
COMB CELL SIZE
EGG GENOTYPE3 AFRICANIZED EUROPEAN
SINGLE DRONE INSEMINATIONS
SDA12
3.9
3.6
SDY10
3.2
2.5
SDY11
2.8
3.5
SDY1
3.6
5.1
YD28
3.3
2.5
MULTIPLE INSEMINATIONS
A26
4.4
4.5
A57
5.8
3.4
B39
3.1
5.1
WEI
3.4
2.0
ANALYSES5
NS
NS
3See Table 3-1 for explanation of genotypes.
Mann-Wh itney U test, one-tailed, alpha = 0.05


TABLE 3-2. Effect of comb cell size and egg genotype on bee pupal
weights (mg). Means + SD, (sample size).
COMB CELL SIZE
EGG GENOTYPES AFRICANIZED3 EUROPEAN5 % INCREASE
AFRICANIZED QUEEN X
AFRICANIZED DRONE
A26c
A57c
B39c
COMBINED
AFRICANIZED QUEEN X
EUROPEAN DRONE
SDA12d
EUROPEAN QUEEN X
AFRICANIZED DRONE
SDY10d
SDYIld
COMBINED
EUROPEAN QUEEN X
EUROPEAN DRONE
YD28d
WE1C
SDYld
COMBINED
105.4 +4.6 121.7 +5.5 15.5
(60) (80)
117.2+6.8 128.6+4.4 9.7
(30) (40)
116.5 +3.6 123.1 +6.3 5.7
(30) (40)
111.1+7.6 123.8+6.2 11.4
(120) (160)
112.9+4.4 122.4+4.4 8.4
(30) (40)
114.6 +3.7 133.7 +3.3 16.7
(10) (23)
115.5 +3.2 138.9 +4.9 20.2
(30) (30)
115.3 +3.3 136.7 +5.0 18.6
(40) (53)
126.8+4.2 138.7+3.5 9.4
(30) (30)
125.1 + 4.3 143.3 + 2.9 14.5
(30) (30)
115.4+4.2 135.0+6.9 17.0
(20) (19)
123.3 + 6.3 139.5 +5.4 13.1
(80) (79)
3Width between opposite sides of the hexagonal cell is 4.8 mm.
bWidth between opposite sides of the hexagonal cell is 5.4 mm.
'"Natural matings, multiple inseminations.
dSingle drone insemination.


"He must be a dull man who can examine the
exquisite structure of a comb, so beautifully
adapted to its end, without enthusiastic admiration."
Charles Darwin 1859


o r
zb 0
/;


97
larvae, approximately 15 hours old, were transferred (grafted) into
beeswax queen-cell cups that had been primed with diluted royal jelly
and then introduced into cell-producing colonies for development. Three
days prior to adult emergence, sealed queen cells were put into an
incubator (35 +1 C.). Emerged, virgin queens were marked for
individual identification, using color-coded, plastic, numbered discs
glued to the queen's thorax (Smith 1972). Virgin queens were maintained
separately in small, two-frame colonies during the period of maturation
and after artificial insemination in order to maximize the number of
spermatozoa that migrate to the spermatheca (Woyke 1979).
Queens were artificially inseminated one week after emergence using
the apparatus designed by Harbo (1979) and Mackensen (Mackensen and
Roberts 1948; Mackensen and Tucker 1970). Queens were inseminated with
2.5 ul of wild-type semen on each of two occasions, 3 days apart, to
increase the percentage of spermatozoa entering the spermatheca (Bolten
and Harbo 1982; Mackensen 1964).
Four colony treatments were established:
1. Africanized queen with Africanized worker bees,
2. Africanized queen with European worker bees,
3. European queen with Africanized worker bees and
4. European queen with European worker bees.
Each experimental colony was in a five-frame hive (23 liters) that
contained three empty combs of European cell size. A different
geometric design was painted over the entrance of each hive in order to
aid in orientation and reduce drifting of foragers between colonies (von
Frisch 1967).


72
TABLE 4-3. Analyses for the comparison of queen development
times for both Africanized and European honey bee
genotypes. Letters A V represent different
treatments; see Table 4-1 for explanation.
Kolmogorov-Smirnov one-tailed test, chi-square
distribution, df = 2, alpha = 0.05 (Siegel 1956).
A
X
B
NSa
C
X
D
NSa
E
X
F
G
X
H
***
I
X
J
**
K
X
L
**
M
X
N
**
0
X
P
*
Q
X
R
*
S
X
T
***
U
X
V
***
aSmall sample size, chi-square distribution is conservative.
b = P<0.05
** = P<0.01
*** = P<0.001.


86
The European queen lines evaluated were a diverse representation of
the European population from North America, whereas the Africanized
queen lines may only reflect a small portion of the Africanized
population. The location for the sources of the Africanized lines was
limited to feral colonies found in one area of eastern Venezuela. A
greater diversity of Africanized lines needs to be evaluated in order to
be able to generalize about queen pupal weights and oviposition rates
for the population as a whole.


84
Spearman's rank correlation coefficient was determined for queen
pupal weights and queen cell lengths (Table 5-4). In general there was
no significant correlation between queen pupal weight and queen cell
length (alpha = 0.05). However, one Africanized line (A26) in cell-
producing colony 2 had a significant correlation (P<0.05) and one
European line (YK) in cell-producing colony 4 had a significant
correlation (P<0.01).
The effect of cell-producing colonies on queen pupal weights and
queen cell lengths is presented in Table 5-5. There was no significant
difference for Africanized queen line A26 in four different cell-
producing colonies (one-way analysis of variance, alpha = 0.05); nor was
there a significant difference for Africanized queen line A57 in two
different cell-producing colonies (Mann-Whitney U test, two-tailed,
alpha = 0.05). There was no significant difference in pupal weights for
the European queen line YK in four different cell-producing colonies
(one-way analysis of variance, alpha = 0.05), but there was a
significant effect of cel 1-producing colonies on queen cell length
(P<0.001). When cell-producing colony 2 was removed from the analysis,
there was no significant difference in queen cell length.
Discussion
If we assume for both Africanized and European honey bees that
queen weight is correlated with egg production or fecundity (Boch and
Jamieson 1960), we would then expect that egg laying rates would follow
the same ranking as presented in Table 5-2 for queen pupal weights.
Based on these pupal weights, we would predict that there would be no
difference in egg laying rates for the Africanized and European honey
bee populations. In fact, when egg laying rates for Africanized and


125
introduced into the mating nuclei was 50% and 61% respectively. There
was no significant difference in acceptance (Fisher's exact probability
test, alpha = 0.05).
Discussion
Evidence for Hybridization
There appears to be no effective reproductive isolating mechanism
operating between the Africanized and European populations. The mating
of both Africanized and European queens with Africanized drones was
equally successful as judged by the number of spermatozoa in the
spermatheca. Offspring from the hybrid crosses were viable with no
apparent difference in mortality as determined by the uniformity of the
brood pattern. Kerr and Bueno (1970) report that there may be a
difference in ejaculation response between Africanized and European
drones that may provide a potential isolating mechanism. Even if this
exists, European queens were still able to successfully mate with
Africanized drones without any apparent problem, as determined by both
the numbers of spermatozoa in their spermatheca and the age when
oviposit ion began.
Although the same queen pheromone is produced by three sympatric
Asiatic species of Apis (Butler, Calam and Callow 1967; Shearer, Boch,
Morse and Laigo 1970), reproductive isolation occurs between the three
species because there is no overlap in times of drone flight (Koeniger
and Wijayagunasekera 1976). The situation between Africanized and
European populations of honey bees is quite different with respect to
the time of flight of the queens and drones. Data from Venezuela show
that flight times for Africanized and European drones completely overlap
during the approximately three hours of mating flight activity with only


14
between the effects of egg genotype and the colony-level parameters of
comb cell size and nurse bee genotype on worker bee development time.
These experiments were conducted during July-October 1979.
Methods.
Table 2-1 summarizes the experimental design. Four experimental
colony treatments were established as follows:
i.Africanized comb cell size Africanized nurse bees (A55)
ii.Africanized comb cell size European nurse bees (H2)
iii.European comb cell size, Africanized nurse bees (A41)
iv.European comb cell size, European nurse bees (IBR877).
Each experimental colony was a five-frame hive (22 liters) with
four empty combs and one comb with honey and pollen and approximately 2
kg of young adult bees (Africanized or European, depending upon
treatment). Because natural nectar and pollen resources were available
irregularly throughout the experimental period (16 weeks), the colonies
were supplemented with honey and pollen as necessary.
European comb was built from commercially-produced beeswax
foundation that had been fastened into standard wooden frames.
Africanized comb was naturally built (not from foundation) by
Africanized bees in empty standard wooden frames to facilitate
manipulation and colony inspection.
The queens in colonies with Africanized nurse bees (A55 and A41)
were Africanized queens produced by standard queen rearing methods
(Laidlaw 1979) and then naturally mated to Africanized drones. Mating
occurred in an area of eastern Venezuela that had a large feral
population of Africanized honey bees with no known European honey bees
present (near San Jose de Buja, Monagas, Venezuela). The feral colonies


105
European colonies a growth rate advantage. Unfortunately, brood
mortality for both Africanized and European honey bees during the
initial colony growth phase has not been investigated under identical
conditions.
Based on the results from these studies and those evaluating other
colony demographic parameters, e.g., worker development times (Chapter
II) and queen maturation rates (Chapter IV), it must be concluded that
differences in reproductive rates between Africanized and European honey
bees in South America cannot be attributed to intrinsic demographic
factors. Reproductive rates in honey bees are a function of at least
two other factorsresource availability and resource utilization
efficiency. Chapter VIII presents a hypothesis to explain the success
of Africanized honey bees based on differences in resource utilization
efficiency. This hypothesis is based on differences between Africanized
and European bees with respect to foraging behavior, brood production
efficiency as a function of bee size, and resource-induced absconding.


145
flowers, Nunez (1973, 1979a, 1982) analyzed foraging behaviors of
Africanized and European honey bees. Differences between the two
populations were observed that were appropriate to having evolved under
either temperate or tropical resource availability patterns. Hoarding
cage studies also demonstrated differences in response between
Africanized and European honey bees, suggesting differences in resource
utilization behavior (Rinderer, Bolten, Harbo and Collins 1982).
Increased numbers of bees in a colony are important for successful
foraging, colony defense and reproduction (see Wilson 1971). When
floral resources (nectar and pollen) are limited, a greater number of
individual bees can be produced from a given amount of food if brood
production is more efficient and/or if bees are smaller. There are no
data available that were collected under identical conditions that allow
comparison of brood production efficiency between Africanized and
European honey bees, as measured by the ratio of developing brood to
adult population for a range of different adult populations (Michener
1964; Moeller 1961). However, their smaller size may result in
increased brood production efficiency in Africanized bees, i.e., less
food is necessary to produce smaller bees (Chapter III; Fletcher and
Tribe 1977a; Tribe and Fletcher 1977). Therefore, with a limited food
supply, Africanized honey bees could increase their population at a
greater rate than European honey bees. With more successful foraging
behavior and smaller bee size, Africanized colonies can grow rapidly
under conditions where European colonies may not be able to survive.
Another important difference between Africanized and European honey
bees is the strategy used to survive during periods of food shortage.
Honey bee colonies may either hoard sufficient quantities of food


153
The Africanized honey bees studied in Venezuela are only a small
sample of the total Africanized honey bee population in South and
Central America and represent only a fraction of the variation within
the population, particularly if we consider that Africanized honey bees
are a result of hybridization. Nevertheless, the results presented in
the foregoing chapters demonstrate that at least some portion of the
Africanized honey bee population is similar to the European honey bee
population with respect to the demographic parameters analyzed.
Potential Impact of Africanized Honev Bees in North America
The selective advantage of Africanized honey bees in South America
will be lost as they disperse north into temperate regions. European
honey bees will have the selective advantage in temperate regions
because of their particular behavioral repertoire which is better
adapted to temperate conditions. However, because the populations can
interbreed successfully, negative characteristics of the Africanized
population, e.g., their stinging behavior, may become genetically
introgressed into the European population of North America and therefore
widespread wherever honey bees can survive (Chapter VII). A more
optimistic scenario is that the large population of European honey bees
in Mexico will slow the spread of African genes because of competition
for available resources as well as through hybridization. Therefore,
through selection, hybridization, and competition, the impact of
Africanized honey bees may be minimized in North America (Chapter VII).


66
cells were introduced into the colonies, resulting in poor survival or
acceptance of the grafted larvae (5-50% for Africanized colonies,
compared with 35-95% for European colonies). In addition, Africanized
colonies were difficult to manage because of excessive stinging that
occurred when manipulating the strong colonies that were necessary for
proper queen production.
Page and Erickson (1984) found evidence that nurse bee colonies
preferentially raised queens from more closely related larvae. However,
in the present study, no evidence for kin recognition was observed.
Africanized and European nurse bee colonies reared Africanized and
European queens with equal frequency (Table 4-2).
Rate of Maturation
Attractiveness of queens to drones is a function of the amount of
pheromone (9-oxodec-trans-2-enoic acid) produced in the mandibular
glands of the queens (Butler 1971; Boch, Shearer and Young 1975). In
England, using European genotypes, Butler (1971) tethered virgin queens
of various ages 6 meters above the ground in areas where drones were
flying. He determined that queens younger than 5 to 6 days old seldom
elicited a positive drone response. Maximum positive responses from
drones were observed in queens 8 or more days old. Butler's results
differ from those presented in this study and may be attributed to
either differences in experimental conditions or genetic differences
between the queen lines studied rather than to differences due to any
tropical or temperate conditions. The response of drones to queens
reported in this chapter was evaluated in a drone congregation area
which may account for the differences between the studies.


76
TABLE 4-7. Unsealed (egg and larval periods combined) development times
(in days) for Africanized and European queens: median,
(sample size).
EGG GENOTYPES
NURSE BEE GENOTYPE3
AFRICANIZED (A26)
EUROPEAN (Y5)
EFFECT OF EGG
GENOTYPE5
19
7.5 (17)
7.5 (19)
*
28
7.2 (14)
7.5 (12)
*
19 & 28
7.5 (31)
7.5 (31)
*
EFFECT OF NURSE BEE
GENOTYPE0
NS
NS
^Cell-producing colonies, European nurse bees, European comb cell size.
bKolmogorov-Smirnov one-tailed test, chi-square distribution, df = 2,
* = P<0.05.
cKolmogorov-Smirnov two-tailed test, chi-square distribution, df = 2,
alpha = 0.05.


83
Queen cell lengths were measured at the time the queen pupal
weights were determined. A calipers was used to determine the external
length from the base to the apex of the queen cell.
Results *
Queen pupal weights for four Africanized and five European lines
are presented in Table 5-1. European queen pupal weights were
significantly larger than Africanized queen pupal weights for two
different pairwise comparisons (YK vs. A26 and YK vs. A57; P<0.05 to
P<0.001; Mann-Whitney U test one-tailed). Africanized queen pupal
weights were significantly larger in one pairwise comparison (A62 vs. N;
P<0.02; Mann-Whitney U test two-tailed). For three pairwise
comparisons there was no statistical difference (A26 vs. WE, A57 vs.
YD, and A61 vs. GK; Mann-Whitney U test, one-tailed, alpha = 0.05).
Because different cell-producing colonies had no significant effect on
queen pupal weights (see below), the means for the nine queen lines can
be ranked and analyzed (Table 5-2). There was no significant difference
between the Africanized and European honey bee populations for queen
pupal size (Mann-Whitney U test, one-tailed, alpha = 0.05).
Queen cell lengths for Africanized and European lines are presented
in Table 5-3. In six out of eight pairwise comparisons, there was no
significant difference in queen cell lengths between Africanized and
European queens (Mann-Whitney U test, one-tailed, alpha = 0.05). For
the pair in cell-producing colony 2, the European line was significantly
larger than the Africanized line (P<0.05). For the pair in cell-
producing colony 4, the Africanized line was significantly larger than
the European line (P<0.02; Mann-Whitney U test, two-tailed).


BIOLOGY OF AFRICANIZED AND EUROPEAN HONEY BEES,
Apis mail ifera, IN VENEZUELA
By
ALAN B. BOLTEN
A DISSERTATION PRESENTED TO THE GRADUATE SCHOOL
OF THE UNIVERSITY OF FLORIDA IN
PARTIAL FULFILLMENT OF THE REQUIREMENTS
FOR THE DEGREE OF DOCTOR OF PHILOSOPHY
UNIVERSITY OF FLORIDA


19
Africanized worker bees developed faster than European bees (ACEG x
BDFH). The unsealed larval period was 4.3 +0.4 days compared with 4.9
+0.4 days, PcO.OOl; the sealed larval and pupal period was 11.6 +0.5
days compared with 11.9 + 0.4 days, P<0.01; and the total development
time was 18.9 + 0.3 days compared with 19.8 + 0.4 days, P<0.001. There
was no significant effect of comb cell size or nurse bee genotype on
development times. These differences in total development time are
similar to the differences found between three different lines of
Africanized and three different lines of European honey bees compared in
another study (19.2 days compared with 20.0 days, Table A1).
Comb cell sizes for the experimental colonies and egg sample frames
are presented in Table 2-7. Temperatures recorded for all experimental
colonies varied from 35-36C.
When differences in development times between Africanized and
European egg genotypes were compared for each stage of development, the
greatest difference was observed in the unsealed larval stage (Table 2-
8). However, this was not a result of a differential acceleration of
development during the unsealed larval period for the Africanized honey
bees. The proportion of unsealed larval development time to total
development time and the proportion of sealed brood development time to
total development time were compared for the Africanized and European
honey bee populations following angular transformations of the
proportions (Sokal and Rohlf 1969). These proportions were not
significantly different between the Africanized and European honey bees.
The differences recorded for the unsealed brood stage between the
Africanized and European honey bees may be an artifact of the experiment
for two reasons. First, the 24-hour observation interval may obscure


161
TABLE B-3. Comparison of Africanized and European comb cell diameter
and comb cell volume: mean + SD, range, CV, (sample size).
DIAMETER3 VOLUME5
(mm) (ml x 10-3) CORRELATIONS0
AFRICANIZED COMB CELLS 4.8 +0.1
4.6 4.9
CV =2.1
(50)
184.6 + 15.8
160 215
CV = 8.6
(50)
EUROPEAN COMB CELLS
5.4 + 0.05
5.4 5.5
CV = 1.0
(30)
264.3 + 23.5
225 300
CV = 8.9
(30)
NS
NSd
ANALYSES
P<0.001 P<0.001
determined by measuring 10 horizontal, consecutive cells; cell-wall
thickness not considered.
5Cell volume determined by filling cells with water with a pipette.
cSpearman's rank correlation coefficient, alpha = 0.05.
dNegative correlation, P<0.01.
et-test, one-tailed.


64
different as a result of the distribution around the median (P<0.05,
Kolmogorov-Smirnov one-tailed test* chi-square distribution, df = 2).
Development of Attractiveness to Drones
The response of drones to tethered, virgin queens is summarized in
Table 4-8. There were no differences between Africanized and European
queens with respect to either the earliest age at which a positive drone
response (Rank 1) was observed or the earliest age at which a maximum
drone response (Rank 4) was observed. Both Africanized and European
virgin queens were able to attract drones (Rank 1) on the day they
emerged. Africanized virgin queens can maximally attract drones (Rank
4) by the fourth day post-emergence; European virgin queens can elicit a
Rank 4 response by the fifth day post-emergence. This difference was
not significant (Kolmogorov-Smirnov one-tailed test, chi-square
distribution, df = 2, alpha = 0.05). The data in Table 4-8 have been
combined for the two Africanized and two European queen lines. However,
the queens within a population (Africanized or European) or within a
line within a population were not uniform with respect to drone response
or the rate of maturation. There were differences between the two
Africanized lines and between individuals within the same line for the
earliest age for a Rank 4 response. This same variation between lines
and within lines existed for the European population.
Time Post-Emergence to Initiation of Ovinos it ion
Table 4-9 presents the data for time post-emergence to the
initiation of oviposition for both Africanized and European queens.
Africanized queens began oviposition at 8.5 days post-emergence whereas
European queens begin at 7.5 days (P<0.05, Kolmogorov-Smirnov two-tailed
test, chi-square distribution, df = 2).


Qj-j^pyfrp jjj
INTERACTION OF MATERNAL GENOTYPE, EGG GENOTYPE AND COMB CELL SIZE ON
HONEY BEE WORKER SIZE AND SIZE VARIATION
Introduction
In the evolution of eusociality in bees (Apoidea), there is a
considerable decrease in size variation of the workers within a colony.
Worker size variation within a colony of primitively eusocial sweat bees
(Halictidae) or bumble bees (Apidae) is much greater than the size
variation of workers within colonies of highly eusocial stingless bees
(Mel iponinae: Mel ipona and Trigonal or honey bees (Apinae: Apis)
(Brian 1952; Kerr and Hebling 1964; Medler 1965; Michener 1974). For
example, the coefficient of variation (CV) for worker weights in a
bumble bee colony may be as high as 31-37% (calculated from Brian 1952
for Bombus aarorum. Table B1) whereas the CV for worker weights within
a honey bee (Apis me!1ifera) colony is only 4-7% (Table B-2).
An effect of the reduction of size variation is that the mechanism
for the division of labor of workers within a colony shifts from being
size dependent to primarily age dependent (Michener 1974). In the
primitively eusocial bumble bees, division of labor is size related
(Brian 1952); large workers may be twice the size (linear measurements)
of small workers within the same colony (Medler 1965). In highly
eusocial stingless bees and honey bees, division of labor is primarily
age dependent (Free 1965; Gary 1975; Kerr and Hebling 1964; Lindauer
1953; Seeley 1982). In honey bees, the workers proceed through a series
35


65
Discuss ion
Queen Development
Development time from oviposition to emergence for Africanized
queens in this study was 14.5 days, which is the same as the development
period reported for both Africanized bees in French Guiana (Winston
1979c) and their parental population, A. m. adanson i i (=scutellata) in
South Africa (Anderson, Buys and Johannsmeier no date; Fletcher 1978;
Fletcher and Tribe 1977c). The European queens in the present study
developed in 15.0 days, which is about one day shorter than expected
from previous reports (Jay 1963; Laidlaw 1979). Therefore, the
difference in development times for Africanized and European queens was
not as great as expected and underscores the importance of making
comparisons under the same experimental conditions.
Queens have approximately a 25% shorter development period than
worker bees. Differences in total development times between queens and
worker bees are primarily due to a much shorter sealed development
stage, i.e., 7.5 days compared with 12 days for European bees and 7.0
days compared with 12 days for Africanized bees. The sealed development
stage in worker bees is approximately 60% of the total development time,
whereas in queens it is approximately 50%.
European queens took 3.4-5.6% longer than Africanized queens to
develop. This difference is similar to the 5.3% difference in
development times between European and Africanized worker bees from the
same two egg sources (A26 and Y5) (see Chapter II).
There was no effect of nurse bee genotype on queen development
times. However, queens were produced more successfully in European
colonies. Africanized nurse bees were easily disturbed when the grafted


23
problems with their analysis. First as already pointed out using the
duration of the unsealed stage has inherent problems because it is not a
precise development stage. Second comparisons based on data collected
under different experimental conditions are not valid. Third, their
logic is perhaps circular with respect to the question of larval size
and larval development times. In the present study, development time
was not size-related for either Africanized or European honey bees.
Africanized honey bees that developed in European comb had the same
development times as those that developed in Africanized comb even
though Africanized bees reared in European comb were significantly
larger (16%; Chapter III). The same relationship was true for European
honey bees with a 17% increase in size of bees from European comb
compared with bees from Africanized comb. And fourth, their comparison
is in itself incorrect. Rather than compare the differences in unsealed
development times between African and European populations to determine
if the African population has a relatively shorter duration as unsealed
larvae, they should have used the proportion of unsealed development
period to total development period in order to compare African and
European populations. In the present study, the relative times spent as
an unsealed larvae to the total development time for both the
Africanized and European genotypes were not significantly different.
The differences in development time between Africanized and European
populations appear constant throughout development without any
developmental acceleration during the larval stage for either
Africanized or European honey bees.


CHAPTER VII
SUCCESSFUL HYBRIDIZATION BETWEEN AFRICANIZED AND EUROPEAN HONEY BEES
IN VENEZUELA WITH IMPLICATIONS FOR NORTH AMERICA
Introduction
In 1956 African honey bees, Apis mel1 ifera scutel1 ata, formerly
classified as A. m. adansonii (Ruttner 1976a, 1976b, 1981), were
imported into southeastern Brazil (Kerr 1967). Their hybridized
descendents, known as Africanized honey bees (Goncalves 1982), have
rapidly spread throughout tropical South and Central America as far
north as Honduras and El Salvador (Rinderer 1986). The dispersion from
their original importation site into new areas has been rapid200-500
km per year (Taylor 1977, 1985; Winston 1979a). As Africanized honey
bees have dispersed into new areas, they have rapidly increased in
number (Otis 1982a) and have attained dramatic population densities
(Michener 1975): 4-8 colonies/km^ (Taylor 1985), or as high as 107.5
colonies/km in the cerrado habitats in the Brazilian states of Goias
and Mato Grosso (Kerr 1971 cited in Michener 1975). There are now
probably more than ten million feral colonies of Africanized honey bees
in South and Central America (Winston, Taylor and Otis 1983). Their
success in these habitats, compared with European populations of honey
bees, may be attributed to their foraging behavior which is more suited
to the resource patterns of the tropics (Nunez 1973, 1979a, 1982;
Rinderer, Bolten, Collins and Harbo 1984; Rinderer, Collins and Tucker
1985; Winston and Katz 1982). As a result of both their foraging
118


TABLE 4-8. Drone response to tethered virgin queens: median
day of response post-emergence, range, (sample
size).
DAY OF RESPONSE LEVEL
RANK 1
RANK 2
RANK 3
RANK 4
AFRICANIZED QUEEN
0a
3.5
3.5
4.0
GENOTYPES

1-5
1-5
1-5
(2)
(6)
(6)
(5)
EUROPEAN QUEEN
0a
1.5
4.0
4.5
GENOTYPES

1-5
2-5
4-5
(1)
(4)
(3)
(2)
ANALYSES6
NS
NS
NS
aDay 0 = day of adult emergence.
bKolmogorov-Smirnov one-tailed test, chi-square distribution,
df = 2, alpha = 0.05.


100
were now introduced into European colonies; queens that had been in
European colonies during the first experiment were now introduced into
Africanized colonies.
After day 12 of oviposition, each experimental colony was inspected
and the amounts of unsealed brood (eggs and larvae) and sealed brood
(pre-pupae and pupae) for each colony were determined. To facilitate
measuring the amounts of unsealed and sealed brood, a 2.5 x 2.5 cm grid
was placed over each frame and the amount of brood within each square
was estimated. The number of developing brood cells was determined by
multiplying the brood area (in cm^) by 4.25 (the number of comb cells
O
per crrr). The number of worker bees present at day 12 was estimated by
assuming a constant rate of mortality of adults from day 0 to day 17.
At the end of day 17 of oviposition, colonies were closed and
killed with potassium cyanide. Unsealed and sealed brood, numbers of
adult bees, amount of pollen, and frame weights were determined.
The number of adult bees present at day 17 was estimated by
determining the mean individual bee weight from three samples of 150-200
bees taken from each colony. The total weight of bees in each colony
was then divided by the mean individual weight to get an estimate of the
total number of bees in each colony (Otis 1982b). The accuracy of this
technique was determined by comparing the estimates with the actual
counts (Table D1). A mean difference of only 1.5% was observed.
Data were analyzed using the Mann-Whitney U test (alpha = 0.05).
Correlations were evaluated using Spearman rank correlation coefficients
(alpha = 0.05).


93
not possible to determine to what extent reproductive differences
between the two populations are a result of differences in intrinsic
demographic parameters and/or environmental differences due to temperate
vs. tropical resources and climatic conditions. In addition,
experimental conditions were very different. For example, an important
variable affecting reproductive rates in honey bees is brood-nest
crowding (Baird and Seeley 1983; Simpson 1966, 1973; Simpson and Riedel
1963). Experimental Africanized colonies in South America were
maintained in 22-liter hives (Otis 1980; Winston 1979b), whereas,
experimental European colonies in North America, with which they were
compared, were maintained in 42-liter hives (Winston 1980a).
Reproductive rates in honey bees are a result of an interaction of
at least three factors: colony demography, resource availability, and
resource utilization efficiency. As part of a larger investigation
comparing intrinsic demographic factors between Africanized and European
honey bees to determine which aspects of demography, if any, are
responsible for the success of Africanized honey bees in South America,
this study evaluated one aspect of demographyqueen fecundity. Queen
egg laying rate is one of the primary demographic parameters that
affects colony growth rates (Brian 1965; Moeller 1961; Wilson 1971).
Although differences in egg laying rates between Africanized queens and
European queens have been reported (Fletcher 1978; Michener 1972, 1975;
Ribbands 1953), they cannot be compared because the data were collected
under different resource and experimental conditions. Therefore, the
present study was undertaken to compare egg laying and brood production
rates for both Africanized and European queens under identical, tropical
conditions in Venezuela. The experimental design allowed for a


94
comparison between the two honey bee populations during the first brood
cycle. Differences in initial colony growth rates between Africanized
and European honey bees may be an important factor in determining
differences in reproductive rates.
Egg laying and brood production rates for Africanized and European
honey bees were evaluated at both the queen and worker bee levels. The
interactions of both Africanized and European queens with both
Africanized and European worker bees were evaluated because of potential
behavioral and/or physiological differences between Africanized and
European nurse bees with respect to affecting brood production rates
and/or the queen's oviposition behavior. In order to compare egg laying
and brood production rates between Africanized and European honey bees,
four variables needed to be controlled.
First is colony size, because egg laying rates are positively
correlated with the number of worker bees in a colony (Moeller 1958).
In order to evaluate initial colony growth, a colony size was selected
that contained the number of worker bees within the range reported for
both Africanized and European swarms (Fell et. al. 1977; Otis 1980;
Rinderer, Collins, Bolten and Harbo 1981; Rinderer, Tucker and Collins
1982; Winston 1980a).
Second is hive cavity volume, which must be controlled in order to
avoid effects of differential brood-nest crowding on oviposition rates
(Brian 1965). A hive cavity volume was selected that represents natural
nest cavity volumes chosen by these two honey bee populations (Rinderer,
Collins, Bolten and Harbo 1981; Rinderer, Tucker and Collins 1982;
Seeley 1977; Seeley and Morse 1976; Winston, Taylor and Otis 1983).


85
European honey bee queens were compared there was no significant
difference between queens from the two populations (Chapter VI). There
were, however, significant differences in pupal weights between
individual queen lines both between and within each population (Table 5-
1). There were also significant differences in egg laying rates between
individual queen lines both between and within the two populations
(Chapter VI).
In this study, Africanized queens were reared in European colonies
because of low acceptance (survival) of grafted cells in Africanized
colonies (Chapter IV). Because queen-worker caste differentiation in
honey bees is regulated by larval feeding (Beetsma 1979), rearing
Africanized queens in European colonies may have obscured differences in
pupal weights between Africanized and European queens. Possibly,
European worker bees may rear larger Africanized queens than would
Africanized worker bees because European worker bees are themselves
larger (Chapter III), and may feed the developing queen larvae
differently. Although virgin European queens have been reported to
weigh more than virgin Africanized queens208 vs. 199 mg (Goncalves,
Kerr and Nocoes 1972 cited in Michener 1975)there was no indication of
conditions under which the queens were reared.
Further analysis of queen weights between Africanized and European
honey bees is needed, preferably in a 2 x 2 experimental design:
Africanized and European queens reared in both Africanized and European
cell-producing colonies. In addition, the relationship between queen
pupal weights and brood production needs to be evaluated for both
Africanized and European honey bee lines to determine if the same
relationship exists for both populations.


170
Dietz, A. 1978. An anatomical character suitable for separating drone
honey bees of Apis mellifera 1 iqustica from Apis me11 ifera
adansoni1. Pages 102-106 in Apicultura em clima quente.
Apimondia, Florianopolis, Brazil.
Dietz, A., R. Krell and F.A. Eischen. 1985. Preliminary investigation
on the distribution of Africanized honey bees in Argentina.
Apidologie 16:99-108.
DuPraw, E.J. 1965. Non-Linnean taxonomy and the systematics of honey
bees. Syst. Zool. 14:1-24.
Eckert, J.E. 1934. Studies on the number of ovarioles in queen
honeybees in relation to body size. J. Econ. Ent. 27:629-635.
Ewel, J.J., and A. Madriz. 1968. Zonas de vida de Venezuela.
Ministerio de Agricultura y Cria, Caracas.
Fell, R.D., J.T. Ambrose, D.M. Burgett, D. De Jong, R.A. Morse and T.D.
Seeley. 1977. The seasonal cycle of swarming in honeybees. J.
Apic. Res. 16:170-173.
Fletcher, D.J.C. 1975. New perspectives in the causes of absconding in
the African bee (Apis mel1 ifera adansonii L.), Part I. S. Afr. Bee
J. 47:11-14.
Fletcher, D.J.C. 1976. New perspectives in the causes of absconding in
the African bee (Apis mellifera adansoni i L.), Part II. S. Afr.
Bee J. 48:6-9.
Fletcher, D.J.C. 1977a. A preliminary analysis of rapid colony
development in Apis mellifera adansonii. Pages 144-145 in Proc.
Eighth Int. Congress, Int. Union for Study of Social Insects.
Wageningen, Netherlands.
Fletcher, D.J.C. 1977b. Evaluation of introductions of European honey
bees into southern and eastern Africa. Pages 146-147 in Proc.
Eighth Int. Congress, Int. Union for Study of Social Insects.
Wageningen, Netherlands.
Fletcher, D.J.C. 1978. The African bee, Apis mellifera adansonii, in
Africa. Ann. Rev. Ent. 23:151-171.
Fletcher, D.J.C., and G.D. Tribe. 1977a. Swarming potential of the
African bee, Apis mellifera adansonii L. Pages 25-34 in D.J.C.
Fletcher (ed.) African bees: Taxonomy, biology and economic use.
Apimondia, Pretoria.
Fletcher, D.J.C., and G.D. Tribe. 1977b. Natural emergency queen
rearing by the African bee gl. adansonii and its relevance for
successful queen production by beekeepers, II. Pages 161-168 in
D.J.C. Fletcher (ed.), African bees: Taxonomy, biology and
economic use. Apimondia, Pretoria.


44
when comb cell size variation increased 300%, bee size variation
increased only 50%.
The genetic variation of worker bees within a colony is great
because queens mate on the average with as many as 17 drones (Adams,
Rothman, Kerr and Paulino 1977). There is some degree of mixing of
spermatozoa in the spermatheca resulting in spermatozoa from at least 5
to 6 drones being used during one time interval (Page and Metcalf 1982).
Evidence that maternal inheritance reduces size variation in genetically
diverse worker offspring can be demonstrated by evaluating the size
variation of offspring from single-drone and multiple-drone inseminated
queens. The progeny of queens that were inseminated by spermatozoa from
single drones (SDA12, SDY10, SDY11, SDY1 and YD28) were expected to be
less variable than multiply-inseminated queens (A26, A57, B39 and WEI)
because all eggs from the former queens would have been fertilized by a
genetically identical male gamete. (Drones are haploid; all spermatozoa
are produced by mitosis and are therefore genetically identical.)
Evaluating the coefficient of variation (CV) for each treatment of
genotype and comb cell size, there is no difference between the
variation of progeny from single-drone inseminations versus those from
multiple inseminations, as shown in Table 3-6 (Mann-Whitney U test, one-
tailed, alpha = 0.05).
Additional evidence of maternal inheritance reducing size variation
in genetically heterogeneous offspring comes from analyzing the results
of the reciprocal cross. Because of the influence of maternal
inheritance, subspecific differences in size between Africanized and
European populations were not reflected in increased size variation of
the hybrids compared with the parental types (Table 3-4).


I certify that I have read this study and that in my opinion it
conforms to acceptable standards of scholarly presentation and is fully
adequate, in scope and quality, as a dissertation for the degree of
Doctor of Philosophy.
1
Thomas C. Emmel, Chairman
Professor of Zoology
I certify that I have read this study and that in my opinion it
conforms to acceptable standards of scholarly presentation and is fully
adequate, in scope and quality, as a dissertation for the degree of
Doctor of Philosophy.
latlran Reiskind
Associate Professor of Zoology
I certify that I
conforms to acceptable
adequate, in scope and
Doctor of Philosophy.
have read this study and that in my opinion it
standards of scholarly presentation and is fully
quality, as a dissertation for the degree of
Malcolm T. Sanford
Associate Professoryof Entomology
and Nematology
This dissertation was submitted to the Graduate Faculty of the
Department of Zoology in the College of Liberal Arts and Sciences and to
the Graduate School and was accepted as partial fulfillment of the
requirements for the degree of Doctor of Philosophy.
August 1986
Dean, Graduate School


73
TABLE 4-4. Analyses of queen development times for the
Africanized egg genotype in the different cell-
producing colonies. Letters represent different
cell-producing colonies; see Table 4-1 for
explanation. Kolmogorov-Smirnov two-tailed test
chi-square distribution df = 2 alpha = 0.05
(Siegel 1956).
A x C
NS
A x G
NS
A x I
NS
A x K
NS
A x M
NS
A x 0
NS
A x Q
NS
C x G
NS
C x I
NS
C x K
NS
C x M
NS
C x 0
NS
C x Q
NS
G x I
NS
G x K
NS
G x M
NS
G x 0
NS
G x Q
NS
I x K
NS
I x M
NS
I x 0
NS
I x Q
NS
K x M
NS
K x 0
NS
K x Q
NS
M x 0
NS
M x Q
NS
0 x Q
NS


VQUEEN PUPAL WEIGHTS 79
Introduction ...... . 79
Methods 80
Results 83
Discussion ....... ...... 84
VIEGG LAYING AND BROOD PRODUCTION RATES
DURING THE FIRST BROOD CYCLE ...... 92
Introduction 92
Methods 96
Results 101
Discussion ...... 102
VIISUCCESSFUL HYBRIDIZATION BETWEEN AFRICANIZED
AND EUROPEAN HONEY BEES IN VENEZUELA WITH
IMPLICATIONS FOR NORTH AMERICA 118
Introduction 118
Methods 123
Results 124
Discussion 125
VIIIDISCUSSION: FACTORS CONTRIBUTING TO THE SELECTION
ADVANTAGE OF AFRICANIZED HONEY BEES IN SOUTH AMERICA
THE RESOURCE UTILIZATION EFFICIENCY HYPOTHESIS 133
Success of Introduced Populations of Honey Bees 133
Factors Affecting Honey Bee Reproductive Rates ....... 135
Factors Contributing to the Selective Advantage
of Africanized Honey Bees in South America 140
Potential Impact of Africanized Honey Bees
in North America 153
APPENDICES
A WORKER BEE DEVELOPMENT TIMES AND MORTALITY
DURING DEVELOPMENT .......... 156
B HONEY BEE SIZE, COMB CELL SIZE AND
SIZE VARIATION 159
C CHANGES IN QUEEN PUPAL WEIGHT WITH AGE ...... 165
D ACCURACY OF TECHNIQUE USED TO ESTIMATE
NUMBER OF BEES IN A COLONY 167
LITERATURE CITED ... ......... 168
BIOGRAPHICAL SKETCH .... ........ 182
vi


TABLE A-l
. Comparison of worker bee development time (in days) for
Africanized and European honey bees: median, (range), mean
+ SD, (n = sample size). All development measured in
European comb cell size with European nurse bees.
EGG
GENOTYPES UNSEALED BRtfOD SEALED BROOD
TOTAL
DEVELOPMENT3
AFRICANIZED
A53
(n = 25)
5.0
(4-5)
4.8 + 0.4
11.0
(11-12)
11.4 + 0.5
19.0
(19-20)
19.2 + 0.4
A26
(n = 9)
5.0
(5)
5.0 + 0
11.0
(11-12)
11.3 + 0.5
19.0
(19-20)
19.3 + 0.5
A25
(n = 19)
5.0
(4-5)
4.6 + 0.5
11.0
(11-12)
11.4 + 0.5
19.0
(19)
19.0 + 0
COMBINED
(n = 53)
5.0
(4-5)
4.8 + 0.4
11.0
(11-12)
11.4 + 0.5
19.0
(19-20)
19.2 + 0.4
EUROPEAN
W18
(n = 28)
5.0
(4-5)
4.8 + 0.4
12.0
(11-13)
12.0 + 0.3
20.0
(19-20)
19.8 + 0.4
HI
(n = 19)
5.0
(5-6)
5.4 + 0.5
12.0
(11-13)
12.1 + 0.4
20.0
(20-21)
20.5 + 0.5
Y(A5)
(n = 26)
5.0
(4-5)
4.8 + 0.4
12.0
(11-12)
12.0 + 0.2
20.0
(19-20)
19.8 + 0.4
COMBINED
(n = 73)
5.0
(4-6)
5.0 + 0.5
12.0
(11-13)
12.0 + 0.3
20.0
(19-21)
20.0 + 0.5
ANALYSES5
NS
P<0.001
P<0.001
3Total development = time from oviposit ion to adult emergence.
bKolmogorov-Smirnov one-tailed test, chi-square distribution, df = 2,
alpha = 0.05 (Siegel 1956). Combined samples used for analyses.
156


30
TABLE 2-5. Sealed brood development times. Hypotheses were tested
using Kolmogorov-Smirnov one-tailed test chi-square
distribution, df = 2, alpha = 0.05 (Siegel 1956).
HI: Worker bee development is faster for Africanized genotypes than for
European genotypes.
A X B NS
C X D *a
E X F NS
G X H NS
A X H NS
AC X BD *
EG X FH *
AE X BF *
CG X DH **
ACEG X BDF **
H2: Worker bee development is more rapid in Africanized comb cells than
in European comb cells.
AXE NS
C X G NS
B X F NS
D X H NS
AC X EG NS
BD X FH NS
H3: Worker bee development is more rapid with Africanized nurse bees
than with European nurse bees
A X C NS
E X G NS
B X D NS
F X H NS
AE X CG NS
BF X DH NS
H4: Worker bee development is more rapid with Africanized comb cells
and Africanized nurse bees than with European comb cells and
European nurse bees.
A X G NS
B X H NS
a = P<0.05
** = P<0.01.


53
TABLE 3-5. Maternal effect: hypotheses and analyses (Mann-
Whitney U test, one-tailed, alpha = 0.05).
Letters refer to experimental treatments, see
Table 3-4. The analyses of the following
hypotheses (a postiori) demonstrate that the pupal
weights of hybrids from a reciprocal cross are
different from each other (H4) but are the same as
their respective queen mothers (H2 and H6).
HI:
B
<
R
***a
H2:
B
<
H
NS
H3:
B
<
J
***
B
<
L
***
H4:
H
<
J
***
H
<
L
***
H5:
H
<
R
***
H6:
J
<
R
NS
L
<
R
NS
a*** = P<0.001.


8
All Africanized and European honey bee comparisons were made at the
same time under identical experimental conditions. Field and
experimental methods are described for each of the experiments in the
appropriate chapters.
A few experiments with European honey bees only were conducted in
the U.S.A. to confirm techniques developed and used while in Venezuela.
These experiments were undertaken either at the U.S. Department of
Agriculture Bee Breeding and Stock Center Laboratory in Baton Rouge,
Louisiana, or at the bee research facilities of the Institute of Food
and Agricultural Sciences at the University of Florida, Gainesville.


57
2) post-emergence development of queen attractiveness to drones; and 3)
time from adult emergence to initiation of oviposit ion.
In the studies reported here, queen development and maturation were
evaluated under controlled conditions. Total development time is
defined as the time from oviposition to adult emergence. These
experimental conditions avoid the problems of previous studies that
evaluated queen development and maturation in colonies that were
swarming (e.g., Otis 1980). Under natural swarming conditions, queens
are very often confined within their cells by worker bees and prevented
from emerging for 1-10 days after pupal eclosin (Otis 1980).
Confinement makes calculations of development times difficult, and,
because maturation proceeds during confinement, maturation time
calculated from emergence to beginning of oviposition would be under
estimated.
Methods
Queen Development Times
The Africanized egg source (A26) and the Africanized cell-producing
colonies (A37 and A43) were established from queens removed from feral
colonies found in an area of eastern Venezuela where there were no known
European honey bees. They were identified as Africanized honey bees by
their behavior and characteristic comb cell size (4.5-5.0 mm wide
between opposite sides of the hexagon, see Chapter III). The European
egg source (Y5) and the European cell-producing colonies (19, 27 28, F,
H and HI) came from European queens commercially produced in the
southeastern U.S.A. and shipped to Venezuela. European colony IBR was a
stock supplied by the U.S. Department of Agriculture Bee Breeding and
Stock Center Laboratory, Baton Rouge, Louisiana, USA.


171
Fletcher D.J.C., and G.D. Tribe. 1977c. Natural emergency queen
rearing by the African bee A. ul. adansonii and its relevance for
successful queen production by beekeepers I. Pages 132-140 in
D.J.C. Fletcher (ed.) African bees: Taxonomy biology and
economic use. Apimondia Pretoria.
Free, J.B. 1965. The allocation of duties among worker honeybees.
Symp. Zool. Soc. (London) 14:39-59.
Fyg, W. 1959. Normal and abnormal development in the honeybee. Bee
World 40:57-66, 85-96.
Garofalo, C.A. 1977, Brood viability in normal colonies of Apis
mellifera. J. Apic. Res. 16:3-13.
Gary, N.E. 1975. Activities and behavior of honey bees. Pages 185-264
in Dadant and Sons (eds.), The hive and the honey bee. Dadant and
Sons, Inc., Hamilton, Illinois.
Glushkov, N.M. 1958. Experiments on combs with enlarged cells. Apic.
Absts. 9:102.
Goncalves, L.S. 1974. Comments on the aggressiveness of the
Africanized bees in Brazil. Am. Bee J. 114:448-450.
Goncalves, L.S. 1975. Do the Africanized bees of Brazil only sting?
Am. Bee J. 115:8-10.
Goncalves, L.S. 1982. The economic impact of the Africanized honey bee
in South America. Pages 134-137 in M.D. Breed, C.D. Michener and
H.E. Evans (eds.), The biology of social insects. Westview Press,
Boulder, Colorado.
Gould, J.L. 1982. Why do honey bees have dialects? Behav. Ecol.
Sociobiol. 10:53-56.
Grout, R.A. 1937. The influence of size of brood cell upon the size
and variability of the honeybee. Research Bulletin 218,
Agricultural Experiment Station, Iowa State College of Agriculture
and Mechanic Arts, Ames.
Hall, H.G. In press. DNA differences found between Africanized and
European honeybees. Proc. Natl. Acad. Sci., USA.
Harbo, J.R. 1979. Storage of honeybee spermatozoa at -196C. J. Apic.
Res. 18:57-63.
Harbo J.R., and A.B. Bolten. 1981. Development times of male and
female eggs of the honey bee. Ann. Ent. Soc. Am. 74:504-506.
Harbo, J.R., A.B. Bolten, T.E. Rinderer and A.M. Collins. 1981.
Development periods for eggs of Africanized and European honeybees.
J. Apic. Res. 20:156-159.


113
O
TABLE 6-8. Comparison of brood production in cm for Africanized and
European queens during two periods of the first brood cycle.
DAY 12
DAY 17
USB3 SBb TBC
USB SB TB
AFRICANIZED NURSE BEES
AFRICANIZED QUEENS
A62 (W81) 1053
A57 (W42) 416
EUROPEAN QUEEN
GK30 (Y63) 1775
EUROPEAN NURSE BEES
AFRICANIZED QUEENS
A62 (W85) 906
A57 (W41) 1481
EUROPEAN QUEEN
GK30 (Y61) 1614
962
666
2015
1082
1484
493
1301
818
2785
1311
819
2594
1330
1756
3086
823
487
1729
1968
434
1018
1354
1256
1788
2274
728
2342
1580
1519
3099
ANALYSES^
AFR WORKERS x EUR WORKERS NS NS NS NS NS NS
3USB = unsealed brood (eggs and larvae).
bSB = sealed brood (pre-pupae and pupae).
^TB = total brood.
dMann-Whitney U test, one-tailed, alpha = 0.05. AFR = Africanized;
EUR = European.


67
In addition, Africanized drones (not present in Butler's study) may
have a lower response threshold to queen pheromone and therefore would
respond to queens with less pheromone present than would European
drones. This hypothesis is suggested by the observation that there are
differences in the sensory receptors on the antennae of Africanized
drones compared with European drones (Dietz 1978). Further comparison
between Africanized and European drones is needed to determine any
differences in the threshold of response and whether or not this would
give the Africanized drones a mating advantage.
Another factor that needs to be considered when using this
behavioral bioassay (drone response) to compare rate of maturation of
queens 1s that the pheromone is not continually produced but rather is
pulsed in its production (R. Boch, pers. comm.). This factor may help
to explain some of the variation of responses produced by queens within
the same line. For example, in a few trials, a queen elicited a
decreased response compared with the previous response she had elicited.
In the present study, the time post-emergence to the initiation of
oviposition for Africanized queens was 8.5 days and for European queens
was 7.5 days. There are no other data available that allow for valid
comparisons. For example, Otis (1980) reports that the mean interval
from emergence to initiation of oviposition was 7.8 days for Africanized
queens in French Guiana. However, these data were collected by
observing queen maturation in colonies that had swarmed and, therefore,
the time from emergence to oviposition would be shortened because of a
variable period of queen confinement [110 days (Otis 1980)] within the
cells. In another set of data, Otis (1980) reports the mean maturation
interval from eclosin to oviposition was 9.7 days. However, he does


TABLE B-l. Coefficients of variation (CV) of worker bees in honey
bee and bumble bee colonies calculated from data
presented in the references.
CV
REFERENCES
Honey bees (Ap is mel1 ifera)
Weights
Adult (fresh)
0.4-0.6
Abdellatif 1965
4.0-4.53
Bolten (Table B-2)
5.4-7.0b
Bolten (Table B-2)
10.7-11.2
Kerr and Hebling 1964
Adult (dry)
4.8-8.1
Grout 1937
4.1-4.53
Bolten (Table B-2)
4.1-6.7
Bolten (Table B-2)
Linear Measurements
Length forewing
1.5-1.6
Grout 1937
Width forewing
2.2-2.5
Grout 1937
Length proboscis
1.6-1.9
Grout 1937
Bumble bees (Bombus)
Weights
Adult (fresh) 31.0-36.7 Brian 1952
(Bombus agrorum)
Linear Measurements0*
Length radial cell 7.4-13.8 Medler 1965
(Bombms f.erv.idiig)
^European genotypes.
Africanized genotypes in South America.
cCV may be high as a result of variable engorgement during 4 hour delay
from emergence to weighing.
Different linear measurements for Bombus are significantly correlated
(P<0.01, Medler 1962).
159


18
record time of developmental changes was outweighed by the negative
effects of disturbance on development rates.
Temperature of the brood nest in the space between the two
monitored combs was recorded periodically Ijy placing a thermometer into
that area through a hole in the hive cover. This was only an
approximate measure of temperature because development occurs within the
cells where temperature is less affected by the ventilating air currents
within the hive.
Results
Table 2-2 presents the development times for unsealed brood, sealed
brood, and total development times from oviposition to adult emergence
for Africanized and European egg genotypes in each of the four
experimental treatments. Table 2-3 summarizes the argumentation for
evaluating the interactions of egg genotypes with the colony-level
parameters of comb cell size and nurse bee genotype on worker bee
development times. The best tests to use to compare differences between
the Africanized and European egg genotypes are evaluating pairs in each
of the four experimental nurse bee colonies (A55, H2, A41, and IBR877),
i.e., AxB, CxD, ExF, and GxH. In these comparisons, the colony-level
parameters (colony population, position within the brood nest,
temperature, comb cell size, and nurse bee genotype) are identical and
allow for only differences in egg genotype to be compared. Tables 2-4,
2-5 and 2-6 summarize the results from the statistical analyses for the
unsealed brood, sealed brood, and total development times, respectively.
Data were analyzed with the Kolmogorov-Smirnov one-tailed test using the
chi square distribution, df = 2 (Siegel 1956).


146
(primarily honey) to sustain them during periods of resource dearth or
the colonies can abscond (relocate or migrate) to another area where
conditions may be better. Hoarding large surpluses of honey is
characteristic of European honey bees in temperate regions. However,
hoarding behavior may be disadvantageous in the tropics because colonies
with large food surpluses may be more easily discovered by predators and
less easily protected. Because predation has been a major evolutionary
force for tropical honey bee populations (Seeley 1983; Seeley, Seeley
and Akratanakul 1982), resource-induced absconding may be a better
evolutionary alternative to hoarding.
Resource-induced absconding occurs when an entire colony abandons a
nest and is quite common in tropical species of Apis (A. florea, A.
dorsata, and A. cerana) and tropical populations of A. mel1ifera during
periods of resource dearth (Winston, Otis and Taylor 1979; Winston,
Taylor and Otis 1983; Woyke 1976). Africanized honey bees have a high
rate of resource-induced absconding in South America (Winston, Otis and
Taylor 1979). Resource-induced absconding behavior may not be
advantageous in temperate regions because colonies may not have enough
time once they settle in a new area to store sufficient food to
successfully overwinter (Butler 1974). European honey bees generally do
not abscond in either temperate or tropical regions (Butler 1974;
Fletcher 1978; Winston, Otis and Taylor 1979; Winston, Taylor and Otis
1983).
Whether resource-induced absconding behavior is a more successful
strategy in the tropics than is hoarding behavior requires further study
to determine the advantages and disadvantages of each strategy under
tropical conditions. Both may be viable strategies and may not be


103
also no significant difference between Africanized and European worker
bees on brood production rates or egg laying rates of either Africanized
or European queens.
Differences in egg laying and brood production rates could be
evaluated by eliminating potential differences in foraging success
between Africanized and European bees by providing surplus honey to each
experimental colony and by controlling comb cell size. Differences in
brood production caused by differences in resource utilization
efficiency as a result of either smaller bee size or increased foraging
success were therefore, not evaluated. For a given amount of food, a
greater number of smaller, Africanized bees can be produced compared
with larger, European bees (Fletcher and Tribe 1977a; Tribe and Fletcher
1977). However, this advantage for Africanized bees with respect to
their smaller size, would only be present if resources were limited.
Therefore, surplus honey was provided for each colony to reduce the
effects of limited resources and the effects of differential foraging
success between the two honey bee populations. Controlling comb cell
size eliminated brood production differences based on bee size. For
example, a given number of nurse bees may be able to rear more smaller
bees than larger ones. In order that queen-worker bee interactions
could be evaluated, European comb size was selected because of the
difficulties both European queens and worker bees have with Africanized
comb as discussed earlier.
Egg laying rates observed during these experiments were within the
range reported for Africanized bees in French Guiana (Winston and Taylor
1980) but lower than the maximum reported for either population
(Fletcher 1978; Ribbands 1953). Several factors can account for this


141
Although Africanized queens have been reported to have greater egg
laying rates than European queens (Fletcher 1978; Michener 1972, 1975;
Ribbands 1953), under identical experimental conditions in Venezuela,
there was no significant difference in queen fecundity during the
initial colony growth period (Chapters V and VI). Also, European honey
bee workers live longer (Winston and Katz 1981), giving European honey
bee colonies a growth rate advantage with respect to this demographic
parameter.
Because of the relationship of worker longevity to colony growth
rates, initial colony growth rates may be affected by the age structure
of bees in a swarm. Colonies established from swarms with older bees
will have a more rapid decline in population, which will adversely
affect colony growth because egg laying rates are a function of the
number of bees in a colony (Moeller 1958). The age structure of
Africanized swarms has been evaluated (Winston and Otis 1978) but there
are no data for both Africanized and European swarms under similar
conditions.
Another parameter affecting colony growth rate is brood mortality,
but there are no data available that simultaneously compare Africanized
and European honey bees under identical conditions. Experimental and
environmental conditions are particularly important with respect to this
parameter. Rates of brood mortality can be as high as 50% and are
affected by season, resource availability and colony adult population
(Garofalo 1977; Merrill 1924; Woyke 1977). These high rates of brood
mortality and/or brood cannibalism may function to regulate protein
balance in honey bee colonies during protein (pollen) shortages (Weiss
1984). Therefore, earlier studies comparing brood mortalities in


TABLE 5-4. Correlation of queen cell length (mm) and queen pupal
weight (mg): mean + SD, (sample size). Measurement
made on day 11.25 post-ov iposit ion.
QUEEN CELL QUEEN PUPAL
QUEEN GENOTYPE LENGTH WEIGHT CORRELATIONS3
AFRICANIZED
A26
(CB1)b
2.58 + 0.1
257.1 + 7.6
NS
(9)
(9)
A26
(CB2)
2.62 + 0.2
255.8 + 9.2
*
(9)
(9)
A26
(CB3)
2.54 + 0.1
262.3 + 6.4

(3)
(3)
A26
(CB5)
2.67 + 0.1
243.0 + 13.4

(3)
(3)
A57
(CB4)
2.58 + 0.1
259.1 + 10.8
NS
(7)
(7)
A57
(CB6)
2.50 + 0.02
248.6 + 14.3
NS
(4)
(4)
A62
(CB7)
2.78 + 0.04
291.8 + 7.8
NS
(5)
(5)
A61
(CB8)
2.79 +0.1
237.2 + 21.7
NS
(5)
(5)
EUROPEAN
YK
(CB1)
2.50 + 0.1
286.4 + 12.0
NS
(9)
(9)
YK
(CB2)
2.70 + 0.1
284.4 + 20.6
NS
(9)
(9)
YK
(CB4)
2.44 + 0.1
272.2 + 17.8
**
(11)
(11)
YK
(CB5)
2.57 + 0.1
282.6 + 12.0

(3)
(3)
WE
(CB3)
2.48 + 0.1
268.4 + 8.8
NS
(8)
(8)
YD
(CB6)
2.57 + 0.1
264.0 + 15.4
NS
(5)
(5)
N
(CB7)
2.81 + 0.1
264.3 + 14.0

(3)
(3)
GK
(CB8)
2.76 + 0.1
232.9 + 0.4

(2)
(2)
aSpearman's rank correlation coefficients, alpha = 0.05;
* = P<0.05; ** = P<0.01.
bCB = cell-producing colony number; refer to Table 5-1 for
explanation.


150
nest cavities than did honey bees from southern Europe. Southern
European winters would be much less severe than those in northern
Europe, and therefore the need for larger nest cavities to store large
food surpluses is less important.
For tropical honey bee populations, there may be a selective
advantage for smaller nest cavity volumes, e.g., to facilitate
protection against infestation from wax moths (Galleria mellonella and
Achroia grisella) (Fletcher 1976). Africanized honey bees utilize a
wider variety of nest sites than do European honey bees, including
smaller cavity volumes (Fletcher 1976). The negative factors associated
with smaller nest cavities may be absent in the tropics because there is
less need to store large surpluses to survive periods of resource
dearthperiods are generally shorter and less costly with respect to
energetic demands for maintaining proper brood nest temperature. In
addition, honey bees that evolved in the tropics commonly abscond during
periods of resource dearth as opposed to hoarding surplus food.
Although nest cavity choice for Africanized and European honey bees
has not been studied under identical conditions, nest cavities selected
by Africanized honey bees in Venezuela were not smaller than cavities
selected by European honey bees in temperate regions (Rinderer, Collins,
Bolten and Harbo 1981; Rinderer, Tucker and Collins 1982). Because of
the importance of nest cavity volume to reproductive rates, nest cavity
volume for both populations needs to be investigated under identical
conditions.
Density-Dependent Factors Regulating Queen Rearing
Other parameters that might account for differences in reproductive
rates between Africanized and European honey bees may be certain


5
European honey bees exist Factors determining honey bee size and
potential problems of Africanized honey bee identification based on size
are analyzed in Chapter III.
The cuticular hydrocarbon composition of Africanized honey bees is
significantly different from that of European honey bees (Carlson and
Bolten 1984). The differences are particularly striking for the 35, 37,
39, 41 and 43 carbon alkenes and alkadienes that total over 22% of the
hydrocarbons extracted from Africanized bees but only 1-3% of the
hydrocarbons extracted from European bees. Because hydrocarbon
composition is not affected by honey bee size or diet, using hydrocarbon
analysis to distinguish between Africanized and European honey bees has
great potential. However, further research to determine heritability
patterns for different hydrocarbon components is needed.
Differences between Africanized and European honey bees have also
been demonstrated for foraging behavior (Nunez 1973, 1979a, 1982;
Rinderer, Bolten, Collins and Harbo 1984; Rinderer, Collins and Tucker
1985; Winston and Katz 1982), egg development times (Harbo, Bolten,
Rinderer and Collins 1981), selection preferences for nest cavity sizes
(Fletcher 1976; Michener 1972; Rinderer, Collins, Bolten and Harbo 1981;
Rinderer, Tucker and Collins 1982), hoarding behavior (Rinderer, Bolten,
Harbo and Collins 1982), worker bee longevity (Winston and Katz 1981),
morphometric analysis (Daly and Balling 1978), and allozyme patterns
(Nunamaker and Wilson 1981; Sylvester 1982). Africanized honey bee
populations in South America are reported to have a high colony
reproductive (swarming) rate compared with European honey bee
populations in North America (Otis 1980, 1982a; Winston 1979b, 1980a;
Winston, Dropkin and Taylor 1981; Winston, Taylor and Otis 1983).


163
TABLE B-5. Changes in European worker bee pupal weight (mg) with
changes in pupal age: mean + SD, (sample size).
Fresh weights were measured in Gainesville, Florida.
AGE (DAYS POST-OVIPOSITION)
11.5 12.5 13.5 14.5 15.5 16.5 17.5
145.8
145.5
141.9
140.9
141.1
139.1
138.0
+ 4.8
+ 3.8
+ 3.8
+ 4.2
+ 4.6
+ 4.0
+ 4.8
(10)
(45)
(31)
(38)
(31)
(32)
(34)
A
B
C
D
E
F
G
ANALYSES
CDEF
NS
A x B
NS
B x C
NS
C x D
NS
D x E
NS
E x F
NS
F x G
NS
^One-way analysis of variance, alpha = 0.05.
bt-test, two-tailed, alpha = 0.05.


69
several days before the swarm departs, allowing time for their ovaries
to recess. Therefore, maturation for Africanized queens may be delayed
in order for the swarms with new queens to be able to migrate long
distances. Rather than selection operating to shorten the maturation
interval, selection may be operating to delay maturation to enable long
swarm migration distances.
The variation in queen maturation rates (see Tables 4-8 and 4-9)
observed both within a population and within a queen line suggests that
the physiological parameters involved in the process of maturation may
be genetically determined. The rate of maturation is an important
economic characteristic for commercial queen producers to consider in
their selection programs. Reducing the time from emergence to
initiation of oviposition can significantly increase the number of
queens produced in each mating colony during the queen-producing season.


96
experimental queens were produced under identical conditions and were
the same age.
Estimates of daily egg laying rates derived from total brood
production may not be accurate (Merrill 1924) because mortality of
unsealed brood (eggs and larvae) may be quite high, up to 50% (Garofalo
1977; Merrill 1924; V/oyke 1977), particularly during the initial period
of colony growth (Winston, Dropkin and Taylor 1981). Therefore, both
daily egg laying rates and brood production during the first brood cycle
were analyzed.
Methods
D.fU.l.y, Egg_.Lay1ng_R.ates
Three different Africanized queen mothers (A57, A61 and A62) were
established from feral colonies that were found in an area of eastern
Venezuela with no known European honey bees. They were confirmed as
Africanized honey bees by their behavior and comb cell size (Chapter
III). Three different European queen mothers were shipped to Venezuela
from the U.S.A.; two (GK and YD) were from the U.S. Department of
Agriculture Bee Breeding and Stock Center Laboratory in Baton Rouge,
Louisiana, and the third (YK) was from a commercial queen producer in
southeastern U.S.A.
Experimental queens were produced from the queen mothers using
standard queen rearing techniques (Laidlaw 1979). Eggs from the queen
mothers were collected by restricting queens to a portion of comb for 4
to 6 hours under 8 x 8 cm push-in cages that had queen excluder material
soldered to the tops (Harbo, Bolten, Rinderer and Collins 1981). Combs
with the egg samples were then placed into a populous colony where the
eggs were incubated until the larvae hatched and were fed. Young


181
Winston, M.L. and S.J. Katz. 1982. Foraging differences between
cross-fostered honeybee workers (Apis mel1 ifera) of European and
Africanized races. Behav. Ecol. Sociobiol. 10:125-129.
Winston, M.L., and G.W. Otis. 1978. Ages of bees in swarms and
afterswarms of the Africanized honeybee. J. Apic. Res. 17:123-129.
Winston, M.L., G.W. Otis and O.R. Taylor. 1979. Absconding behaviour
of the Africanized honeybee in South America. J. Apic. Res. 18:85-
94.
Winston, M.L., and O.R. Taylor. 1980. Factors preceding queen rearing
in the Africanized honeybee (Apis mellifera) in South America.
Insectes Sociaux 27:289-304.
Winston, M.L., O.R. Taylor and G.W. Otis. 1983. Some differences
between temperate European and tropical African and South American
honeybees. Bee World 64:12-21.
Woyke, J. 1969. African honey bees in Brazil. Am. Bee J. 109:342-344.
Woyke, J. 1971. Correlations between the age at which honeybee brood
was grafted, characteristics of the resultant queens, and results
of insemination. J. Apic. Res. 10:45-55.
Woyke, J. 1973. Experiences with Apis mellifera adansonii in Brazil
and in Poland. Apiacta 8:115-116.
Woyke, J. 1976. Brood-rearing efficiency and absconding in Indian
honeybees. J. Apic. Res. 15:133-143.
Woyke, J. 1977. Cannibalism and brood-rearing efficiency in the
honeybee. J. Apic. Res. 16:84-94.
Woyke, J. 1979. Effects of the access of worker honeybees to the queen
on the results of the instrumental insemination. J. Apic. Res.
19:136-143.
Woyke, J. 1984. Correlations and interactions between population,
length of worker life and honey production by honeybees in a
temperate region. J. Apic. Res. 23:148-156.
Zeuner, F.E., and F.J. Manning. 1976. A monograph on fossil bees
(Hymenoptera: Apoidea). Bull. Brit. Mus. (Nat. Hist.), Geol.
27:151-268.
Zmarlicki, C. and R.A. Morse. 1963. Drone congregation areas. J.
Apic. Res. 2:64-66.


4
22 meters) and remain disturbed for a greater period of time (28 versus
3 minutes) (Stort 1971 cited in Goncalves 1974). Differences in defense
behavior between Africanized and European bees do not appear to be a
function of either quantitative differences in pheromone production
(Crewe and Hastings 1976) or numbers of olfactory structures on the
antennae (Stort and Barelii 1981).
There is a difference in natural comb cell size between Africanized
and European populations. The width between opposite sides of the
hexagonal cells for the Africanized population in Brazil averaged 5.0 mm
compared with 5.4 mm for the European population in Canada (Michener
1972). In a recent study, cells built by Africanized swarms in
Venezuela were 4.8-4.9 mm wide and those built by European swarms in
Louisiana, U.S.A. were 5.2-5.3 mm wide (Rinderer, Tucker and Collins
1982). Adult Africanized bees are smaller than European bees (62 mg
compared with 93 mg, unengorged) (Otis 1982b; Otis, Winston and Taylor
1981). However, as Africanized honey bees disperse into areas with
extensive European honey bee populations, size differences between the
two populations may become less distinct. Increased hybridization
between the two populations could result in bees with an Africanized
genome developing in European comb cells, resulting in larger
Africanized bees. Therefore, methods used to identify Africanized honey
bees based on size parameters, for example, morphometric analysis, may
become less reliable. As Daly, Hoelmer, Norman and Allen (1982) point
out, there is a "difficulty in using phenotype characters to identify
genetically different, but closely related populations" (p. 593). This
will be more evident as Africanized honey bees disperse into areas of
Central America and particularly Mexico, where large populations of


TABLE 5-1. Comparison of Africanized and European queen pupal weights
(mg): mean + SD, (sample size), (genotype).
87
AFRICANIZED EUROPEAN
CELL BUILDER3 GENOTYPES GENOTYPES ANALYSES5
1
257.1
+ 7.6
(9)
(A26)
2
255.8
+ 9.2
(9)
(A26)
3
262.3
+ 6.4
(3)
(A26)
4
260.0
+ 9.0
(11)
(A57)
5
243.0
+ 13.4
(3)
(A26)
6
248.6
+ 14.3
(4)
(A57)
7
291.8
+ 7.9
(5)
(A62)
8
237.2
+ 21.7
(5)
(A61)
286.4
(9)
+ 12.0
(YK)
***
284.4
(9)
+ 20.6
(YK)
**
266.7
(9)
+ 9.7
(WE)
NS
272.5
(13)
+ 16.5
(YK)
*
282.6
(3)
+ 12.0
(YK)
*
264.0
(5)
+ 15.4
(YD)
NS
257.3
(4)
+ 18.2
(N)
c
232.9
(2)
+ 0.4
(GK)
NS
aCe11-producing colonies; European nurse bees and European comb
cell size.
bMann-Whitney U test, one-tailed, alpha = 0.05;
* = P<0.05, ** = P<0.01, *** = P<0.001.
difference in wrong direction for one-tailed test; two-tailed
test results in a P<0.02.


117
Table 6-12. Comparison of egg laying rates during daily egg laying
rate experiment and brood production experiment.
AFRICANIZED EUROPEAN
NURSE BEES NURSE BEES
QUEENS
ELRa
BP12b
BP17C
ELR
BP12
BP17
AFRICANIZED
A62 (W81)

714
696
916


A62 (W85)
799



612
447
A57 (W42)

383
328
700
__
A57 (W41)
694



697
568
EUROPEAN
GK (Y63)

918
772
737
GK (Y61)
824

*
829
775
ANALYSES^
ELR
X
BP12
NS
ELR
X
BP 17
NS
BP12
X
BP 17
P<0.05
aDaily egg laying rate experiment, means.
bBrood production experiment; egg laying rate estimated for first
12 days of brood cycle by dividing total brood at day 12 by 12.
cBrood production experiment; egg laying rate estimated for first
17 days of brood cycle by dividing total brood at day 17 by 17.
dSpearman rank correlation, alpha = 0.05; Africanized and
European nurse bees combined.


Harbo, J.R., and T.I. Szabo. 1984. A comparison of instrumentany
inseminated and naturally mated queens. J. Apic. Res. 23:31-36.
172
Harp, E.R. 1973. A specialized system for multiple rearing of quality
honeybee queens. Am. Bee J. 113:256-258, 261.
Heinrich, B. 1979a. Bumblebee economics. Harvard Univ. Press,
Cambridge, Mass.
Heinrich, B. 1979b. Thermoregulation of African and European honeybees
during foraging, attack, and hive exits and returns. J. Exp. Biol.
80:217-229.
Holdridge, L.R. 1964. Life zone ecology. Tropical Science Center, San
Jose, Costa Rica.
Hoopingarner, R., and C.L. Farrar. 1959. Genetic control of size of
queen honey bees. J. Econ. Ent. 52:547-548.
Jay, S.C. 1963. The development of honeybees in their cells. J. Apic.
Res. 2:117-134.
Jaycox, E.R., and S.G. Parise. 1980. Homesite selection by Italian
honey bee swarms, Apis mellifera 1iqustica (Hymenoptera: Apidae).
J. Kans. Ent. Soc. 53:171-178.
Jaycox, E.R., and S.G. Parise. 1981. Homesite selection by swarms of
black-bodied honey bees, Apis mellifera caucasica and A. m. carnica
(Hymenoptera: Apidae). J. Kans. Ent. Soc. 54:697-703.
Jeffree, E.P. 1955. Observations on the decline and growth of honey
bee colonies. J. Econ. Ent. 48:723-726.
Johansson, T.S.K., and M.P. Johansson. 1973. Methods for rearing
queens. Bee World 54:149-175.
Kerr, W.E. 1967. The history of the introduction of African bees to
Brazil. S. Afr. Bee J. 39:3-5.
Kerr, W.E., and D. Bueno. 1970. Natural crossing between Apis
roe.11 if era adansonii and Apis mell if era 1 igus.lica. Evolution
24:145-155.
Kerr, W.E., L.S. Goncalves, L.F. Blotta and H.B. Made!. 1972.
Biologa comprada entre as abelhas italianas (Apis mel1 ifera
1iqustica) Africana (Apis mellifera adansonii) e suas hibridas.
Pages 151-185 in Anais do Io Congresso Brasileiro de Apicultura.
Congresso Brasileiro de Apicultura, Florianopolis, Brazil.
Kerr, W.E., and N.J. Hebling. 1964. Influence of the weight of worker
bees on division of labor. Evolution 18:267-270.
Koeniger, N., and H.N.P. Wljayagunasekera. 1976. Time of drone flight
in the three Asiatic honeybee species. J. Apic. Res. 15:67-71.


42
sealed with strongly convex cappings similar to the way cells containing
drones are sealed in order to accomodate their larger size. Africanized
bee pupae (123.8 +6.2 mg) that developed in European comb cells were
smaller than European bee pupae (139.5 + 54 mg) that also developed in
European comb cells (BDF x NPR, P<0.001). For each of the nine
genotypes investigated, worker bee pupae that developed in Africanized
comb cells were smaller than pupae that developed in European comb
cells, PcO.OOl. There is a 43% increase in comb cell volume between
Africanized and European combs (Table B-3), but the Africanized and
European genotypes only increased in pupal weight by 11.4% and 13.1%,
respectively (Table 3-2). These results show that both genotype and
comb cell size affect worker bee size.
Table 3-4 presents the results for the pupal weights of the
reciprocal F^ hybrids and their respective maternal lines. Data from
only European comb cells were used in order to observe genotype effects
without the constraint of the small Africanized comb cells on European
genotypes. Table 3-5 summarizes the results of the statistical
analyses. The pupal weights of the hybrids from this reciprocal F^
cross were significantly different from each other (H x J; H x L;
P<0.001), but were the same as their respective maternal line (B x H; J
x R; L x R).
Discussion
Reduction of Bee Size Variation
Bee size is a result of not only the interaction of egg genotype
and comb cell size but also the maternal genotype. This can be seen by
evaluating the reciprocal hybrid crosses. The genotype component for
bee size is not a result of "simple" inheritance because pupal weights


37
Honey bee size and size variation is a result of both genetic and
environmental factors. Research has focused primarily on the extrinsic
factors that affect bee size (e.g., comb cell size, nutrition and
temperature). Honey bee worker sizes and honey bee comb cell sizes have
been shown to be inter-related: because of the manner by which comb
cells are constructed (Darwin 1859/1958), worker body size affects the
diameter of cells they construct (Baudoux 1933; Glushkov 1958), and
worker bee size is correlated to the size of cells in which they are
reared (Baudoux 1933; Buchner 1955; Glushkov 1958; Grout 1937; Michailov
1927-28 cited in Alpatov 1929; Tuenin 1927).
This interaction between comb cell size and egg genotype may at
first appear to provide a mechanism for both regulating bee size and
reducing size variation among bees within a colony. However, the comb
cell itself can become a source of variation in bee size. Although comb
cell size appears quite uniform, especially when first constructed, the
cells become variable in size as the number of generations reared in
them increases, because pupal cocoons adhere to the cell walls, reducing
cell diameter (Abdellatif 1965; Alpatov 1929; Buchner 1955; Grout 1937).
For example, there is a 25% reduction in cell volume between cells from
new and old combs (Table B-3). Comb cell volume has a greater variance
than cell diameter and is not correlated with diameter (Table B-3).
In addition to comb cell size, there are other extrinsic factors
that affect development and resultant bee size, e.g., quantity and
quality of larval food, and temperature and humidity at which the larvae
and pupae are reared (Buchner 1955; Fyg 1959; Jay 1963; Kulzhinskaya
1956; Michailov 1927-28 cited in Alpatov 1929). These same factors not
only affect absolute size but are sources of size variation.


120
(Rinderer, Collins,. Bolten and Hanbo 1981; Rinderer, Tucker and Collins
1982); allozyme patterns (Nunamaker and Wilson 1981; Sylvester 1982);
cuticular hydrocarbon composition (Carlson and Bolten 1984, and
unpublished data); adult bee size and comb cell size (Chapter III;
Michener 1975); and morphometric relationships (Daly and Balling 1978).
This apparent lack of evidence for hybridization has been
attributed primarily to some degree of reproductive isolation between
the Africanized and European populations (Kerr and Bueno 1970; Taylor
1985). Three isolating mechanisms have been suggested: assortative
mating (Kerr and Bueno 1970); physiological incompatibility with respect
to the drone ejaculation response (Kerr and Bueno 1970); and differences
in drone and presumably the queen flight times between the Africanized
and European populations (Taylor 1985; Taylor, Kingsolver and Otis in
press). At best, these mechanisms may be only partially effective and
are not likely to account for the apparent lack of hybridization. For
example, Kerr and Bueno (1970) present data to support assortative
mating even though 35% and 42% of the matings evaluated were hybrid.
With respect to differences in queen and drone flight times, data from
Venezuela suggest that mean peak drone flight times for Africanized and
European populations are separated by only 23 minutes and that drones
from both populations are present in the mating areas at all times
during the approximately three hour flight period (Taylor, Kingsolver
and Otis in press).
A more probable argument for the maintenance of the African
characteristics is based on selection advantages for the African
genotype in tropical habitats of South America, which are characterized
by resource distribution patterns similar to the ones in Africa where


11
nest; colony size, which affects both brood nest temperature and feeding
frequency and quality; and nectar and pollen resources, which also
affect feeding quantity and quality. Valid comparison of development
times between genotypes or populations can only be made when these
factors are controlled under similar experimental conditions.
The importance of slight temperature differences on development
time cannot be overstated. Development times for European worker bees
in Wisconsin, U.S.A., averaged 20.5 days but ranged from 20-24 days,
depending on differences in temperature in different areas of the brood
nest (Milum 1930). Harbo and Bolten (1981) showed that fertilized eggs
kept at 34.8C hatched about 1.4 hours sooner than those kept at 34.3C.
This difference in egg hatch time for only a 0.5C difference in
temperature can be extrapolated to approximately 10 hours for the entire
development period [calculated from Harbo and Bolten (1981)]. However,
normal temperatures within a brood nest can vary to a much greater
extent (Milum 1930; Jay 1963). For example, when Tribe and Fletcher
(1977) determined the development rates of African honey bees in South
Africa, they recorded that temperatures in the brood nest varied from
26-34C.
In an incubator with controlled temperature and humidity, eggs from
Africanized genotypes hatched significantly sooner than eggs from
European genotypes, 69.6 + 1.06 hours compared with 73.3 + 1.14 hours
(Harbo, Bolten, Rinderer and Collins 1981). Egg development requires
that only temperature and humidity be controlled and can therefore be
evaluated independently of colony-level parameters. However,
differences between Africanized and European genotypes for total worker
bee development periods need to be evaluated within a colony in order to


17
or forty eggs selected to be monitored for development formed either a
3x10 or a 4x10 cell area. A reference point which facilitated locating
the designated development sample was indicated by colored pins inserted
five cells to the left of each original egg row.
The two combs (one comb from each egg source) were placed in the
center of each experimental colony at the same time. The cells with the
eggs to be monitored faced each other in order to reduce any effects of
brood nest position. Two frames of brood were removed from each
experimental colony to make room for the experimental frames. This also
reduced the amount of brood being reared in each experimental colony,
insuring that the monitored eggs would be optimally fed.
The pairs of frames were put into the four experimental colonies on
four successive days because of the 24 hours needed to collect each set
of eggs. Once the eggs were put into the experimental colonies, they
were inspected every day at 0800 hours. The survivorship and
developmental status of each original test group egg was recorded.
The sample size of eggs monitored was selected to minimize the time
each colony would need to be opened for observation in order to minimize
disturbance. When a colony was disturbed, bees flew from the comb and
ran on the bottom of the hive, resulting in temperature fluctuations and
interrupted feeding of larvae. Colonies were carefully opened, using
minimal amounts of smoke. Adult bees were not shaken off the combs but
rather gently pushed aside to observe the development stage within the
cells. Inspections more frequent than every 24 hours also increased the
level of disturbance, especially in the Africanized colonies (A55 and
A4I). The advantage of more frequent monitoring to more accurately


149
Nest Sites and Cavity Volume
Fletcher (1976) suggested another adaptive advantage that
Africanized bees have is their ability to utilize a greater variety of
nest sites. Fletcher may be confusing cause with effect when he
suggests that it is this ability that "has enabled them to establish
themselves in areas not previously inhabited by honey-bees at all"
(1976, p. 6). A more likely explanation for the success of Africanized
bees in those areas would be their ability to utilize the particular
nectar and pollen resources available. That is, without a more
efficient utilization of resources, Africanized honey bees would not be
able to exploit these other habitats irrespective of their ability to
utilize a greater variety of nest sites. As discussed above,
Africanized honey bees are more successful foragers than are European
honey bees under resource conditions typical of tropical habitats.
Nest cavity volume is another factor affecting reproduction in
honey bees. One of the stimuli for reproductive swarming is brood-nest
crowding (Baird and Seeley 1983; Simpson 1966, 1973; Simpson and Riedel
1963; Winston and Taylor 1980). Colonies inhabiting smaller cavities
become crowded more rapidly and have a higher tendency to swarm.
Colonies established in large cavities would be less crowded and have a
lower rate of swarming. In temperate regions, small cavities would be
selected against because there would be less volume available for
storing surplus honey to enable the colony to overwinter. Therefore,
Seeley proposed that nest-cavity volume may "regulate mature colony size
at an optimum between small colonies with low survivorship and large
colonies with low fertility" (Seeley 1977, p. 226). Jaycox and Parise
(1980, 1981) found that honey bees from northern Europe selected larger


16
The experimental colonies were placed in an apiary under a roof
with completely open sides. The roof served two purposes. First the
colonies were in complete shade which reduced any effects from
differences in ambient temperature and sunlight. Second, colonies could
be opened and inspected in order to monitor development with a minimum
of disturbance, especially during rain.
Eggs were collected from queens of the two designated egg source
lines (A26 and Y5), that were established in modified colonies similar
to those used by queen producers in the U.S.A. (Harp 1973). These
colonies consisted of five standard frames with the middle frame
isolated from the four others by queen excluder side and top panels.
The excluder panels have a mesh size that restricts the queen from
passing through because of her wider thorax, but allows worker bees to
pass through to feed and communicate with the queen. Thus, the queen
was isolated on a specific comb so that eggs could be collected that
would then be placed into one of the four experimental colonies to
evaluate development time.
The comb used had the appropriate comb cell size for the
experimental colony into which it would be placed. Comb cell size was
measured in each experimental nurse bee colony and for each Africanized
and European egg comb put into each experimental colony.
The queens were caged on each comb for 24 hours so that a large,
uniform egg sample could be collected. Eggs were monitored only from
the center of each frame, which insured a more uniform temperature
during development as well as uniform brood nest position. The large
egg sample also insured that the monitored eggs were in a normal
environment, surrounded by similarly-aged developing bees. The thirty


169
Bodenheimer, F.S., and A. Ben-Nerya. 1937. One-year studies on the
biology of the honey-bee in Palestine. Ann. Appl. Biol. 24:
385-403.
Bolten, A.B., and J.R. Harbo. 1982. Numbers of spermatozoa in the
spermatheca of the queen honeybee after multiple inseminations with
small volumes of semen. J. Apic. Res. 21:7-10.
Brian A.D. 1952. Division of labour and foraging in Bombus agrorum
Fabricius. J. Anim. Ecol. 21:223-240.
Brian, M.V. 1965. Social insect populations. Academic Press, London.
Brother Adam. 1966. In search of the best strains of bees.
Ehrenwirth, Munich.
Buchmann, S.L. 1982. Africanized bees confirmed in Panama. Am. Bee J.
122:322.
Buchner, R. 1955. Effect on the size of workers of restricted space
and nutrition during larval development. Ap1c. Absts. 6:15.
Butler, C.G. 1971. The mating behaviour of the honeybee (Apis
mellifera L.). J. Ent. (A) 46:1-11.
Butler, C.G. 1974. The world of the honeybee. Collins, London.
Butler, C.G., D.H. Calam and R.K. Callow. 1967. Attraction of Apis
mellifera drones by the odours of the queens of two other species
of honeybees. Nature 213:423-424.
Cantwell, G.E. 1974. The African (Brazilian) bee problem. Am. Bee J.
114:368-372.
Carlson, D.A., and A.B. Bolten. 1984. Identification of Africanized
and European honey bees, using extracted hydrocarbons. Bull. Ent.
Soc. Am. 30:32-35.
Collins, A.M., T.E. Rinderer, J.R. Harbo and A.B. Bolten. 1982. Colony
defense by Africanized and European honey bees. Science 218:72-74.
Crewe, R.M., and H. Hastings. 1976. Production of pheromones by
workers of Apis mellifera adansoni1. J. Apic. Res. 15:149-154.
Daly, H.V., and S.S. Balling. 1978. Identification of Africanized
honeybees in the Western Hemisphere by discriminant analysis. J.
Kans. Ent. Soc. 51:857-869.
Daly, H.V., K. Hoelmer, P. Norman and T. Allen. 1982. Computer-
assisted measurement and identification of honey bees (Hymenoptera:
Apidae). Ann. Ent. Soc. Am. 75:591-594.
Darwin, C. 1958. The origin of species. The New American Library,
Inc. New York. (Original work published 1859).


129
Competition with an established population of honey bees for limited
floral nectar and pollen resources will be much greater than Africanized
bees have previously experienced in any areas in South America. This
competition will greatly slow their dispersal. In many areas of Mexico
and southern U.S.A., pollen and nectar resources are already close to
being saturated by the existing honey bee population. In addition,
under temperate resource conditions, the foraging behavior of
Africanized honey bees (which is more appropriate to tropical resource
patterns) will be at a disadvantage relative to the foraging behavior of
the European honey bee population, which is characterized by greater
colony recruitment (Rinderer, Bolten, Collins and Harbo 1984; Rinderer,
Collins and Tucker 1985; Visscher and Seeley 1982).
The selective advantage of the foraging and/or thermoregulatory
behavior of European honey bees has been demonstrated in temperate
regions. African honey bee queens were introduced into North America
during the late 1800's and early 1900's when the beekeeping industry in
the U.S.A. was developing (Morse et al. 1973) and more recently, into
Louisiana (Cantwell 1974; Morse et al. 1973; Taber 1961). However, due
to hybridization and selection against the African genotype, the impact
of these introductions of African bees is undetectable today. There
have also been unsuccessful introductions of African and Africanized
bees into Europe (Cantwell 1974; Morse et al. 1973; Woyke 1973).
Therefore, these factorsa large, established population of European
honey bees, and both a foraging and thermoregulatory behavior in
European bees better adapted to temperate conditionsprecludes using
South and Central America as a model for North America in predicting the
impact, as well as the rate of spread, of Africanized honey bees.


BIOLOGY OF AFRICANIZED AND EUROPEAN HONEY BEES,
Apis mail ifera, IN VENEZUELA
By
ALAN B. BOLTEN
A DISSERTATION PRESENTED TO THE GRADUATE SCHOOL
OF THE UNIVERSITY OF FLORIDA IN
PARTIAL FULFILLMENT OF THE REQUIREMENTS
FOR THE DEGREE OF DOCTOR OF PHILOSOPHY
UNIVERSITY OF FLORIDA

"He must be a dull man who can examine the
exquisite structure of a comb, so beautifully
adapted to its end, without enthusiastic admiration."
Charles Darwin 1859

ACKNOWLEDGEMENTS
I would like to thank the members of my committee: Dr. Thomas C.
Emmel, my chairman, for his continued support, guidance and
encouragement throughout my graduate education; Dr. Malcolm T. Sanford
for stimulating discussions and his thorough editing; and Dr. Jonathan
Reiskind for his enthusiasm and helpful suggestions. I appreciate the
comments made by Drs. James Nation and Frank Nordlie on the
dissertation. I am also grateful to Professor Frank Robinson for
introducing me to the excitement and challenges of honey bee research
and management. Drs. John Harbo, Anita Collins and Tom Rinderer were
excellent field companions, sharing their knowledge of honey bee
research techniques, and creating a stimulating research environment,
both in Venezuela and during my work in Baton Rouge. I particularly
want to thank Dr. John Harbo for the instrumental inseminations and
acknowledge his collaboration on both the bee size and egg laying rate
experiments. I would also like to thank Dr. Orley Taylor for giving me
the opportunity to study Africanized honey bees.
This research was supported by the U.S. Department of Agriculture
Cooperative Agreement No. 58-7B30-8-7 with the University of Kansas (0.
R. Taylor, principal investigator). The Ministerio de Agricultura y
Cria de Venezuela provided research facilities near Maturin. I would
like to thank Med. Vet. Ricardo Gomez Rodriguez for his hospitality and

logistic support. Laboratory facilities at the Universidad de Oriente
in Jusepin were made available by Professor Dick Pulido.
The research presented in this dissertation and the commitment to
complete the writing could not have been accomplished without the
collaboration, companionship, encouragement and insights of my wife,
Karen Bjorndal, who shared not only the excitement and successes but
also the frustrations and discomforts of Africanized honey bee research.
Finally, I would like to thank my parents, who have always
supported and encouraged my work.
iv

TABLE OF CONTENTS
Page
ACKNOWLEDGEMENTS 111
ABSTRACT v11
CHAPTERS
I INTRODUCTION 1
Evolutionary Origin and Distribution of Honey Bees 1
Importation of African Honey Bees into Brazil and
Their Dispersal Throughout South and Central America . 1
Characteristics of Africanized Honey Bees 3
Purpose of My Research 6
Identification of Honey Bees Used in My Research 7
When and Where Research Was Conducted 7
II WORKER BEE DEVELOPMENT TIMES .... 9
Introduction .... 9
Methods 14
Results 18
Discussion 20
IIIINTERACTION OF MATERNAL GENOTYPE, EGG GENOTYPE AND COMB
CELL SIZE ON HONEY BEE WORKER SIZE AND SIZE VARIATION ... 35
Introduction ............ 35
Methods ..... 38
Results 41
Discussion 42
IVQUEEN DEVELOPMENT AND MATURATION .... 55
Introduction 55
Methods 57
Results ................. 63
Discussion 65
v

VQUEEN PUPAL WEIGHTS 79
Introduction ...... . 79
Methods 80
Results 83
Discussion ....... ...... 84
VIEGG LAYING AND BROOD PRODUCTION RATES
DURING THE FIRST BROOD CYCLE ...... 92
Introduction 92
Methods 96
Results 101
Discussion ...... 102
VIISUCCESSFUL HYBRIDIZATION BETWEEN AFRICANIZED
AND EUROPEAN HONEY BEES IN VENEZUELA WITH
IMPLICATIONS FOR NORTH AMERICA 118
Introduction 118
Methods 123
Results 124
Discussion 125
VIIIDISCUSSION: FACTORS CONTRIBUTING TO THE SELECTION
ADVANTAGE OF AFRICANIZED HONEY BEES IN SOUTH AMERICA
THE RESOURCE UTILIZATION EFFICIENCY HYPOTHESIS 133
Success of Introduced Populations of Honey Bees 133
Factors Affecting Honey Bee Reproductive Rates ....... 135
Factors Contributing to the Selective Advantage
of Africanized Honey Bees in South America 140
Potential Impact of Africanized Honey Bees
in North America 153
APPENDICES
A WORKER BEE DEVELOPMENT TIMES AND MORTALITY
DURING DEVELOPMENT .......... 156
B HONEY BEE SIZE, COMB CELL SIZE AND
SIZE VARIATION 159
C CHANGES IN QUEEN PUPAL WEIGHT WITH AGE ...... 165
D ACCURACY OF TECHNIQUE USED TO ESTIMATE
NUMBER OF BEES IN A COLONY 167
LITERATURE CITED ... ......... 168
BIOGRAPHICAL SKETCH .... ........ 182
vi

Abstract of Dissertation Presented to the Graduate School
of the University of Florida in Partial Fulfillment of the
Requirements for the Degree of Doctor of Philosophy
BIOLOGY OF AFRICANIZED AND EUROPEAN HONEY BEES,
Apis me!1 ifera, IN VENEZUELA
By
Alan B. Bolten
August 1986
Chairmanj Thomas C. Emmel
Major Department: Zoology
To determine factors responsible for the greater success of
Africanized honey bees, Apis mel1 ifera, in tropical regions of South
America, demographic parameters affecting colony reproductive rates were
evaluated for Africanized and European honey bees under identical
conditions in Venezuela. Worker bee development time was evaluated as
an interaction between egg genotype, comb cell size and nurse bee
genotype. Africanized worker bees developed faster than European bees:
18.9 and 19.8 days, respectively. There was no significant effect of
comb cell size or nurse bee genotype. Mortality for different
developmental stages was recorded. The relationship of worker bee
development time to colony growth rate is discussed.
Africanized queens develop in 14.5 days post-oviposit ion compared
with 15.0 days for European queens. Queen pupal weights were not
significantly different. Post-emergence maturation rates for
Africanized and European queens were similiar as determined by both the

ages when queens attracted drones and the ages when oviposit ion was
initiated.
Daily egg laying rates and brood production during initial colony
growth were not significantly different for Africanized and European
queens. Africanized and European worker bees did not differentially
affect egg laying and brood production rates.
Differences in reproductive rates between Africanized and European
honey bees in South America cannot be attributed to differences in
intrinsic demographic factors. A hypothesis based on differences in
resource utilization efficiency is presented to explain the success of
Africanized bees compared with European bees in South America.
Results from reciprocal crosses indicate that bee size is a
function of egg genotype, comb cell size and maternal genotype. The
importance of maternal inheritance for reducing worker bee size
variation within a colony is discussed. Advantages of smaller worker
bee size are evaluated for Africanized bees.
There are no effective reproductive isolating mechanisms operating
between Africanized and European honey bee populations. Both
Africanized and European queens mated with equal success with
Africanized drones as measured by the numbers of spermatozoa in the
spermatheca. The potential impact of Africanized bees on North America
is analyzed with respect to hybridization and genetic introgression,
resource competition, and selection advantages for European bees in
temperate regions.
v i i i

CHAPTER I
INTRODUCTION
Evolutionary Origin and Distribution of Honey Bees
Honey bees of the genus Apis have their greatest diversity in Asia
(Michener 1979). Earliest fossil evidence for the genus is from
01igocene deposits in Europe (Zeuner and Manning 1976). The
evolutionary relationships of the four generally recognized species of
Apis are reviewed by Michener (1974). Three of the species (A. cerana.
A. dorsata and A. florea) are native only to Asia (Michener 1979;
Ruttner 1975). The western honey bee (A. me!1 ifera) is native to
Africa, western Asia and Europe and may have evolved in tropical or
subtropical Africa (Wilson 1971) or the Near East (Ruttner 1975). The
widely different climatic conditions and floral resources under which
populations of A. mel1ifera evolved have resulted in a number of
geographically recognizable subspecies (Alpatov 1929, 1933; Br. Adam
1966; Dupraw 1965; Ruttner 1968, 1975, 1976a, 1976b; Smith 1961; Wafa,
Rashad and Mazeed 1965).
Importation of African Honey Bees into Brazil and Their Dispe.rs.a_l
Throughout South and Central America
European honey bees (A. mel1ifera mel1ifera and A. m. 1igustica)
had been introduced into Brazil by 1845 (Gerstaker cited in Pellet 1938
Woyke 1969). A. m. mel1ifera is native to Europe in the regions west
and north of the Alps and extending east into Central Russia; A. m.
1

2
1 iaustica is native to the Italian peninsula (Ruttner 1975). Because
these European honey bee populations were not very successful in
tropical and subtropical habitats of Brazil (Michener 1972) researchers
believed that they could improve Brazil's honey production by breeding a
honey bee better adapted to local conditions (Woyke 1969). With this
intention, honey bee queens from South Africa (A. m. scute!lata,
formerly classified as adansonii, see Ruttner 1976a, 1976b, 1981) were
imported into southeastern Brazil in 1956 (Kerr 1967). The following
year, swarms escaped and hybridized with established European honey
bees. The descendents from this hybridization are known as Africanized
honey bees (Goncalves 1982). Details of the introduction and subsequent
spread throughout South America have been extensively reviewed
(Goncalves 1974, 1975, 1982; Kerr 1967; Michener 1972, 1975; Taylor
1977, 1985; Taylor and Williamson 1975; Woyke 1969).
In the 30 years since African honey bees were imported into
southeastern Brazil, their hybridized offspring have rapidly dispersed
throughout tropical South and Central America and are now as far north
as Honduras and El Salvador (Rinderer 1986). The dispersion from their
original importation site into new areas has been rapid200-500 km per
year (Taylor 1977, 1985; Winston 1979a). As Africanized honey bees have
colonized new areas, they have achieved dramatic population densities
(Michener 1975). There are now probably more than ten million feral
colonies in South and Central America (Winston, Taylor and Otis 1983).
Their success in these new habitats, compared with the lack of success
of European honey bee populations, may be attributed to their foraging
behavior which is more suited to the resource patterns of the tropics
(Nunez 1973, 1979a, 1982; Rinderer, Bolten, Collins and Harbo 1984;

3
Rinderer, Collins and Tucker 1985; Winston and Katz 1982). As a result
of both foraging success and the length of time throughout the year that
resources are available in the tropics, Africanized honey bees have a
high annual reproductive rate, which is responsible for both their rate
of dispersal into new areas and their high colony densities. Net
reproductive rates for Africanized bees are estimated to be 16 colonies
per colony per year based on demographic data collected in French Guiana
(Otis 1980, 1982a), compared with 0.92-0.96 (Seeley 1978) or, when
afterswarms are considered, 3-3.6 (Winston 1980a; Winston, Taylor and
Otis 1983) colonies per colony per year for European honey bees in North
America.
Characteristics of Africanized Honey Bees
The most well known characteristic that differentiates Africanized
honey bees from European honey bees is their stinging behavior (Collins,
Rinderer, Harbo and Bolten 1982; Stort 1974, 1975a, 1975b, 1975c, 1976).
Because of their stinging behavior, Africanized bees are a health hazard
for both humans and domestic animals (Taylor 1986). Collins, Rinderer,
Harbo and Bolten (1982) compared the colony defense behavior of
Africanized honey bees in Venezuela with European bees under identical
conditions in Venezuela and with a population of European bees in
Louisiana, U.S.A. Africanized honey bees responded more rapidly and in
much greater numbers, resulting in 5.9 times as many stings in a target
compared with European honey bees in Venezuela and 8.2 times as many
stings compared with European bees in Louisiana. Two additional
components of Africanized honey bee defense behavior increase their
potential as a health hazard. Compared to European bees, Africanized
bees pursue a source of disturbance for a greater distance (160 versus

4
22 meters) and remain disturbed for a greater period of time (28 versus
3 minutes) (Stort 1971 cited in Goncalves 1974). Differences in defense
behavior between Africanized and European bees do not appear to be a
function of either quantitative differences in pheromone production
(Crewe and Hastings 1976) or numbers of olfactory structures on the
antennae (Stort and Barelii 1981).
There is a difference in natural comb cell size between Africanized
and European populations. The width between opposite sides of the
hexagonal cells for the Africanized population in Brazil averaged 5.0 mm
compared with 5.4 mm for the European population in Canada (Michener
1972). In a recent study, cells built by Africanized swarms in
Venezuela were 4.8-4.9 mm wide and those built by European swarms in
Louisiana, U.S.A. were 5.2-5.3 mm wide (Rinderer, Tucker and Collins
1982). Adult Africanized bees are smaller than European bees (62 mg
compared with 93 mg, unengorged) (Otis 1982b; Otis, Winston and Taylor
1981). However, as Africanized honey bees disperse into areas with
extensive European honey bee populations, size differences between the
two populations may become less distinct. Increased hybridization
between the two populations could result in bees with an Africanized
genome developing in European comb cells, resulting in larger
Africanized bees. Therefore, methods used to identify Africanized honey
bees based on size parameters, for example, morphometric analysis, may
become less reliable. As Daly, Hoelmer, Norman and Allen (1982) point
out, there is a "difficulty in using phenotype characters to identify
genetically different, but closely related populations" (p. 593). This
will be more evident as Africanized honey bees disperse into areas of
Central America and particularly Mexico, where large populations of

5
European honey bees exist Factors determining honey bee size and
potential problems of Africanized honey bee identification based on size
are analyzed in Chapter III.
The cuticular hydrocarbon composition of Africanized honey bees is
significantly different from that of European honey bees (Carlson and
Bolten 1984). The differences are particularly striking for the 35, 37,
39, 41 and 43 carbon alkenes and alkadienes that total over 22% of the
hydrocarbons extracted from Africanized bees but only 1-3% of the
hydrocarbons extracted from European bees. Because hydrocarbon
composition is not affected by honey bee size or diet, using hydrocarbon
analysis to distinguish between Africanized and European honey bees has
great potential. However, further research to determine heritability
patterns for different hydrocarbon components is needed.
Differences between Africanized and European honey bees have also
been demonstrated for foraging behavior (Nunez 1973, 1979a, 1982;
Rinderer, Bolten, Collins and Harbo 1984; Rinderer, Collins and Tucker
1985; Winston and Katz 1982), egg development times (Harbo, Bolten,
Rinderer and Collins 1981), selection preferences for nest cavity sizes
(Fletcher 1976; Michener 1972; Rinderer, Collins, Bolten and Harbo 1981;
Rinderer, Tucker and Collins 1982), hoarding behavior (Rinderer, Bolten,
Harbo and Collins 1982), worker bee longevity (Winston and Katz 1981),
morphometric analysis (Daly and Balling 1978), and allozyme patterns
(Nunamaker and Wilson 1981; Sylvester 1982). Africanized honey bee
populations in South America are reported to have a high colony
reproductive (swarming) rate compared with European honey bee
populations in North America (Otis 1980, 1982a; Winston 1979b, 1980a;
Winston, Dropkin and Taylor 1981; Winston, Taylor and Otis 1983).

However those investigations have not been conducted under similar
environmental or experimental conditions. Therefore, comparisons of
reproductive rates between Africanized and European honey bees using
those data are inappropriate for either identifying differences in
reproductive rates for tropical and temperate honey bee populations or
for identifying factors responsible for the success of Africanized bees
in tropical regions.
Purpose of Mv Research
African and European honey bee populations evolved under very
different resource and climatic conditions. The presence of both
Africanized and European honey bees in Venezuela provided the
opportunity to study both populations under identical conditions in the
tropics. Differences between the two honey bee populations that make
Africanized bees more successful in tropical regions could then be
evaluated. The underlying assumption of my research was that the life
history of Africanized honey bee populations in South America (as well
as the parental population in Africa) is characterized by a high
reproductive rate. Demographic features expected to be correlated with
this high rate of colony reproduction include short worker bee
development times, small worker bee size, rapid queen development and
maturation, and increased egg laying and brood production rates.
Predictions involving these demographic characteristics led to a series
of experiments that are presented and discussed in the following
chapters.
In addition, the question of reproductive isolation versus
hybridization and differential selection between the two populations in
tropical conditions was experimentally evaluated. Whether there is

7
hybridization or reproductive isolation between Africanized and European
honey bee populations could result in very different scenarios for the
potential impact of Africanized honey bees on North America,
particularly the U.S.A.
Identification of Honey Bees Used in My Research
For the experiments presented here, Africanized honey bee colonies
were established from queens removed from feral colonies in an area in
eastern Venezuela where there were no known European honey bees. They
were identified as Africanized bees primarily by their distinctly
smaller comb cell size as compared with European honey bees.
European honey bees used in the experiments were from commercially
produced queens from three different queen breeders in the U.S.A.
Additional European lines were obtained from the U.S. Department of
Agriculture Bee Research Laboratories in Madison, Wisconsin, and Baton
Rouge, Louisiana. All of these European queens were either naturally
mated or instrumentally inseminated in the U.S.A. and then shipped to
Venezuela.
When and Where Research Was Conducted
All field research with Africanized and European honey bees was
conducted from December 1978 through February 1980 at the Ministerio de
Agricultura y Cria de Venezuela Africanized Honey Bee Research
facilities near Maturin, Monagas. The area originally was a Tropical
Dry Forest [sensu Holdridge Life Zone System (Holdridge 1964; Ewel and
Madriz 1968)]. The forest had been partially cleared, and the area was
grazed by cattle.

8
All Africanized and European honey bee comparisons were made at the
same time under identical experimental conditions. Field and
experimental methods are described for each of the experiments in the
appropriate chapters.
A few experiments with European honey bees only were conducted in
the U.S.A. to confirm techniques developed and used while in Venezuela.
These experiments were undertaken either at the U.S. Department of
Agriculture Bee Breeding and Stock Center Laboratory in Baton Rouge,
Louisiana, or at the bee research facilities of the Institute of Food
and Agricultural Sciences at the University of Florida, Gainesville.

CHAPTER II
WORKER BEE DEVELOPMENT TIMES
Introduction
The presence of both Africanized and European honey bees, Apis
mel1 ifera, in South America allows for comparisons to be made under
identical conditions between a population that has evolved in the
tropics and one that has evolved in temperate regions. Africanized
honey bee populations in South America are reported to have a high
colony reproductive (swarming) rate compared with European honey bee
populations in North America (Otis 1980, 1982a; Seeley 1978; Winston
1979b, 1980a; Winston, Dropkin and Taylor 1981; Winston, Taylor and Otis
1983). However, these investigations have not been conducted under
similar environmental or experimental conditions. Therefore,
comparisons of reproductive rates between Africanized and European honey
bees using these data are inappropriate either for identifying
differences in reproductive rates between tropical and temperate honey
bee populations or for identifying factors responsible for the success
of Africanized bees in tropical regions.
Reproductive rates in honey bees are a function of colony growth
rates which are a result of an interaction of at least three factors:
resource availability, resource utilization efficiency (foraging
success, brood production efficiency, and bee size), and colony
demographic parameters. Worker bee longevity is the only demographic
9

10
parameter that has been compared between Africanized and European bees
under identical conditions. The greater longevity of European honey
bees (Winston and Katz 1981) gives European bees a colony growth rate
advantage. Other demographic characteristics that affect reproductive
rates of Africanized and European honey bees (for example, worker bee
development times, brood mortality, queen development and maturation
periods, queen fecundity and brood production rates) have not been
evaluated for Africanized and European bees under similar conditions.
As part of a larger study evaluating these demographic parameters, this
study compares worker bee development periods for Africanized and
European honey bees in Venezuela.
Smith (1958a) and Tribe and Fletcher (1977) reported that the total
development period (from oviposition to adult emergence) for worker bees
of Apis mellifera adansonii (now classified as &. m, scutellata: Ruttner
1976a, 1976b, 1981) from South Africa was between 18.6-20 days. Similar
development times for the Africanized honey bee populations (descendents
of A. m. scutellata) in Brazil have been presented (Kerr, Goncalves,
Blotta and Maciel 1972; Wiese 1972). Worker bee development times for
European populations (primarily A. m. mel1ifera, 1ioustica, carnica and
caucasica) from Europe and North America range from 20-24 days (Jay
1963).
The differences in development times between African (and
Africanized) and European genotypes, which range from 1.4 to 5.4 days,
are difficult to evaluate because they are based on data collected under
very different experimental conditions. Jay (1963) summarized a number
of factors that affect development times: seasonal variation in
temperature; temperature differences in different areas within the brood

11
nest; colony size, which affects both brood nest temperature and feeding
frequency and quality; and nectar and pollen resources, which also
affect feeding quantity and quality. Valid comparison of development
times between genotypes or populations can only be made when these
factors are controlled under similar experimental conditions.
The importance of slight temperature differences on development
time cannot be overstated. Development times for European worker bees
in Wisconsin, U.S.A., averaged 20.5 days but ranged from 20-24 days,
depending on differences in temperature in different areas of the brood
nest (Milum 1930). Harbo and Bolten (1981) showed that fertilized eggs
kept at 34.8C hatched about 1.4 hours sooner than those kept at 34.3C.
This difference in egg hatch time for only a 0.5C difference in
temperature can be extrapolated to approximately 10 hours for the entire
development period [calculated from Harbo and Bolten (1981)]. However,
normal temperatures within a brood nest can vary to a much greater
extent (Milum 1930; Jay 1963). For example, when Tribe and Fletcher
(1977) determined the development rates of African honey bees in South
Africa, they recorded that temperatures in the brood nest varied from
26-34C.
In an incubator with controlled temperature and humidity, eggs from
Africanized genotypes hatched significantly sooner than eggs from
European genotypes, 69.6 + 1.06 hours compared with 73.3 + 1.14 hours
(Harbo, Bolten, Rinderer and Collins 1981). Egg development requires
that only temperature and humidity be controlled and can therefore be
evaluated independently of colony-level parameters. However,
differences between Africanized and European genotypes for total worker
bee development periods need to be evaluated within a colony in order to

12
allow for normal feeding and growth. Worker bee development rates are a
result of an interaction between the egg genotype and the colony. There
are three colony-level factors that need to be considered when comparing
total development time of Africanized and European worker bees.
First is the effect of comb cell size. There is a difference in
natural comb cell size between Africanized and European populations.
The width between opposite sides of the hexagonal cells for the African
population in Africa measured 4.77-4.94 mm (Smith 1958a). Cells for the
Africanized population in Brazil averaged 5.0 mm (range 4.8-5.4 mm)
(Michener 1972), but cells of the Africanized population in Venezuela
averaged 4.8 mm (range 4.5-5.0 mm) (Chapter III; Rinderer, Tucker and
Collins 1982). Cells from the European population from Ontario, Canada,
averaged 5.4 mm (range 5.2-5.7 mm) (Michener 1972), and those from
Louisiana, U.S.A., averaged 5.2-5.3 mm (range 5.2-5.4 mm) (Rinderer,
Tucker and Collins 1982). Adult bee size is a function of comb cell
size (Grout 1937); adult Africanized bees are smaller than European bees
(62 mg compared with 93 mg, unengorged) (Otis 1982b; Otis, Winston and
Taylor 1981).
Abdellatif (1965) suggested that larvae in smaller comb cells
received less food which caused them to elongate and become sealed
earlier. Also, Tribe and Fletcher (1977) suggested that the difference
in development time for African and European genotypes may be a function
of the small African bee size. Therefore, the effect of comb cell size
needs to be considered when comparing development times of Africanized
and European honey bees.
The second colony-level factor is the effect of nurse bee genotype.
There may be behavioral differences and/or physiological differences in

13
the way In which nurse bees from the two populations interact with the
developing larvae. Mel'nichenko (1962) suggested that differences
between nurse bee genotypes might affect developmental rates as well as
size of developing larvae. For European honey bees, Lindauer (1953)
calculated that each developing larva requires over 2785 adult bee
visits taking a total of 10.3 hours. This appears to provide sufficient
opportunity for possible genotype differences, either quantitative or
qualitative, to affect development rates. In addition to potential
qualitative or quantitative differences in feeding of larvae, nurse bees
of different genotypes may also maintain different brood nest
temperatures. Therefore, development times for Africanized and European
worker bees were evaluated in both Africanized and European colonies.
The third colony-level factor affecting worker development is
colony size (number of worker bees in a colony). Colony size affects
both brood nest temperature and larval feeding rates, which, as already
discussed, are two major factors affecting development times.
In addition to these colony-level parameters, resource conditions
also affect development time. Nelson and Sturtevant (1924) reported
that development of European bees was more rapid with increased larval
feeding associated with a nectar flow. Ribbands (1953) and Jay (1963)
both summarized evidence of the effect of food on larval development
rates. Therefore, all comparisons of worker development times were
conducted simultaneously to avoid any differences due to resource
conditions.
This paper reports the results from a comparison of the development
times of Africanized and European honey bees under identical conditions
in Venezuela. The experimental design allowed for the discrimination

14
between the effects of egg genotype and the colony-level parameters of
comb cell size and nurse bee genotype on worker bee development time.
These experiments were conducted during July-October 1979.
Methods.
Table 2-1 summarizes the experimental design. Four experimental
colony treatments were established as follows:
i.Africanized comb cell size Africanized nurse bees (A55)
ii.Africanized comb cell size European nurse bees (H2)
iii.European comb cell size, Africanized nurse bees (A41)
iv.European comb cell size, European nurse bees (IBR877).
Each experimental colony was a five-frame hive (22 liters) with
four empty combs and one comb with honey and pollen and approximately 2
kg of young adult bees (Africanized or European, depending upon
treatment). Because natural nectar and pollen resources were available
irregularly throughout the experimental period (16 weeks), the colonies
were supplemented with honey and pollen as necessary.
European comb was built from commercially-produced beeswax
foundation that had been fastened into standard wooden frames.
Africanized comb was naturally built (not from foundation) by
Africanized bees in empty standard wooden frames to facilitate
manipulation and colony inspection.
The queens in colonies with Africanized nurse bees (A55 and A41)
were Africanized queens produced by standard queen rearing methods
(Laidlaw 1979) and then naturally mated to Africanized drones. Mating
occurred in an area of eastern Venezuela that had a large feral
population of Africanized honey bees with no known European honey bees
present (near San Jose de Buja, Monagas, Venezuela). The feral colonies

15
from which the Africanized queen mothers were extracted were also from
this area. The colonies were identified as Africanized honey bees by
both their behavior and their small comb cell size characteristic of the
Africanized population (4.5-5.0 mm, see Chapter III).
The queens in the colonies with European nurse bees (H2 and IBR877)
were European queens that had been mated to European drones in the
U.S.A. and transported to Venezuela. Line H2 was from a commerical
queen producer in the southeastern U.S.A.; IBR877 was an outbreed line
from the U.S. Department of Agriculture Bee Breeding and Stock Center
Laboratory in Baton Rouge, Louisiana, U.S.A.
The source for the Africanized egg genotype (A26) was a queen
removed from a feral colony of Africanized honey bees in the San Jose de
Buja area. The colony was identified as Africanized by its behavior and
characteristic comb cell size. The source of the European egg genotype
(Y5) was a queen commercially produced in the southeastern U.S.A. and
shipped to Venezuela.
Because adult longevity is from 2 to 5 weeks for European honey
bees (Woyke 1984) and 2 to 3 weeks (or less) for Africanized honey bees
(Winston 1979bj Winston and Katz 1981), experimental colonies were
established 10 weeks prior to the start of the experiment. This was
sufficient time to insure that at the beginning of the developmental
trial all the adult bees present within the experimental colonies had
developed in those colonies, and, therefore, were offspring of a known
genetic line having developed within a known comb cell size. Before
development times were measured, the worker bee populations in the
experimental colonies were equalized as much as possible by removing
random samples of bees from the most populous colonies.

16
The experimental colonies were placed in an apiary under a roof
with completely open sides. The roof served two purposes. First the
colonies were in complete shade which reduced any effects from
differences in ambient temperature and sunlight. Second, colonies could
be opened and inspected in order to monitor development with a minimum
of disturbance, especially during rain.
Eggs were collected from queens of the two designated egg source
lines (A26 and Y5), that were established in modified colonies similar
to those used by queen producers in the U.S.A. (Harp 1973). These
colonies consisted of five standard frames with the middle frame
isolated from the four others by queen excluder side and top panels.
The excluder panels have a mesh size that restricts the queen from
passing through because of her wider thorax, but allows worker bees to
pass through to feed and communicate with the queen. Thus, the queen
was isolated on a specific comb so that eggs could be collected that
would then be placed into one of the four experimental colonies to
evaluate development time.
The comb used had the appropriate comb cell size for the
experimental colony into which it would be placed. Comb cell size was
measured in each experimental nurse bee colony and for each Africanized
and European egg comb put into each experimental colony.
The queens were caged on each comb for 24 hours so that a large,
uniform egg sample could be collected. Eggs were monitored only from
the center of each frame, which insured a more uniform temperature
during development as well as uniform brood nest position. The large
egg sample also insured that the monitored eggs were in a normal
environment, surrounded by similarly-aged developing bees. The thirty

17
or forty eggs selected to be monitored for development formed either a
3x10 or a 4x10 cell area. A reference point which facilitated locating
the designated development sample was indicated by colored pins inserted
five cells to the left of each original egg row.
The two combs (one comb from each egg source) were placed in the
center of each experimental colony at the same time. The cells with the
eggs to be monitored faced each other in order to reduce any effects of
brood nest position. Two frames of brood were removed from each
experimental colony to make room for the experimental frames. This also
reduced the amount of brood being reared in each experimental colony,
insuring that the monitored eggs would be optimally fed.
The pairs of frames were put into the four experimental colonies on
four successive days because of the 24 hours needed to collect each set
of eggs. Once the eggs were put into the experimental colonies, they
were inspected every day at 0800 hours. The survivorship and
developmental status of each original test group egg was recorded.
The sample size of eggs monitored was selected to minimize the time
each colony would need to be opened for observation in order to minimize
disturbance. When a colony was disturbed, bees flew from the comb and
ran on the bottom of the hive, resulting in temperature fluctuations and
interrupted feeding of larvae. Colonies were carefully opened, using
minimal amounts of smoke. Adult bees were not shaken off the combs but
rather gently pushed aside to observe the development stage within the
cells. Inspections more frequent than every 24 hours also increased the
level of disturbance, especially in the Africanized colonies (A55 and
A4I). The advantage of more frequent monitoring to more accurately

18
record time of developmental changes was outweighed by the negative
effects of disturbance on development rates.
Temperature of the brood nest in the space between the two
monitored combs was recorded periodically Ijy placing a thermometer into
that area through a hole in the hive cover. This was only an
approximate measure of temperature because development occurs within the
cells where temperature is less affected by the ventilating air currents
within the hive.
Results
Table 2-2 presents the development times for unsealed brood, sealed
brood, and total development times from oviposition to adult emergence
for Africanized and European egg genotypes in each of the four
experimental treatments. Table 2-3 summarizes the argumentation for
evaluating the interactions of egg genotypes with the colony-level
parameters of comb cell size and nurse bee genotype on worker bee
development times. The best tests to use to compare differences between
the Africanized and European egg genotypes are evaluating pairs in each
of the four experimental nurse bee colonies (A55, H2, A41, and IBR877),
i.e., AxB, CxD, ExF, and GxH. In these comparisons, the colony-level
parameters (colony population, position within the brood nest,
temperature, comb cell size, and nurse bee genotype) are identical and
allow for only differences in egg genotype to be compared. Tables 2-4,
2-5 and 2-6 summarize the results from the statistical analyses for the
unsealed brood, sealed brood, and total development times, respectively.
Data were analyzed with the Kolmogorov-Smirnov one-tailed test using the
chi square distribution, df = 2 (Siegel 1956).

19
Africanized worker bees developed faster than European bees (ACEG x
BDFH). The unsealed larval period was 4.3 +0.4 days compared with 4.9
+0.4 days, PcO.OOl; the sealed larval and pupal period was 11.6 +0.5
days compared with 11.9 + 0.4 days, P<0.01; and the total development
time was 18.9 + 0.3 days compared with 19.8 + 0.4 days, P<0.001. There
was no significant effect of comb cell size or nurse bee genotype on
development times. These differences in total development time are
similar to the differences found between three different lines of
Africanized and three different lines of European honey bees compared in
another study (19.2 days compared with 20.0 days, Table A1).
Comb cell sizes for the experimental colonies and egg sample frames
are presented in Table 2-7. Temperatures recorded for all experimental
colonies varied from 35-36C.
When differences in development times between Africanized and
European egg genotypes were compared for each stage of development, the
greatest difference was observed in the unsealed larval stage (Table 2-
8). However, this was not a result of a differential acceleration of
development during the unsealed larval period for the Africanized honey
bees. The proportion of unsealed larval development time to total
development time and the proportion of sealed brood development time to
total development time were compared for the Africanized and European
honey bee populations following angular transformations of the
proportions (Sokal and Rohlf 1969). These proportions were not
significantly different between the Africanized and European honey bees.
The differences recorded for the unsealed brood stage between the
Africanized and European honey bees may be an artifact of the experiment
for two reasons. First, the 24-hour observation interval may obscure

20
exact timing of developmental changes. Second, the process of sealing
is not a precise developmental stage and may take from six hours
(Lindauer 1953) to 24 hours (Jay 1963). When the unsealed and sealed
brood stages are combined, the proportional differences between the two
populations are the same as for egg development times and total
development times (Table 2-8).
Mortality for different developmental stages for each colony
treatment is presented in Table 2-9 (see also Table A-2). Mortality was
high (26-37%) for larvae in the experimental colony (H2) with European
nurse bees on Africanized comb cell size. The high mortality during the
larval stage may be a result of the reduced ability of larger European
nurse bees to feed the developing larvae in smaller Africanized comb
cells. There was also a high egg mortality recorded for European eggs
in A41 and IBR87734 and 70%, respectively. Woyke (1977) reports
normal mortality may be as high as 10-50% depending on the season.
Garofalo (1977) also reports varying mortalities depending on both the
size of the colony and the time of year: eggs 10-25%, larvae 11-37%,
pupae 5-7%, and all developmental stages combined 25-53%.
Discussion
This study is the first to evaluate worker bee development times
between Africanized and European honey bees as an interaction between
egg genotype and colony-level parameters. Differences in worker bee
development times were independent of the colony-level parameters of
comb cell size and nurse bee genotype but were dependent on egg genotype
differences between the Africanized and European populations. The
difference in development times between these two populations was not as

21
large as expected from previous reports, which underscores the
importance of making comparisons under identical conditions.
The proportional difference (5.7%) in egg development times between
Africanized (A26) and European (Y5) honey bees reported by Harbo,
Bolten, Rinderer and Collins (1981) is identical to the proportional
difference in total development time reported in the present study
(5.7%, see Table 2-8). Egg development time is a function of the
inherent rate characteristic of the particular genotype because colony-
level parameters (e.g., feeding) are not involved (Harbo, Bolten,
Rinderer and Collins 1981). Using egg development to evaluate
differences in total development between genotypes (or populations) is
advantageous because egg development times are easier to evaluate, take
less time, have fewer variables to control (temperature and humidity
only), and can be evaluated in an incubator rather than in a colony,
avoiding problems associated with disturbing the colony during
observations. It must be noted, however, that by using egg development
times one can only extrapolate proportional differences between
genotypes for total development time but cannot extrapolate the absolute
total development time.
A prerequisite for high reproductive rates would be a rapid colony
growth rate. However, the importance of worker development time to the
rate of colony growth (increase in numbers of bees in a colony) has
apparently been misunderstood, e.g., see Fletcher (1977a, 1978),
Fletcher and Tribe (1977a), Tribe and Fletcher (1977), Winston (1979b),
Winston, Dropkin and Taylor (1981), Winston and Katz (1982), Winston,
Taylor and Otis (1983). The difference in worker development times
observed for Africanized and European honey bees is not a factor

22
contributing to either differences in rate of colony population increase
or to differences in reproductive rates between the two honey bee
populations.
The importance attributed to worker development time on the rate of
colony growth may be a result of confusing colony population increase
(increase in the number of bees in the colony) with general population
growth models designed for other species in which all individuals are
potential reproductives. For honey bees, individual (or worker bee)
development time is not equal to generation time. Organism growth
models must be used to evaluate colony growth even though the number of
individual worker bees within the hive increases. The hive is the
organism. Worker bee development time does not affect the rate of
colony growth. Worker development time affects only the length of time
between a given change in egg laying rate and its resulting change in
population increase or decrease. Africanized bees develop in 19 days
and begin their population increase (^growth) on the 19th day of the
colony cycle, compared with the 20th day for European bees. This
difference is trivial compared to potential differences from other
demographic factors that do affect rates of colony growth. Egg laying
and brood production rates, worker bee longevity, brood mortality, and
resource availability are factors that do affect the rate of colony
population increase and, therefore, affect the reproductive rates.
Tribe and Fletcher (1977) have suggested that African worker bees
have a shorter unsealed development stage because they do not grow as
large as European honey bees. They compare their data for African bees
with data for European bees in the literature and conclude that African
bees have a 20-30% shorter unsealed larval stage. There are four

23
problems with their analysis. First as already pointed out using the
duration of the unsealed stage has inherent problems because it is not a
precise development stage. Second comparisons based on data collected
under different experimental conditions are not valid. Third, their
logic is perhaps circular with respect to the question of larval size
and larval development times. In the present study, development time
was not size-related for either Africanized or European honey bees.
Africanized honey bees that developed in European comb had the same
development times as those that developed in Africanized comb even
though Africanized bees reared in European comb were significantly
larger (16%; Chapter III). The same relationship was true for European
honey bees with a 17% increase in size of bees from European comb
compared with bees from Africanized comb. And fourth, their comparison
is in itself incorrect. Rather than compare the differences in unsealed
development times between African and European populations to determine
if the African population has a relatively shorter duration as unsealed
larvae, they should have used the proportion of unsealed development
period to total development period in order to compare African and
European populations. In the present study, the relative times spent as
an unsealed larvae to the total development time for both the
Africanized and European genotypes were not significantly different.
The differences in development time between Africanized and European
populations appear constant throughout development without any
developmental acceleration during the larval stage for either
Africanized or European honey bees.

TABLE 2-1. Experimental matrix for evaluating interaction of egg
genotype, comb cell size, and nurse bee genotype on worker
development times. A H represent each treatment.
AFRICANIZED EUROPEAN
EGG GENOTYPE (A26) EGG GENOTYPE (Y5)
AFRICANIZED COMB CELLS
AFRICANIZED NURSE
BEES (A55) A B
EUROPEAN NURSE
BEES (H2) C D
EUROPEAN COMB CELLS
AFRICANIZED NURSE
BEES (A41) E F
EUROPEAN NURSE
BEES (IBR877) G H

25
TABLE 2-2
Interaction of egg genotype, comb cell size, and nurse
bee genotype on worker bee development time (days):
median, (range), mean + SD, (n = sample size).
AFRICANIZED EGG GENOTYPE (A26)
USa
SBb
TDTC
AFRICANIZED COMB CELLS
AFRICANIZED NURSE
BEES (A55)
4.0
(4-5)
4.2 + 0.4
(n = 30)
12.0
(11-12)
11.6 + 0.5
(n = 30)
19.0
(18-20)
18.8 + 0.5
(n = 30)
EUROPEAN NURSE
BEES (H2)
4.0
(4-5)
4.4 + 0.5
(n = 29)
12.0
(11-12)
11.6 + 0.5
(n = 29)
19.0
(19-20)
19.1 + 0.2
(n = 29)
EUROPEAN COMB CELLS
AFRICANIZED NURSE
BEES (A41)
4.0
(4-5)
4.2 + 0.4
(n = 30)
12.0
(11-12)
11.7 + 0.5
(n = 30)
19.0
(18-19)
18.9 + 0.2
(n = 30)
EUROPEAN NURSE
BEES (IBR877)
4.0
(4-5)
4.3 + 0.5
(n = 26)
12.0
(11-12)
11.7 + 0.5
(n = 26)
19.0
(19)
19.0 + 0
(n = 26)
TOTALS
4.0
(4-5)
4.3 + 0.4
(n = 115)
12.0
(11-12)
11.6 + 0.5
(n = 115)
19.0
(18-20)
18.9 + 0.3
(n = 115)
aUS = unsealed brood (unsealed larval development period only).
bSB = sealed brood (pre-pupae and pupae).
CTDT = total development time (oviposition to adult emergence).

26
TABLE 2-2~extended.
EUROPEAN EGG GENOTYPE (Y5)
US
SB
TDT
5.0
(4-5)
4.9 + 0.3
(n = 37)
12.0
(11-12)
11.8 + 0.4
(n = 37)
20.0
(19-20)
19.6 + 0.5
(n = 37)
5.0
(4-5)
4.9 + 0.4
(n = 22)
12.0
(12)
12.0 + 0
(n = 22)
20.0
(19-20)
19.9 + 0.4
(n = 22)
5.0
(4-6)
5.0 + 0.4
(n = 19)
12.0
(12-13)
12.0 + 0.2
(n = 19)
20.0
(19-21)
20.0 + 0.3
(n = 19)
5.0
(4-6)
4.7 + 0.8
(n = 7)
12.0
(12-13)
12.1 + 0.4
(n = 7)
20.0
(19-21)
19.8 + 0.7
(n = 7)
5.0
(4-6)
4.9 + 0.4
(n = 85)
12.0
(11-13)
11.9 + 0.4
(n = 85)
20.0
(19-21)
19.8 + 0.4
(n = 85)

27
TABLE 2-3. Summary of hypotheses and tests for evaluating development
times; letters represent treatments (see Table 2-1).
HI: Worker bee development is faster for Africanized genotypes than
for European genotypes.
A x B
C x D
E x F
G x H
A x H
AC x BD
EG x FH
AE x BF
CG x DH
ACEG x BDFH
African'ized comb cell size; Africanized nurse bees
Africanized comb cell size; European nurse bees
European comb cell size; Africanized nurse bees
European comb cell size; European nurse bees
Africanized comb cell size and nurse bees compared with
European comb cell size and nurse bees
Africanized comb cell size; both nurse bee genotypes
combined
European comb cell size; both nurse bee genotypes combined
Africanized nurse bees; both comb cell sizes combined
European nurse bees; both comb cell sizes combined
Both comb cell size and both nurse bee genotype variables
combined
H2: Worker bee development is more rapid in Africanized comb cells
than in European comb cells.
A x E Africanized egg genotype; Africanized nurse bees
C x G Africanized egg genotype; European nurse bees
B x F European egg genotype; Africanized nurse bees
D x H European egg genotype; European nurse bees
AC x EG Africanized egg genotype; both nurse bee genotypes
combined
BD x FH European egg genotype; both nurse bee genotypes combined
H3: Worker bee development is more rapid with Africanized nurse bees
than with European nurse bees.
A x C
E x G
B x D
F x H
AE x CG
BF x DH
Africanized egg genotype; Africanized comb cell size
Africanized egg genotype; European comb cell size
European egg genotype; Africanized comb cell size
European egg genotype; European comb cell size
Africanized egg genotype; both comb cell sizes combined
European egg genotype; both comb cell sizes combined

TABLE 2-3continued.
H4: Worker bee development is more rapid with Africanized comb cells
and Africanized nurse bees than with European comb cells and
European nurse bees.
A x G
B x H
Africanized egg genotype
European egg genotype

29
TABLE 2-4. Unsealed brood development times. Hypotheses were tested
using Kolmogorov-Smirnov one-tailed test chi-square
distribution, df = 2, alpha = 0.05 (Siegel 1956).
HI: Worker bee development is faster for Africanized genotypes than for
European genotypes.
A X B ***a
C X D **
E X F ***
G X H NS
A X H NS
AC X BD ***
EG X FH ***
AE X BF ***
CG X DH ***
ACEG X BDFH ***
H2: Worker bee development is more rapid in Africanized comb cells than
in European comb cells.
AXE NS
C X G NS
B X F NS
D X H NS
AC X EG NS
BD X FH NS
H3: Worker bee development is more rapid with Africanized nurse bees
than with European nurse bees.
A X C NS
E X G NS
B X D NS
F X H NS
AE X CG NS
BF X DH NS
H4: Worker bee development is more rapid with Africanized comb cells
and Africanized nurse bees than with European comb cells and
European nurse bees.
A X G NS
B X H NS
a ** = P<0.01
*** = Pco.001.
bAnalysis may be NS because test used is conservative for small sample
sizes using chi-square distribution.

30
TABLE 2-5. Sealed brood development times. Hypotheses were tested
using Kolmogorov-Smirnov one-tailed test chi-square
distribution, df = 2, alpha = 0.05 (Siegel 1956).
HI: Worker bee development is faster for Africanized genotypes than for
European genotypes.
A X B NS
C X D *a
E X F NS
G X H NS
A X H NS
AC X BD *
EG X FH *
AE X BF *
CG X DH **
ACEG X BDF **
H2: Worker bee development is more rapid in Africanized comb cells than
in European comb cells.
AXE NS
C X G NS
B X F NS
D X H NS
AC X EG NS
BD X FH NS
H3: Worker bee development is more rapid with Africanized nurse bees
than with European nurse bees
A X C NS
E X G NS
B X D NS
F X H NS
AE X CG NS
BF X DH NS
H4: Worker bee development is more rapid with Africanized comb cells
and Africanized nurse bees than with European comb cells and
European nurse bees.
A X G NS
B X H NS
a = P<0.05
** = P<0.01.

31
TABLE 2-6. Total worker bee development times. Hypotheses were tested
using Kolmogorov-Smirnov one-tailed test, chi-square
distribution, df = 2, alpha = 0.05 (Siegel 1956).
HI: Worker bee development is faster for Africanized genotypes than for
European genotypes.
A X B
***a
C X D
***
E X F
***
G X H
***
A X H
**
AC X BD
***
EG X FH
***
AE X BF
***
CG X DH
***
ACEG X BDFH
***
H2: Worker bee development is more rapid in Africanized comb cells than
in European comb cells.
AXE NS
C X G NS
B X F NS
D X H NS
AC X EG NS
BD X FH NS
H3: Worker bee development is more rapid with Africanized nurse bees
than with European nurse bees
A X C NS
E X G NS
B X D NS
F X H NS
AE X CG NS
BF X DH NS
H4: Worker bee development is more rapid with Africanized comb cells
and Africanized nurse bees than with European comb cells and
European nurse bees.
A X G NS
B X H NS
a ** = P<0.01
*** = P<0.001.

32
TABLE 2-7. Comb cell size for worker development time experiment: comb
measurements = mm for 10 consecutive, horizontal cells, mean
+ SD, (sample size).
COMB CELL SIZE
NURSE BEE AFRICANIZED EGG EUROPEAN EGG
COLONY GENOTYPE GENOTYPE
AFRICANIZED COMB CELL SIZE3
AFRICANIZED NURSE
BEES (A55)
47.5 + 0.58
(4)
49.8 + 0.50
(4)
EUROPEAN NURSE
BEES (H2)
EUROPEAN COMB CELL SIZE5
48.2 + 0.96
(4)
48.5 + 0.58
(4)
AFRICANIZED NURSE
BEES (A41)
54.0 + 0.0
(3)
54.0 + 0.0
(3)
EUROPEAN NURSE
BEES (IBR877)
53.3 + 0.58
(3)
53.3 + 0.58
(3)
45.8 + 0.50
(4)
48.5 + 0.58
(4)
54.0 + 0.0
(3)
53.7 + 0.58
(3)
3Natural comb built without foundation.
bBuilt from foundation.

TABLE 2-8. Comparison of differences in development times (in days)
for Africanized and European honey bees for different
developmental stages.
DEVELOPMENTAL STAGES
EGG HRS
(DAYS)3
UNSEALED
BROODb
SEALED
BROOD0
UNSEALED
& SEALED
TOTAL
DEVELOPMENT
AFRICANIZED EGG
GENOTYPE (A26)
69.6
(2.90)
4.3
11.6
15.9
18.80
EUROPEAN EGG
GENOTYPE (Y5)
73.6
(3.07)
4.9
11.9
16.8
19.87
% DIFFERENCEd
5.7
14.0
2.6
5.7
5.7
aFrom Harbo, Bolten, Rinderer and Collins (1981); data used are
their Africanized #3 = A26 and their European #5 = Y5.
Unsealed larval period only.
^Pre-pupae and pupae.
d% Difference = C(Y5)-(A26)/(A26)] x 100.

34
TABLE 2-9. Mortality during different developmental stages.
AFRICANIZED EGG GENOTYPE (A26) EUROPEAN EGG GENOTYPE (Y5)
E1 E2 l1 L2 SB N
AFRICANIZED COMB
CELL SIZE
AFRICANIZED
NURSE BEES
A55 1 5 0 3 1 40 1 0 0 2 0 40
EUROPEAN
NURSE BEES
H2 1 0 3 7 0 40 1 0 0 13 0 36
EUROPEAN COMB
CELL SIZE
AFRICANIZED .
NURSE BEES
A41 0 0 0 0 0 30 8 2 0 0 0 29
EUROPEAN
NURSE BEES
IBR877 00220 30 13 6100 27
^Mortality during first 24 hours in test colony (acceptance).
^Mortality between 24-72 hours (before hatching).
^Mortality between 72-96 hours (at time of hatching).
Mortality during older larval stages before sealing.
^Mortality during the pupal stage.
fN = total eggs monitored.

Qj-j^pyfrp jjj
INTERACTION OF MATERNAL GENOTYPE, EGG GENOTYPE AND COMB CELL SIZE ON
HONEY BEE WORKER SIZE AND SIZE VARIATION
Introduction
In the evolution of eusociality in bees (Apoidea), there is a
considerable decrease in size variation of the workers within a colony.
Worker size variation within a colony of primitively eusocial sweat bees
(Halictidae) or bumble bees (Apidae) is much greater than the size
variation of workers within colonies of highly eusocial stingless bees
(Mel iponinae: Mel ipona and Trigonal or honey bees (Apinae: Apis)
(Brian 1952; Kerr and Hebling 1964; Medler 1965; Michener 1974). For
example, the coefficient of variation (CV) for worker weights in a
bumble bee colony may be as high as 31-37% (calculated from Brian 1952
for Bombus aarorum. Table B1) whereas the CV for worker weights within
a honey bee (Apis me!1ifera) colony is only 4-7% (Table B-2).
An effect of the reduction of size variation is that the mechanism
for the division of labor of workers within a colony shifts from being
size dependent to primarily age dependent (Michener 1974). In the
primitively eusocial bumble bees, division of labor is size related
(Brian 1952); large workers may be twice the size (linear measurements)
of small workers within the same colony (Medler 1965). In highly
eusocial stingless bees and honey bees, division of labor is primarily
age dependent (Free 1965; Gary 1975; Kerr and Hebling 1964; Lindauer
1953; Seeley 1982). In honey bees, the workers proceed through a series
35

36
of age-related tasks. However, the sequence and duration of the
different stages are flexible and depend on the needs of the colony.
An advantage of worker size variation within bumble bee colonies
may be efficient utilization of diverse nectar and pollen resources that
may be size dependent. Different sized workers within a colony
specialize on those resources that they can most efficiently exploit
(Heinrich 1979a). However, highly eusocial bees are not at a
disadvantage with respect to resource utilization because they have
evolved complex communication systems that allow foragers to monitor
changing nectar conditions and to recruit workers from the colony to a
particular resource. Therefore, both the species characterized by
workers of highly variable sizes and those species characterized by
uniformly-sized workers have evolved behaviors that enhance the
efficiency of nectar and pollen exploitation.
The difference in intra-colony worker size variation between
primitively eusocial and highly eusocial species of bees is so
significant that Kerr and Hebling postulated that "some controlling
mechanism leads to reduced variances among mature workers [Meliponinae
and Apis], which are therefore of relatively uniform size" (1964, p.
267). Waddington (1981) hypothesized that the evolution and maintenance
of the complex communication systems in Apis, Triaona and Mel ipona
depend upon uniformity of worker bee size within a colony. Differences
in bee size may result in miscommunication because resource
"profitability" may be size dependent. For example, a high quality
resource for a small bee may not be a high quality resource for a larger
bee. However, a regulatory mechanism for reduced size variation has not
been identified.

37
Honey bee size and size variation is a result of both genetic and
environmental factors. Research has focused primarily on the extrinsic
factors that affect bee size (e.g., comb cell size, nutrition and
temperature). Honey bee worker sizes and honey bee comb cell sizes have
been shown to be inter-related: because of the manner by which comb
cells are constructed (Darwin 1859/1958), worker body size affects the
diameter of cells they construct (Baudoux 1933; Glushkov 1958), and
worker bee size is correlated to the size of cells in which they are
reared (Baudoux 1933; Buchner 1955; Glushkov 1958; Grout 1937; Michailov
1927-28 cited in Alpatov 1929; Tuenin 1927).
This interaction between comb cell size and egg genotype may at
first appear to provide a mechanism for both regulating bee size and
reducing size variation among bees within a colony. However, the comb
cell itself can become a source of variation in bee size. Although comb
cell size appears quite uniform, especially when first constructed, the
cells become variable in size as the number of generations reared in
them increases, because pupal cocoons adhere to the cell walls, reducing
cell diameter (Abdellatif 1965; Alpatov 1929; Buchner 1955; Grout 1937).
For example, there is a 25% reduction in cell volume between cells from
new and old combs (Table B-3). Comb cell volume has a greater variance
than cell diameter and is not correlated with diameter (Table B-3).
In addition to comb cell size, there are other extrinsic factors
that affect development and resultant bee size, e.g., quantity and
quality of larval food, and temperature and humidity at which the larvae
and pupae are reared (Buchner 1955; Fyg 1959; Jay 1963; Kulzhinskaya
1956; Michailov 1927-28 cited in Alpatov 1929). These same factors not
only affect absolute size but are sources of size variation.

38
The importance of the genetic component to bee size can be inferred
from the fact that different geographic populations of honey bees differ
with respect to worker bee size (Alpatov 1929; Ruttner 1968, 1975,
1976a, 1976b; Wafa, Rashad and Mazeed 1965). Because honey bee queens
mate with many different drones (Adams, Rothman, Kerr and Paulino 1977;
Peer 1956; Roberts 1944; Taber 1954; Taber and Wendel 1958), the genetic
component becomes an additional factor affecting size variation.
Africanized honey bees in Venezuela (descendents of A. ott..
scutellata) were smaller and had a smaller comb cell diameter (mean 4.8
mm between opposite sides of the hexagonal cells in the comb) compared
with European bees in Venezuela (mean 5.4 mm) (Tables B-3 and B-4;
Rinderer, Tucker and Collins 1982). An opportunity, therefore, existed
to experimentally evaluate the interaction of both genotype and comb
cell size on resultant worker bee size and size variation by studying
both the Africanized and European honey bee populations under identical
experimental conditions. The results from this study provide
information not only on the proximal question involving the factors
affecting bee size but also provide a mechanism by which size variation
may be reduced within a honey bee colony.
Methods
Nine genotypes were evaluated: three Africanized, three European,
and three reciprocal hybrids. The Africanized genotypes (A26, A57,
and B39) were established from queens removed from feral colonies
located in an area in eastern Venezuela with no known European honey
bees. They were identified as Africanized honey bees by their comb cell
sizes which were significantly smaller than European comb cell sizes
(Tables B-3 and B-4; Michener 1972, 1975; Rinderer, Tucker and Collins

39
1982). The European genotypes (YD28 and WEI) were imported into
Venezuela from the U.S. Department of Agriculture Bee Breeding and Stock
Center Laboratory in Baton Rouge, Louisiana, U.S.A., and from a
commercial queen producer from southeastern U.S.A., respectively. Queen
YD28 was artificially inseminated with the spermatozoa from one drone;
queen WEI was naturally mated. The third European genotype (SDY1) was a
daughter from line YK produced by another commercial queen producer from
southeastern U.S.A. and artificially inseminated in Venezuela with a
single drone from the same commercial line.
Two reciprocal hybrid lines were established from artificially
reared queens (Laidlaw 1979) that were instrumentally inseminated with
spermatozoa from single drones: Africanized queen x European drone
(SDA12) and European queen x Africanized drone (SDY10 and SDY11). The
Africanized queen and drone source was A26. The European queen and
drone source was line YK. The hybrid lines were therefore genetically
similar, but were the reciprocal of each other with respect to their
queen and drone sources.
Queens were produced by the standard method of transferring young
larvae from the desired queen line into artificial queen cells which
were then introduced into cell-producing colonies (Laidlaw 1979).
Mature queen cells were put into an incubator (35 + 1C) 72 hours prior
to adult emergence. Newly emerged virgins were marked for individual
identification and then put into individual cages and maintained in a
strong, queenless colony for approximately one week until they were
artificially inseminated.
Drones for instrumental inseminations were produced by caging drone
comb containing sealed drone pupae from the desired drone source lines.

40
As drones emerged, they were placed into special holding cages and
maintained in a colony so that worker bees could feed them until they
matured. This manipulation insured that the drones used for
inseminations were from the desired queen lines.
To collect eggs for the bee size experiments, queens from the nine
genotypes were confined for five hours in their own colonies to a
section of Africanized comb (mean cell size = 4.8 mm), using 8 x 8 cm
push-in cages. These cages were made from 3 mm mesh hardware cloth and
had queen excluder material soldered to the top to enable worker bees to
pass through to tend the queen (Harbo, Bolten, Rinderer and Collins
1981). After five hours, the queens were removed from the combs. The 8
x 8 cm sections of comb with eggs from each queen were cut out and
fitted into special frames. The nine sections were then placed in a
strong Africanized colony (Africanized nurse bees and Africanized comb
cell size) for development. The following day, eggs were collected in
European combs (mean cell size = 5.4 mm) using the same procedure with
the same nine queens except that the nine sections were put into a
European colony (European nurse bees and European comb cell size) for
development. Having all nine egg sources for each comb cell size
treatment (Africanized or European) develop in the same colony
controlled for additional variables affecting development and bee size:
temperature and humidity, nurse bee genotypes and colony size (see
Chapter II).
Fresh pupal weights were compared for each of the nine genotypes
reared in both Africanized and European comb cell sizes. Pupal weights
were measured on the 16th day after oviposition. This age corresponds
to the period during pupal development of least weight change (Melampy

41
and Willis 1939). This was confirmed for fresh pupal weights by
weighing a sample of pupae every 24 hours from day 11.5 post oviposition
to 17.5 days post oviposition (Table B-5). Although Africanized bees
develop one day faster than European bees (Chapter II), pupal weights
can be compared because there is no significant difference in weights
between adjacent days during this period of pupal development (Table B-
5).
Pupal weights were used instead of adult weights in order to reduce
variation resulting from differences in food engorgement and/or feces
accumulation. Pupae were carefully removed from their comb cells by
first removing the cappings and then spreading the cell walls with a
forceps in order that the pupae could easily be removed without
rupturing. Weights (to 1.0 mg) were recorded using Mettler Type H4 and
H6 balances. Comb cell diameters were determined by measuring ten
adjacent cells; three sets of measurements were made from each comb.
Results
Table 3-1 presents the experimental design matrix. The interaction
of egg genotype and comb cell size on worker bee pupal weights for each
of the nine genotypes is summarized in Table 3-2. Table 3-3 presents
the results of the statistical analyses. When Africanized and European
genotypes are reared simultaneously in the same colony (same comb cell
size, nurse bee genotype, temperature and humidity, and colony size),
the weights of the worker bees produced are different. Africanized bee
pupae (111.1 +7.6 mg) that developed in Africanized comb cells were
smaller than European bee pupae (123.3 +6.3 mg) that also developed in
Africanized comb cells (ACE x MOO, P<0.001). When worker bees of
European genotypes are reared in Africanized comb cells, the cells are

42
sealed with strongly convex cappings similar to the way cells containing
drones are sealed in order to accomodate their larger size. Africanized
bee pupae (123.8 +6.2 mg) that developed in European comb cells were
smaller than European bee pupae (139.5 + 54 mg) that also developed in
European comb cells (BDF x NPR, P<0.001). For each of the nine
genotypes investigated, worker bee pupae that developed in Africanized
comb cells were smaller than pupae that developed in European comb
cells, PcO.OOl. There is a 43% increase in comb cell volume between
Africanized and European combs (Table B-3), but the Africanized and
European genotypes only increased in pupal weight by 11.4% and 13.1%,
respectively (Table 3-2). These results show that both genotype and
comb cell size affect worker bee size.
Table 3-4 presents the results for the pupal weights of the
reciprocal F^ hybrids and their respective maternal lines. Data from
only European comb cells were used in order to observe genotype effects
without the constraint of the small Africanized comb cells on European
genotypes. Table 3-5 summarizes the results of the statistical
analyses. The pupal weights of the hybrids from this reciprocal F^
cross were significantly different from each other (H x J; H x L;
P<0.001), but were the same as their respective maternal line (B x H; J
x R; L x R).
Discussion
Reduction of Bee Size Variation
Bee size is a result of not only the interaction of egg genotype
and comb cell size but also the maternal genotype. This can be seen by
evaluating the reciprocal hybrid crosses. The genotype component for
bee size is not a result of "simple" inheritance because pupal weights

43
of genetically similar, reciprocal hybrids are not the same. Because
reciprocal hybrids are phenotypically different from each other, but
phenotypically similar to their maternal lines, maternal genotype must
interact with cell size and egg genotype to determine pupal weight.
This is the first character in honey bees that has been shown to be
influenced by maternal inheritance. Other genetic mechanisms cannot
explain these results. The mechanism for maternal inheritance in worker
size may be through egg size, which has been shown to be inherited
(Roberts and Taber 1965; Taber and Roberts 1963).
Alies (1961) and Mel'nichenko (1962) suggested that differences
between nurse bee genotypes might affect size of developing larvae.
However, McGregor (1938) found that bee size was not affected by nurse
bee genotype. In the experiments presented in this chapter, pupae from
each of the genotypes were reared simultaneously in the same colony for
each comb size treatment. Therefore, differences between the pupal
weights of Africanized and European genotypes cannot be attributed to
either nurse bee differences, cell size, or temperature but must be a
result of both egg and maternal genotype differences.
The importance of maternal inheritance on bee size is that it
reduces worker bee size variation within a colony. If maternal
inheritance were not operating, worker bees of different sizes would be
produced within a colony because of cell size differences and genotype
differences. The effectiveness of maternal inheritance for reducing bee
size variation can be demonstrated by comparing the degree of variation
for the two parameters of bee size (comb cell volume and genotype) with
the degree of worker bee size variation. Abdel 1 at if (1965) showed that

44
when comb cell size variation increased 300%, bee size variation
increased only 50%.
The genetic variation of worker bees within a colony is great
because queens mate on the average with as many as 17 drones (Adams,
Rothman, Kerr and Paulino 1977). There is some degree of mixing of
spermatozoa in the spermatheca resulting in spermatozoa from at least 5
to 6 drones being used during one time interval (Page and Metcalf 1982).
Evidence that maternal inheritance reduces size variation in genetically
diverse worker offspring can be demonstrated by evaluating the size
variation of offspring from single-drone and multiple-drone inseminated
queens. The progeny of queens that were inseminated by spermatozoa from
single drones (SDA12, SDY10, SDY11, SDY1 and YD28) were expected to be
less variable than multiply-inseminated queens (A26, A57, B39 and WEI)
because all eggs from the former queens would have been fertilized by a
genetically identical male gamete. (Drones are haploid; all spermatozoa
are produced by mitosis and are therefore genetically identical.)
Evaluating the coefficient of variation (CV) for each treatment of
genotype and comb cell size, there is no difference between the
variation of progeny from single-drone inseminations versus those from
multiple inseminations, as shown in Table 3-6 (Mann-Whitney U test, one-
tailed, alpha = 0.05).
Additional evidence of maternal inheritance reducing size variation
in genetically heterogeneous offspring comes from analyzing the results
of the reciprocal cross. Because of the influence of maternal
inheritance, subspecific differences in size between Africanized and
European populations were not reflected in increased size variation of
the hybrids compared with the parental types (Table 3-4).

45
Further evidence of the effectiveness of maternal inheritance
reducing size variation can be seen by comparing the size variation
within a colony to the size variation within a population. Alpatov
(1929) found that within honey bee colonies, worker size variation
(e.g., for tongue length) was less than the variation for the local
population of a managed apiary. For seven different apiaries in Russia,
each apiary had an average 22.3% (range 5-42%) increased variation over
the mean colony variation within the apiary. Although the genetic
homogeneity of the apiaries is artificially high as a result of
management practices of the beekeepers compared with the variation of
natural populations of animals (Alpatov 1929), with in-colony variation
was still noticeably reduced.
Evolution of complex communication systems in highly eusocial
species may be responsible for selection for reduced size variation
(Waddington 1981; Waddington, Herbst and Roubik 1986). Foragers within
honey bee colonies have the ability to communicate information to nest
mates about the direction, distance and "profitability" of new resources
(von Frisch 1967) which may be interpreted correctly only if worker bees
within the colony are the same size (Waddington 1981). Profitability of
the resource may be size-dependent as Waddington (1981) suggested. That
is, a high quality resource for a small bee may not be a high quality
resource for a larger bee.
In addition to the profitablity component, correct interpretation
of the distance component of the honey bee waggle dance (von Frisch
1967; Wenner 1962) may also be size-dependent. Different distance
dialects occur not only between subspecies (Boch 1957; Gould 1982) but
also between colonies (Esch 1978 cited in Gould 1982). There is greater

46
variation in individual dialects in colonies that are genetically
heterogeneous compared with colonies that are genetically homogeneous
(Gould 1982). Variation in bee size within a colony may accentuate
differences in distance dialects and increase the possibility of
miscommunication. Therefore, worker size variation within a colony of
honey bees needs to be reduced in order for a communication system that
recruits foragers to a particular floral resource to function correctly
and efficiently with respect to either the profitability (Waddington
1981; Waddington, Herbst and Roubik 1986) or distance component.
Maternal effects operate to reduce bee size variation within a colony of
honey bees, thereby allowing their communication system to function
effectively.
Africanized and European Honev Bee Size Difference
Several hypotheses have been suggested to explain the smaller
worker bee size of the Africanized population. One advantage suggested
for smaller size is more rapid development times, permitting more rapid
colony growth resulting in increased reproductive swarming (Fletcher
1977a; Fletcher and Tribe 1977a; Tribe and Fletcher 1977). However,
cell size and bee size do not affect development times, and, in
addition, worker development times do not affect colony growth rates
(Chapter II).
Fletcher and Tribe (1977a) and Tribe and Fletcher (1977) suggested
that smaller bee size would permit greater numbers of worker bees to be
reared on the same amount of food compared with larger bees. Advantages
of increased worker numbers include frequency of reproductive swarming,
colony defense and foraging success (Wilson 1971). Thus, smaller,
individual bee size maximizes the use of the limited food that

47
characterizes the unreliable nectar availability in Africa (Tribe and
Fletcher 1977). Smaller bee size increases the resource utilization
efficiency of Africanized honey bees and may be a factor in the success
and high reproductive rates of Africanized honey bees compared with
European honey bees in tropical areas of South America (see Chapter
VIII).
I suggest two other hypotheses to explain the advantages of smaller
size in the Africanized population. First smaller size is more
efficient with respect to dissipating heat loads in tropical habitats
(see also Heinrich 1979b). Fletcher (1978) reports that foraging may
stop during the hottest part of the day, which would avoid the
disadvantages of smaller size with respect to gaining a heat load. The
sizes of two other subspecies of honey bees in Africa support this
hypothesis. One of the smallest subspecies in Africa A. oi. 1 itorea, is
found in a very hot and dry area along the coast of Kenya and Tanzania.
One of the largest subspecies, A. m. monticola, is found at higher
elevations and colder temperatures on Mount Kenya.
Secondly, the advantage of smaller bee size may actually lie with
the advantages of smaller cell size. For a given nest cavity volume, a
larger number of worker bees can be produced if cell sizes are smaller.
There is approximately a 25% increase in the number cells for a given
comb area with smaller Africanized comb cells compared with larger
European comb cells. Considering the advantages of increased worker
numbers in a colony (Wilson 1971), the increase in worker numbers as a
result of smaller cell size may be important, particularly if nest
cavity volumes are limited.

48
Because maternal inheritance affects bee size, methods that use
components of size to identify Africanized bees, e.g., morphometric
analysis, may be invalid. Offspring from the cross of a European queen
x Africanized drone (SDY10 and SDY11) are the same weight as offspring
from European queen x European drone (SDY1) (Table 3-4). More
importantly, the offspring from the cross of a European queen x
Africanized drone are significantly different from offspring from
Africanized queen x Africanized drone and Africanized queen x European
drone matings. The European queen x Africanized drone mating represents
the most probable scenario for initial hybridization in North America
(see Chapter VII). That is, a virgin queen from a managed or a feral
European colony mates with Africanized drones and produces offspring
with a 50% Africanized genome. Analyzing the offspring using size as a
component for identification may result in a false negative
identification of Africanized bees. The extent of the problem would
depend upon the degree to which particular linear measurements are
either affected by maternal inheritance and/or are correlated with bee
weight. As Daly, Hoelmer, Norman and Allen (1982) point out, there is a
'difficulty in using phenotype characters to identify genetically
different, but closely related populations" (p. 593).

TABLE 3-1. Effect of comb cell size and egg genotype
on bee pupal weights. Experimental design
matrix (code letters A-R used in tables of
statistical analyses).
COMB CELL SIZE
EGG GENOTYPES AFRICANIZED EUROPEAN
AFRICANIZED QUEEN X
AFRICANIZED DRONE
A26a
A57a
B39a
A
C
E
B
D
F
AFRICANIZED QUEEN X
EUROPEAN DRONE
SDA12b G H
EUROPEAN QUEEN X
AFRICANIZED DRONE
SDY10b I J
SDYllb K L
EUROPEAN QUEEN X
EUROPEAN DRONE
YD28b
WEla
SDYlb
M
0
Q
N
P
R
aNatural matings, multiple inseminations
bSingle drone insemination.

TABLE 3-2. Effect of comb cell size and egg genotype on bee pupal
weights (mg). Means + SD, (sample size).
COMB CELL SIZE
EGG GENOTYPES AFRICANIZED3 EUROPEAN5 % INCREASE
AFRICANIZED QUEEN X
AFRICANIZED DRONE
A26c
A57c
B39c
COMBINED
AFRICANIZED QUEEN X
EUROPEAN DRONE
SDA12d
EUROPEAN QUEEN X
AFRICANIZED DRONE
SDY10d
SDYIld
COMBINED
EUROPEAN QUEEN X
EUROPEAN DRONE
YD28d
WE1C
SDYld
COMBINED
105.4 +4.6 121.7 +5.5 15.5
(60) (80)
117.2+6.8 128.6+4.4 9.7
(30) (40)
116.5 +3.6 123.1 +6.3 5.7
(30) (40)
111.1+7.6 123.8+6.2 11.4
(120) (160)
112.9+4.4 122.4+4.4 8.4
(30) (40)
114.6 +3.7 133.7 +3.3 16.7
(10) (23)
115.5 +3.2 138.9 +4.9 20.2
(30) (30)
115.3 +3.3 136.7 +5.0 18.6
(40) (53)
126.8+4.2 138.7+3.5 9.4
(30) (30)
125.1 + 4.3 143.3 + 2.9 14.5
(30) (30)
115.4+4.2 135.0+6.9 17.0
(20) (19)
123.3 + 6.3 139.5 +5.4 13.1
(80) (79)
3Width between opposite sides of the hexagonal cell is 4.8 mm.
bWidth between opposite sides of the hexagonal cell is 5.4 mm.
'"Natural matings, multiple inseminations.
dSingle drone insemination.

51
TABLE 3-3. Worker bee size: hypotheses and analyses (Mann-
Whitney U test, one-tailed, alpha = 0.05).
Letters refer to experimental treatments, see
Table 3-1.
HI: Africanized bee pupae are smaller than European bee
pupae independent of comb cell size.
ACE
X
MOQ
***a
BDF
X
NPR
***
ACE
X
N3 R
***
BDF
X
MOQ
NS
H2: For a given egg genotype, pupae that develop in
Africanized comb cell size are smaller than pupae
that develop in European comb cell size.
A x
B
***
C x
D
***
E x
F
***
G x
H
***
I X
J
***
K x
L
***
M x
N
***
0 x
P
***
Q x
R
***
ACE
x BDF
***
MOQ
x rPR
***
a*** = p
52
TABLE 3-4. Reciprocal F-, cross. Pupal weights (mg)> mean + SD,
(sample size). Data are from European comb cell size only.
AFRa QUEEN
AFR QUEEN
EUR'
a QUEEN
EUR QUEEN
X
X
X
X
AFR DRONE0
EUR DRONE0
AFR
DRONE0
EUR DRONE*
(A26)
(SDA12)
(SDY10)
(SDY11)
(SDY1)
.21.7 +5.5
122.4 + 4.4
133.7 +3.3
138.9 + 4.9
135.0 + 6.<
(80)
(40)
(23)
(30)
(19)
B
H
J
L
R
aAFR = Africanized; EUR = European.
Naturally mated.
Single-drone, artificial insemination.

53
TABLE 3-5. Maternal effect: hypotheses and analyses (Mann-
Whitney U test, one-tailed, alpha = 0.05).
Letters refer to experimental treatments, see
Table 3-4. The analyses of the following
hypotheses (a postiori) demonstrate that the pupal
weights of hybrids from a reciprocal cross are
different from each other (H4) but are the same as
their respective queen mothers (H2 and H6).
HI:
B
<
R
***a
H2:
B
<
H
NS
H3:
B
<
J
***
B
<
L
***
H4:
H
<
J
***
H
<
L
***
H5:
H
<
R
***
H6:
J
<
R
NS
L
<
R
NS
a*** = P<0.001.

TABLE 3-6. Coefficients of variation for pupal weights from
artificial, single drone inseminations and
natural, multiple matings.
COMB CELL SIZE
EGG GENOTYPE3 AFRICANIZED EUROPEAN
SINGLE DRONE INSEMINATIONS
SDA12
3.9
3.6
SDY10
3.2
2.5
SDY11
2.8
3.5
SDY1
3.6
5.1
YD28
3.3
2.5
MULTIPLE INSEMINATIONS
A26
4.4
4.5
A57
5.8
3.4
B39
3.1
5.1
WEI
3.4
2.0
ANALYSES5
NS
NS
3See Table 3-1 for explanation of genotypes.
Mann-Wh itney U test, one-tailed, alpha = 0.05

CHAPTER IV
QUEEN DEVELOPMENT AND MATURATION
Introduction
African honey bees, Apis mel1ifera scutellata (formerly classified
as adansonii; Ruttner 1976a, 1976b, 1981), were introduced into
southeastern Brazil in 1956 (Kerr 1967; Michener 1975; Woyke 1969). The
following year, swarms escaped and hybridized with the established
European honey bees (primarily A. m. 1iaustica and mellifera) that had
been introduced by 1845 (Gerstaker cited in Pellet 1938; Woyke 1969).
The descendents from this hybridization are known as Africanized honey
bees (Goncalves 1982).
Africanized honey bees in South America have a very high annual
reproductive rate compared with European honey bees in temperate
regions. Based on demographic data collected in French Guiana, the net
reproductive rate for Africanized bees is estimated to be 16 colonies
per colony per year (Otis 1980, 1982a). In comparison, the annual rate
determined for European honey bees in North America was 0.92-0.96
(Seeley 1978) or 3-3.6 when afterswarms are considered (Winston 1980a;
Winston, Taylor and Otis 1983). This dramatic difference in
reproduction between these two honey bee populations may be a result of
length of time throughout the year that resources are available in the
tropics compared with temperate regions (see Chapter VIII) and/or
55

56
demographic characteristics of Africanized honey bees that account for
high reproductive rates.
The reproductive rate of Africanized honey bees results in a swarm-
to-swarm interval of approximately 90 days (Winston 1979b). During that
period a virgin queen emerges, develops pheromones necessary to attract
drones, and mates; ovarian follicles mature; oviposition is initiated;
and the colony population growth period begins prior to the next
swarming. One expected demographic feature for a population with a high
reproductive rate would be a short queen maturation interval (Fletcher
1977a). For the Africanized queens in French Guiana, the maturation
interval from pupal eclosin to initiation of oviposition was 9.7 days
(Otis 1980), over 10% of their swarm-to-swarm interval (calculated from
Winston 1979b). Fletcher and Tribe (1977b) report that in the parental
African population, oviposition begins on the 8th to 9th day after queen
emergence. European queens begin ovipositing between the 6th and 17th
day after emergence (Laidlaw and Eckert 1962; Oertel 1940; Root 1947).
Otis (1980) calculated that the mean interval from pupal eclosin to
oviposition for European queens (10.7 days) was not significantly
different from that of Africanized queens (9.7 days). However,
comparisons between reported values for both Africanized and European
honey bees are inappropriate because the data were collected under very
different experimental conditions. Therefore, this study was undertaken
to determine if the queen maturation interval for Africanized honey bees
is significantly different than that for European honey bees under
identical conditions. Three aspects of queen maturation were evaluated:
1) larval, pupal and total development time from egg to adult emergence;

57
2) post-emergence development of queen attractiveness to drones; and 3)
time from adult emergence to initiation of oviposit ion.
In the studies reported here, queen development and maturation were
evaluated under controlled conditions. Total development time is
defined as the time from oviposition to adult emergence. These
experimental conditions avoid the problems of previous studies that
evaluated queen development and maturation in colonies that were
swarming (e.g., Otis 1980). Under natural swarming conditions, queens
are very often confined within their cells by worker bees and prevented
from emerging for 1-10 days after pupal eclosin (Otis 1980).
Confinement makes calculations of development times difficult, and,
because maturation proceeds during confinement, maturation time
calculated from emergence to beginning of oviposition would be under
estimated.
Methods
Queen Development Times
The Africanized egg source (A26) and the Africanized cell-producing
colonies (A37 and A43) were established from queens removed from feral
colonies found in an area of eastern Venezuela where there were no known
European honey bees. They were identified as Africanized honey bees by
their behavior and characteristic comb cell size (4.5-5.0 mm wide
between opposite sides of the hexagon, see Chapter III). The European
egg source (Y5) and the European cell-producing colonies (19, 27 28, F,
H and HI) came from European queens commercially produced in the
southeastern U.S.A. and shipped to Venezuela. European colony IBR was a
stock supplied by the U.S. Department of Agriculture Bee Breeding and
Stock Center Laboratory, Baton Rouge, Louisiana, USA.

58
Eggs of known ages were collected from the Africanized (A26) and
European (Y5) egg sources by the standard commercial queen-producing
technique of caging the queen on an empty comb within a colony (Harp
1973; Laidlaw 1979; see also Chapter II). After 6 hours, the combs with
the egg samples were moved to strong incubator-colonies for the eggs to
develop and larvae to hatch and be fed. Very young larvae, 12-18 hours
old, were transferred (grafted) into beeswax queen-cell cups primed with
royal jelly and then introduced into queen-cell-producing colonies
(=nurse bee colonies) (Laidlaw 1979). All cell-producing colonies had
large worker bee populations and were intentionally crowded into two
standard Langstroth hive bodies. Queens in the cell-producing colonies
were removed 48 hours before introducing the grafted cells. All young,
unsealed brood was also removed 2-4 hours before introducing the grafted
cells. Twenty grafted Africanized and twenty grafted European cells
were introduced into each cell-producing colony. There were twenty cell
cups to a frame, ten on the top bar and ten on the middle bar. Both
Africanized and European larvae were grafted into the same frame, five
each on the top bar and five each on the middle bar. All cell cups were
equally spaced about 8 mm apart, centered on the bars.
In one experimental trial, the effect of nurse bee genotypes on
queen development was evaluated by comparing queen development times for
both Africanized and European egg genotypes in both Africanized and
European cell-producing colonies. In another trial, development times
for Africanized queens in Africanized and European cell-producing
colonies were compared. In both trials, the Africanized cell-producing
colonies had comb cell sizes characteristic of Africanized honey bees
(4.8 mm wide; Chapter III).

59
The queen-cell-producing colonies were inspected only after the
queen cells had been sealed in order to avoid disturbance which could
affect development times. Once the cells are sealed, cell-producing
colonies only maintain the appropriate temperature for the pupae to
develop normally. On the sixth day after grafting, each sealed cell was
protected by placing a 3 mm wire mesh tube around it to avoid any
problems associated with queens being confined to their cells by worker
bees. In addition, this also prevented any emerged virgin queens from
destroying sealed cells that had not yet emerged. Beginning 24 hours
before any expected queen emergence, the cell-producing colonies were
inspected daily at 0630, 1200 and 1730 hours to record queen emergence.
In two trials, cell-producing colonies were inspected daily at 0630,
1200 and 1730 hours, beginning 24 hours prior to estimated sealing time,
in order to determine unsealed development times.
Development of Attractiveness of Virgin Africanized and European Honey
Bee Queens to Drones
Two Africanized queen mothers (A26 and A57) were removed from feral
colonies in eastern Venezuela. The two European queen mothers (We and
Yk) were shipped to Venezuela from different commercial queen breeders
in southeastern USA.
Queens from the four queen mothers (A26, A57, We and Yk) were
produced as described above. Sealed queen cells were removed from the
cell-producing colony and placed in an incubator (35 + 1C) 48 hours
prior to emergence. After emergence, the queens were marked for
individual identification and maintained in separate cages in a queen
storage colony (Laidlaw 1979).
In order to test for the degree of attractiveness to drones, each
queen was tethered in a clean, plastic screen bag. The mesh size was 1

60
x 1 mm, and each bag was approximately 5 x 10 cm. Bags were
individually suspended on monofilament line about 6.5 meters above the
ground, centered between two poles 20 meters apart. Queens could be
rapidly raised and lowered by a pulley system. Queens were put into the
mesh bags just prior to testing in order to avoid any pheromone
accumulation.
The testing location was in an open field in a drone congregation
area (Zmarlicki and Morse 1963), which was located by walking with a
helium-filled weather balloon with mature queens suspended 10-20 meters
above the ground. The drone congregation area was identified when
hundreds of drones oriented to the tethered queens. Boundaries appeared
to be quite distinct and stable through time. Both Africanized and
European drones were probably present, but the identity of each drone
responding to specific queens during the experiment was not known
because there are no reliable techniques to identify individual
Africanized and European honey bees. How drone congregation areas
become established is not understood, but these areas are probably where
most mating occurs.
Individual queens were tested for drone response on consecutive
days, beginning on the day of emergence. Only one queen at a time was
tested so that the relative attraction of each queen would not be
influenced by other queens being tested simultaneously. Testing lasted
for a maximum of 3 minutes for each queen, even if no drone response was
observed. Periodically, empty bags (blanks) were tested to insure that
drones were responding only to the queens and not orienting to the
experimental set-up and responding to the mesh bags. At no time did
drones respond to the blanks. A random sequence for testing individual

61
queens was established on each day of the experiment. Each queen was
tested more than once on each day and always in a new mesh bag, to avoid
any pheromone accumulation or contamination. Each testing session was
begun by suspending an older queen that had previously been determined
to be maximally attractive, in order to insure that a responding drone
population was available. This process was also repeated if the testing
session was interrupted by rain, extreme cloudiness, or high winds
conditions that normally reduce drone flight activity. The testing took
place between 1400 and 1600 hours.
Drone response was evaluated by assigning one of the following
ranks to the test queen:
Rank 0 = no response
Rank 1 = drones oriented to the test subject but only flew past;
no circling of the test subject
Rank 2 = drones oriented to the test subject and persisted in a
wide circling formation more than 2 m from the subject
Rank 3 = drones oriented to the test subject and formed a loose
comet-like formation down wind more than 0.5 m to the
test subject; formation was volatile, continually
fragmenting and reforming; drones did not land on the
mesh bag
Rank 4 = drones oriented to the test subject and formed a tight
comet-like formation down wind less than 0.5 m from the
test subject; formation was persistent and did not
fragment even as the test subject was lowered; drones
landed on and walked over the mesh bag.

62
These ranking categories were easily discriminated and were not affected
by the absolute numbers of drones flying. No estimates of the drone
population were made.
Time Post-Emergence to the Initiation of Oviposition
Queens were produced as described above from one Africanized egg
source (A26) and one European egg source (We). Twenty-seven Africanized
and twenty-five European mature queen cells (two days prior to
emergence) were each introduced into a four-frame queenless mating
colony. Any natural queen cells in the mating colonies were destroyed
before introducing the experimental queen cells. This insured that the
only queen in the mating colony would be the experimental queen. When
only one queen cell is present worker bees usually do not confine her
to her cell and the problem of calculating maturation time is avoided.
Because Africanized and European queens did not develop at the same
rate the day of queen emergence was determined by the mean time of
emergence for a sample of sister queens from the same graft that were
left to emerge in an incubator at 35 + 1C. On the eleventh day after
the queens emerged the colonies were inspected and the age of the brood
was evaluated to determine the age post-emergence when the queens had
begun ovipositing. Those colonies in which there were no larvae were
inspected three and five days later.
This experiment took place during the dry season. Clear weather
prevailed so that mating flights were not affected by weather
conditions. Both Africanized and European drones were in the area.

63
Res u1ts
Queen Development Times
Table 4-1 presents the experimental matrix for evaluating the
interaction of egg genotype and nurse bee genotype on queen development
times for Africanized and European queens. Table 4-2 presents the total
development times from oviposition to adult emergence for Africanized
queens and European queens in Africanized and European cell-producing
colonies. Table 4-3 presents the analyses for the paired comparisons in
each cell-producing colony. These paired comparisons avoid any
differences between cell-producing colonies because colony size (nurse
bee population), brood area temperature, and quantity and quality of
larval food are factors that affect queen development (Beetsma 1979;
Laidlaw 1979; Johansson and Johansson 1973). Africanized queens develop
in 14.5 days post-oviposition compared with 15.0 days for European
queens (P<0.001, Kolmogorov-Smirnov one-tailed test, chi-square
distribution, df = 2; Siegel 1956). There was no significant effect of
the cell-producing colony on queen development times (Kolmogorov-Smirnov
two-tailed test, chi-square distribution, df = 2, alpha = 0.05) (Tables
4-4 and 4-5).
Table 4-6 presents the development times for the Africanized queens
in Africanized and European cell-producing colonies. There was no
difference in Africanized queen development time between Africanized and
European cell-producing colonies (Kolmogorov-Smirnov one-tailed test,
chi-square distribution, df = 2, alpha = 0.05).
The median unsealed development times from oviposition to sealing
for both the Africanized and European queens was 7.5 days (Table 4-7).
However, the Africanized and European genotypes were significantly

64
different as a result of the distribution around the median (P<0.05,
Kolmogorov-Smirnov one-tailed test* chi-square distribution, df = 2).
Development of Attractiveness to Drones
The response of drones to tethered, virgin queens is summarized in
Table 4-8. There were no differences between Africanized and European
queens with respect to either the earliest age at which a positive drone
response (Rank 1) was observed or the earliest age at which a maximum
drone response (Rank 4) was observed. Both Africanized and European
virgin queens were able to attract drones (Rank 1) on the day they
emerged. Africanized virgin queens can maximally attract drones (Rank
4) by the fourth day post-emergence; European virgin queens can elicit a
Rank 4 response by the fifth day post-emergence. This difference was
not significant (Kolmogorov-Smirnov one-tailed test, chi-square
distribution, df = 2, alpha = 0.05). The data in Table 4-8 have been
combined for the two Africanized and two European queen lines. However,
the queens within a population (Africanized or European) or within a
line within a population were not uniform with respect to drone response
or the rate of maturation. There were differences between the two
Africanized lines and between individuals within the same line for the
earliest age for a Rank 4 response. This same variation between lines
and within lines existed for the European population.
Time Post-Emergence to Initiation of Ovinos it ion
Table 4-9 presents the data for time post-emergence to the
initiation of oviposition for both Africanized and European queens.
Africanized queens began oviposition at 8.5 days post-emergence whereas
European queens begin at 7.5 days (P<0.05, Kolmogorov-Smirnov two-tailed
test, chi-square distribution, df = 2).

65
Discuss ion
Queen Development
Development time from oviposition to emergence for Africanized
queens in this study was 14.5 days, which is the same as the development
period reported for both Africanized bees in French Guiana (Winston
1979c) and their parental population, A. m. adanson i i (=scutellata) in
South Africa (Anderson, Buys and Johannsmeier no date; Fletcher 1978;
Fletcher and Tribe 1977c). The European queens in the present study
developed in 15.0 days, which is about one day shorter than expected
from previous reports (Jay 1963; Laidlaw 1979). Therefore, the
difference in development times for Africanized and European queens was
not as great as expected and underscores the importance of making
comparisons under the same experimental conditions.
Queens have approximately a 25% shorter development period than
worker bees. Differences in total development times between queens and
worker bees are primarily due to a much shorter sealed development
stage, i.e., 7.5 days compared with 12 days for European bees and 7.0
days compared with 12 days for Africanized bees. The sealed development
stage in worker bees is approximately 60% of the total development time,
whereas in queens it is approximately 50%.
European queens took 3.4-5.6% longer than Africanized queens to
develop. This difference is similar to the 5.3% difference in
development times between European and Africanized worker bees from the
same two egg sources (A26 and Y5) (see Chapter II).
There was no effect of nurse bee genotype on queen development
times. However, queens were produced more successfully in European
colonies. Africanized nurse bees were easily disturbed when the grafted

66
cells were introduced into the colonies, resulting in poor survival or
acceptance of the grafted larvae (5-50% for Africanized colonies,
compared with 35-95% for European colonies). In addition, Africanized
colonies were difficult to manage because of excessive stinging that
occurred when manipulating the strong colonies that were necessary for
proper queen production.
Page and Erickson (1984) found evidence that nurse bee colonies
preferentially raised queens from more closely related larvae. However,
in the present study, no evidence for kin recognition was observed.
Africanized and European nurse bee colonies reared Africanized and
European queens with equal frequency (Table 4-2).
Rate of Maturation
Attractiveness of queens to drones is a function of the amount of
pheromone (9-oxodec-trans-2-enoic acid) produced in the mandibular
glands of the queens (Butler 1971; Boch, Shearer and Young 1975). In
England, using European genotypes, Butler (1971) tethered virgin queens
of various ages 6 meters above the ground in areas where drones were
flying. He determined that queens younger than 5 to 6 days old seldom
elicited a positive drone response. Maximum positive responses from
drones were observed in queens 8 or more days old. Butler's results
differ from those presented in this study and may be attributed to
either differences in experimental conditions or genetic differences
between the queen lines studied rather than to differences due to any
tropical or temperate conditions. The response of drones to queens
reported in this chapter was evaluated in a drone congregation area
which may account for the differences between the studies.

67
In addition, Africanized drones (not present in Butler's study) may
have a lower response threshold to queen pheromone and therefore would
respond to queens with less pheromone present than would European
drones. This hypothesis is suggested by the observation that there are
differences in the sensory receptors on the antennae of Africanized
drones compared with European drones (Dietz 1978). Further comparison
between Africanized and European drones is needed to determine any
differences in the threshold of response and whether or not this would
give the Africanized drones a mating advantage.
Another factor that needs to be considered when using this
behavioral bioassay (drone response) to compare rate of maturation of
queens 1s that the pheromone is not continually produced but rather is
pulsed in its production (R. Boch, pers. comm.). This factor may help
to explain some of the variation of responses produced by queens within
the same line. For example, in a few trials, a queen elicited a
decreased response compared with the previous response she had elicited.
In the present study, the time post-emergence to the initiation of
oviposition for Africanized queens was 8.5 days and for European queens
was 7.5 days. There are no other data available that allow for valid
comparisons. For example, Otis (1980) reports that the mean interval
from emergence to initiation of oviposition was 7.8 days for Africanized
queens in French Guiana. However, these data were collected by
observing queen maturation in colonies that had swarmed and, therefore,
the time from emergence to oviposition would be shortened because of a
variable period of queen confinement [110 days (Otis 1980)] within the
cells. In another set of data, Otis (1980) reports the mean maturation
interval from eclosin to oviposition was 9.7 days. However, he does

68
not indicate how he determined when eclosin occurred or if he was
using the terms eclosin and emergence interchangeably. The normal time
from eclosin to emergence for queens is approximately 12 hours (Jay
1963).
Because of their high reproductive rate and resultant short swarm-
to-swarm interval Africanized honey bees were expected to have a rapid
queen maturation interval compared with European honey bees. Fletcher
and Tribe suggest "that in the adansonii f=scute11ata] race natural
selection has worked strongly in favour of minimizing the period between
the loss of a queen [from swarming] and the re-establishment of
oviposition by a new queen" (1977b, p. 167). The surprising result from
this study was that both Africanized and European queens matured at
approximately the same rate, determined both by their attractiveness to
drones and the time from adult emergence to initiation of oviposition.
As Fletcher and Tribe (1977b) suggested, one would expect natural
selection to be operating to minimize the maturation interval for
queens, in order to maximize brood production between swarming periods.
However, Africanized queens may be under a second and possibly more
important selection pressure which may affect their maturation interval.
Africanized swarms may travel great distances (Fletcher 1978; Michener
1975). Otis (1980) confirmed that at least some queens issuing with
afterswarms had already mated. If new queens issuing with these swarms
have mated prior to swarming or mate while enroute, then delayed
maturation, particularly with respect to development of ovarian
follicles, would be advantageous. Follicular development would increase
the queen's weight and make it more difficult for her to fly. Prior to
issuing with the prime swarm, older queens usually stop egg laying

69
several days before the swarm departs, allowing time for their ovaries
to recess. Therefore, maturation for Africanized queens may be delayed
in order for the swarms with new queens to be able to migrate long
distances. Rather than selection operating to shorten the maturation
interval, selection may be operating to delay maturation to enable long
swarm migration distances.
The variation in queen maturation rates (see Tables 4-8 and 4-9)
observed both within a population and within a queen line suggests that
the physiological parameters involved in the process of maturation may
be genetically determined. The rate of maturation is an important
economic characteristic for commercial queen producers to consider in
their selection programs. Reducing the time from emergence to
initiation of oviposition can significantly increase the number of
queens produced in each mating colony during the queen-producing season.

TABLE 4-1. Experimental matrix for the comparison of total
development times (oviposition to adult emergence)
for both Africanized and European honey bee queens.
EGG GENOTYPES
NURSE BEE GENOTYPE3 AFRICANIZED (A26) EUROPEAN (Y5)
AFRICANIZED
A43 A
A37 C
COMBINED E
EUROPEAN
19 G
27 I
28 K
F M
H 0
IBR Q
COMBINED S
B
D
F
H
J
L
N
P
R
T
COMBINED AFRICANIZED
AND EUROPEAN U V
aQueen-cell-producing colony.

TABLE 4-2. Total development times (in days from
oviposit ion to adult emergence) for
Africanized and European honey bee queens:
median (sample size).
EGG GENOTYPES
NURSE BEE GENOTYPE3 AFRICANIZED (A26) EUROPEAN (Y5)
AFRICANIZED
A43
14.0
(1)
15.0
(4)
A37
14.5
(7)
15.0
(4)
COMBINED
14.5
(8)
15.0
(8)
EUROPEAN
19
14.5
(15)
15.0
(19)
27
14.5
(17)
15.0
(13)
28
14.0
(13)
14.5
(12)
F
14.0
(10)
14.5
(9)
H
14.0
(7)
14.5
(8)
IBR
14.0
(8)
14.8
(8)
COMBINED
14.2
(70)
15.0
(69)
COMBINED AFRICANIZED
AND EUROPEAN
14.5
(78)
15.0
(77)
aQueen-cell-producing colony.

72
TABLE 4-3. Analyses for the comparison of queen development
times for both Africanized and European honey bee
genotypes. Letters A V represent different
treatments; see Table 4-1 for explanation.
Kolmogorov-Smirnov one-tailed test, chi-square
distribution, df = 2, alpha = 0.05 (Siegel 1956).
A
X
B
NSa
C
X
D
NSa
E
X
F
G
X
H
***
I
X
J
**
K
X
L
**
M
X
N
**
0
X
P
*
Q
X
R
*
S
X
T
***
U
X
V
***
aSmall sample size, chi-square distribution is conservative.
b = P<0.05
** = P<0.01
*** = P<0.001.

73
TABLE 4-4. Analyses of queen development times for the
Africanized egg genotype in the different cell-
producing colonies. Letters represent different
cell-producing colonies; see Table 4-1 for
explanation. Kolmogorov-Smirnov two-tailed test
chi-square distribution df = 2 alpha = 0.05
(Siegel 1956).
A x C
NS
A x G
NS
A x I
NS
A x K
NS
A x M
NS
A x 0
NS
A x Q
NS
C x G
NS
C x I
NS
C x K
NS
C x M
NS
C x 0
NS
C x Q
NS
G x I
NS
G x K
NS
G x M
NS
G x 0
NS
G x Q
NS
I x K
NS
I x M
NS
I x 0
NS
I x Q
NS
K x M
NS
K x 0
NS
K x Q
NS
M x 0
NS
M x Q
NS
0 x Q
NS

74
TABLE 4-5. Analyses of queen development times for the
European egg genotype in the different cell-
producing colonies. Letters represent different
cell-producing colonies; see Table 4-1 for
explanation. Kolmogorov-Smirnov two-tailed test
chi-square distribution df = 2 alpha = 0.05
(Siegel 1956).
B x D
NS
B x H
NS
B x J
NS
B x L
NS
B x N
NS
B x P
NS
B x R
NS
D x H
NS
D x J
NS
D x L
NS
D x N
NS
D x P
NS
D x R
NS
H x J
NS
H x L
NS
H x N
NS
HxP
NS
H x R
NS
J x L
NS
J x N
NS
J x P
NS
J x R
NS
L x N
NS
L x P
NS
L x R
NS
N x P
NS
N x R
NS
P x R
NS

TABLE 4-6. Total development time (in days from oviposition
to adult emergence) of Africanized queens in
Africanized and European cell-producing colonies
median, (sample size).
NURSE BEE GENOTYPE3 AFRICANIZED EGG GENOTYPE (A26)
AFRICANIZED5
A43
14.4
(10)
A
A37
14.6
(6)
B
A43 & A37
14.4
(16)
C
EUROPEAN
HI
14.4
(16)
D
IBR
14.2
(14)
E
HI & IBR
14.4
(30)
F
ANALYSES0
A x D NS
A x E NS
B x D NS
B x E NS
C x F NS
3Queen-cell-producing colonies.
^Africanized comb cell size.
Kolmogorov-Smirnov one-tailed test, chi-square
distribution, df = 2, alpha = 0.05.

76
TABLE 4-7. Unsealed (egg and larval periods combined) development times
(in days) for Africanized and European queens: median,
(sample size).
EGG GENOTYPES
NURSE BEE GENOTYPE3
AFRICANIZED (A26)
EUROPEAN (Y5)
EFFECT OF EGG
GENOTYPE5
19
7.5 (17)
7.5 (19)
*
28
7.2 (14)
7.5 (12)
*
19 & 28
7.5 (31)
7.5 (31)
*
EFFECT OF NURSE BEE
GENOTYPE0
NS
NS
^Cell-producing colonies, European nurse bees, European comb cell size.
bKolmogorov-Smirnov one-tailed test, chi-square distribution, df = 2,
* = P<0.05.
cKolmogorov-Smirnov two-tailed test, chi-square distribution, df = 2,
alpha = 0.05.

TABLE 4-8. Drone response to tethered virgin queens: median
day of response post-emergence, range, (sample
size).
DAY OF RESPONSE LEVEL
RANK 1
RANK 2
RANK 3
RANK 4
AFRICANIZED QUEEN
0a
3.5
3.5
4.0
GENOTYPES

1-5
1-5
1-5
(2)
(6)
(6)
(5)
EUROPEAN QUEEN
0a
1.5
4.0
4.5
GENOTYPES

1-5
2-5
4-5
(1)
(4)
(3)
(2)
ANALYSES6
NS
NS
NS
aDay 0 = day of adult emergence.
bKolmogorov-Smirnov one-tailed test, chi-square distribution,
df = 2, alpha = 0.05.

TABLE 4-9. Time post-emergence to initiation of oviposition
median, range, (sample size).
AFRICANIZED GENOTYPE (A26)
DAYS POST-EMERGENCE
8.5
7.5-12
(10)
EUROPEAN GENOTYPE (We)
7.5
6-10
(16)
ANALYSIS3
P<0.05
Kolmogorov-Smirnov two-tailed test, chi-square distribution,
df = 2.

CHAPTER V
QUEEN PUPAL WEIGHTS
Introduction
Africanized honey bees in South America are hybridized descendents
of African honey bees (Apis me!1 ifera scute!lata) and European honey
bees (primarily A. ¡n. 1 igustica and A. m.. me! 1 if era! (Goncalves 1982;
Woyke 1969). The annual net reproductive rate of Africanized honey bees
in South America is four to five times greater than that of European
honey bees in temperate regions: 16 colonies per colony per year
compared with 3-3.6 (Otis 1980 1982a; Winston 1980a; Winston, Taylor
and Otis 1983). Differences in reproductive rates between these two
honey bee populations may be a result of: 1) colony demography;
2) temperate vs. tropical climate and floral resources; 3) resource
utilization behaviors; or 4) a combination of factors. Because
Africanized and European honey bees have not been compared under
identical experimental conditions, it is not possible to determine to
what extent reproductive differences are a result of genetic or
environmental parameters.
One demographic parameter associated with rapid colony growth and a
high rate of colony reproduction would be a high oviposition rate (Brian
1965; Moeller 1961; Wilson 1971). In the evolution of social insects,
queen oviposition rates have increased primarily due to one of the
following: increased number of ovarioles, increased length of the
79

80
ovarioles, more rapid egg maturation, and reduction in egg size (Hagan
1954 and Iwata and Sakagami 1966 cited in Wilson 1971; Wilson 1971).
Honey bee queens have a very large number of ovarioles (>300) and, for
European queens, the number of ovarioles has been shown to be an
inherited character (Eckert 1934) which is positively correlated with
queen pupal weight (Hoopingarner and Farrar 1959). Queen weight was
also found to be correlated with brood production (Boch and Jamieson
1960). If it is assumed that both Africanized and European honey bees
have the same relationship between queen weight and brood production,
then weights of queens from the two populations can be compared to
determine potential differences in fecundity.
In honey bees, differentiation between worker and queen castes is
not genetically determined, but rather is regulated by the quantity and
quality of food fed to developing larvae during the first 3 days
(Beetsma 1979). Therefore, a number of factors other than genotype
affect queen size, e.g., age of larvae used to produce queens,
population of the cell-producing colony, quantity and quality of food
fed to developing larvae, and temperature (Beetsma 1979; Johansson and
Johansson 1973; Laidlaw 1979; Weiss 1974; Woyke 1971). Because of
differences in queen rearing methods and experimental conditions,
previous comparisons of size between Africanized and European queens may
be inappropriate. This study was undertaken to compare queen pupal
weights for Africanized and European honey bees under identical
experimental conditions in Venezuela.
Methods
Four Africanized honey bee lines (A26, A57, A61 and A62) were
established from queens removed from feral colonies in an area in

81
eastern Venezuela that had no known European honey bees. They were
identified as Africanized honey bees by their comb cell size, which was
significantly smaller than European comb cell size (Chapter III). Two
European lines (YK and WE) were established from queens shipped to
Venezuela by commericial queen producers in the southeastern U.S.A.
Three additional European lines (YD, N and GK) were established from
queens shipped to Venezuela from the U.S. Department of Agriculture Bee
Breeding and Stock Center Laboratory, Baton Rouge, Louisiana, U.S.A.
Queens were produced from these nine lines by standard queen
rearing methods (Laid!aw 1979). Egg samples from the nine queen mothers
were collected by confining the queens to an empty comb within their own
colonies using an 8 x 8 cm push-in cage made from 3 mm mesh hardware
cloth. -Queen excluder material was soldered to the tops of the push-in
cages, allowing worker bees to move in and out in order to feed and tend
the queen (Harbo, Bol ten, Rinderer and Collins 1981). Both Africanized
and European eggs were collected in European size comb. After
approximately 4-6 hours, the queens were released, and combs containing
the eggs were put into a strong colony in order for the eggs to be
incubated and for the larvae to be fed. Africanized and European eggs
were both put into the same incubator-colony in order to control for any
differences in early larval feeding and temperature.
Young larvae approximately 12-15 hours old were transferred
(grafted) into artificial, beeswax, queen-cell cups and then introduced
into the cell-producing colonies. Twenty larvae from one of the
Africanized lines and twenty larvae from one of the European lines were
grafted into each cell-producing colony. To control for extrinsic
factors affecting queen size, analyses of Africanized and European

82
queens were limited to paired comparisons (one Africanized and one
European line) that were each simultaneously introduced into the same
incubator-colony and then grafted into the same queen-cell-producing
colony. Possible effects from different cell-producing colonies on
queen pupal weight were evaluated by grafting the same queen lines into
different cell-producing colonies.
Only European cell-producing colonies were used because of the
difficulty in producing queens in Africanized colonies. Africanized
cell-producing colonies remained disturbed for a long period of time
after the grafted larvae were introduced which resulted in poor
acceptance (survival) of the larvae (see Chapter IV).
Queen pupal weights are used for comparison because adult weights
vary with respect to engorgment of food, dehydration, feces
accumulation, and differential ovariole development. Although queen
pupal weights vary with age of the pupae, there is a period from the
10th through the 13th day post-oviposition when queen pupal weight is
constant (Table C-l). Queen pupal weight comparisons can therefore be
made during this period (Hoopingarner and Farrar 1959). Although there
is a 0.5 day difference in development time between Africanized and
European queens (Chapter IV), the 3-4 day pupal period during which
there is no significant weight change is of sufficient duration to allow
Africanized and European queens to be accurately and consistently
compared. Africanized and European queen pupae were weighed on the 11th
day post-oviposition. Queen cells from each of the lines were randomly
selected to avoid any position effect from location on the grafting
frame. Weights were measured to the nearest 1.0 mg using either a
Mettler Type H4 or H6 balance.

83
Queen cell lengths were measured at the time the queen pupal
weights were determined. A calipers was used to determine the external
length from the base to the apex of the queen cell.
Results *
Queen pupal weights for four Africanized and five European lines
are presented in Table 5-1. European queen pupal weights were
significantly larger than Africanized queen pupal weights for two
different pairwise comparisons (YK vs. A26 and YK vs. A57; P<0.05 to
P<0.001; Mann-Whitney U test one-tailed). Africanized queen pupal
weights were significantly larger in one pairwise comparison (A62 vs. N;
P<0.02; Mann-Whitney U test two-tailed). For three pairwise
comparisons there was no statistical difference (A26 vs. WE, A57 vs.
YD, and A61 vs. GK; Mann-Whitney U test, one-tailed, alpha = 0.05).
Because different cell-producing colonies had no significant effect on
queen pupal weights (see below), the means for the nine queen lines can
be ranked and analyzed (Table 5-2). There was no significant difference
between the Africanized and European honey bee populations for queen
pupal size (Mann-Whitney U test, one-tailed, alpha = 0.05).
Queen cell lengths for Africanized and European lines are presented
in Table 5-3. In six out of eight pairwise comparisons, there was no
significant difference in queen cell lengths between Africanized and
European queens (Mann-Whitney U test, one-tailed, alpha = 0.05). For
the pair in cell-producing colony 2, the European line was significantly
larger than the Africanized line (P<0.05). For the pair in cell-
producing colony 4, the Africanized line was significantly larger than
the European line (P<0.02; Mann-Whitney U test, two-tailed).

84
Spearman's rank correlation coefficient was determined for queen
pupal weights and queen cell lengths (Table 5-4). In general there was
no significant correlation between queen pupal weight and queen cell
length (alpha = 0.05). However, one Africanized line (A26) in cell-
producing colony 2 had a significant correlation (P<0.05) and one
European line (YK) in cell-producing colony 4 had a significant
correlation (P<0.01).
The effect of cell-producing colonies on queen pupal weights and
queen cell lengths is presented in Table 5-5. There was no significant
difference for Africanized queen line A26 in four different cell-
producing colonies (one-way analysis of variance, alpha = 0.05); nor was
there a significant difference for Africanized queen line A57 in two
different cell-producing colonies (Mann-Whitney U test, two-tailed,
alpha = 0.05). There was no significant difference in pupal weights for
the European queen line YK in four different cell-producing colonies
(one-way analysis of variance, alpha = 0.05), but there was a
significant effect of cel 1-producing colonies on queen cell length
(P<0.001). When cell-producing colony 2 was removed from the analysis,
there was no significant difference in queen cell length.
Discussion
If we assume for both Africanized and European honey bees that
queen weight is correlated with egg production or fecundity (Boch and
Jamieson 1960), we would then expect that egg laying rates would follow
the same ranking as presented in Table 5-2 for queen pupal weights.
Based on these pupal weights, we would predict that there would be no
difference in egg laying rates for the Africanized and European honey
bee populations. In fact, when egg laying rates for Africanized and

85
European honey bee queens were compared there was no significant
difference between queens from the two populations (Chapter VI). There
were, however, significant differences in pupal weights between
individual queen lines both between and within each population (Table 5-
1). There were also significant differences in egg laying rates between
individual queen lines both between and within the two populations
(Chapter VI).
In this study, Africanized queens were reared in European colonies
because of low acceptance (survival) of grafted cells in Africanized
colonies (Chapter IV). Because queen-worker caste differentiation in
honey bees is regulated by larval feeding (Beetsma 1979), rearing
Africanized queens in European colonies may have obscured differences in
pupal weights between Africanized and European queens. Possibly,
European worker bees may rear larger Africanized queens than would
Africanized worker bees because European worker bees are themselves
larger (Chapter III), and may feed the developing queen larvae
differently. Although virgin European queens have been reported to
weigh more than virgin Africanized queens208 vs. 199 mg (Goncalves,
Kerr and Nocoes 1972 cited in Michener 1975)there was no indication of
conditions under which the queens were reared.
Further analysis of queen weights between Africanized and European
honey bees is needed, preferably in a 2 x 2 experimental design:
Africanized and European queens reared in both Africanized and European
cell-producing colonies. In addition, the relationship between queen
pupal weights and brood production needs to be evaluated for both
Africanized and European honey bee lines to determine if the same
relationship exists for both populations.

86
The European queen lines evaluated were a diverse representation of
the European population from North America, whereas the Africanized
queen lines may only reflect a small portion of the Africanized
population. The location for the sources of the Africanized lines was
limited to feral colonies found in one area of eastern Venezuela. A
greater diversity of Africanized lines needs to be evaluated in order to
be able to generalize about queen pupal weights and oviposition rates
for the population as a whole.

TABLE 5-1. Comparison of Africanized and European queen pupal weights
(mg): mean + SD, (sample size), (genotype).
87
AFRICANIZED EUROPEAN
CELL BUILDER3 GENOTYPES GENOTYPES ANALYSES5
1
257.1
+ 7.6
(9)
(A26)
2
255.8
+ 9.2
(9)
(A26)
3
262.3
+ 6.4
(3)
(A26)
4
260.0
+ 9.0
(11)
(A57)
5
243.0
+ 13.4
(3)
(A26)
6
248.6
+ 14.3
(4)
(A57)
7
291.8
+ 7.9
(5)
(A62)
8
237.2
+ 21.7
(5)
(A61)
286.4
(9)
+ 12.0
(YK)
***
284.4
(9)
+ 20.6
(YK)
**
266.7
(9)
+ 9.7
(WE)
NS
272.5
(13)
+ 16.5
(YK)
*
282.6
(3)
+ 12.0
(YK)
*
264.0
(5)
+ 15.4
(YD)
NS
257.3
(4)
+ 18.2
(N)
c
232.9
(2)
+ 0.4
(GK)
NS
aCe11-producing colonies; European nurse bees and European comb
cell size.
bMann-Whitney U test, one-tailed, alpha = 0.05;
* = P<0.05, ** = P<0.01, *** = P<0.001.
difference in wrong direction for one-tailed test; two-tailed
test results in a P<0.02.

TABLE 5-2. Queen pupal weights for the nine lines
analyzed.
POPULATION QUEEN LINE MEAN PUPAL WEIGHT(MG)
European
GK
233
Africanized
A61
237
Africanized
A26
256
European
N
257
Africanized
A57
257
European
YD
264
European
WE
267
European
YK
280
Africanized
A62
292
ANALYSIS3
NS
^ann-Whitney U test one-tailed, alpha = 0.05

TABLE 5-3. Comparison of Africanized and European
lengths (mm): mean + SD, (sample size)
(genotypes).
queen cell
9
AFRICANIZED
CELL BUILDER3 GENOTYPES
EUROPEAN
GENOTYPES
ANALYSES13
1 2.58 + 0.1
(9) (A26)
2.50 + 0.1
(9) (YK)
NS
2 2.62 + 0.2
(9) (A26)
2.70 + 0.1
(9) (YK)
*
3 2.54 + 0.1
(3) (A26)
2.48 + 0.1
(9) (WE)
NS
4 2.58 + 0.1
(7) (A57)
2.44 + 0.1
(11) (YK)
c
5 2.67 + 0.1
(3) (A26)
2.57 + 0.1
(3) (YK)
NS
6 2.50 + 0.02
(4) (A57)
2.57 + 0.1
(5) (YD)
NS
7 2.78 + 0.04
(5) (A62)
2.81 + 0.1
(3) (N)
NS
8 2.79 + 0.1
(5) (A61)
2.76 + 0.1
(2) (GK)
NS
aCen-producing colonies; European nurse bees and European comb
cell size.
bMann-Whitney U test, one-tailed, alpha = 0.05; = P<0.05.
difference in wrong direction for one-tailed test; two-tailed
test results in a P<0.02.

TABLE 5-4. Correlation of queen cell length (mm) and queen pupal
weight (mg): mean + SD, (sample size). Measurement
made on day 11.25 post-ov iposit ion.
QUEEN CELL QUEEN PUPAL
QUEEN GENOTYPE LENGTH WEIGHT CORRELATIONS3
AFRICANIZED
A26
(CB1)b
2.58 + 0.1
257.1 + 7.6
NS
(9)
(9)
A26
(CB2)
2.62 + 0.2
255.8 + 9.2
*
(9)
(9)
A26
(CB3)
2.54 + 0.1
262.3 + 6.4

(3)
(3)
A26
(CB5)
2.67 + 0.1
243.0 + 13.4

(3)
(3)
A57
(CB4)
2.58 + 0.1
259.1 + 10.8
NS
(7)
(7)
A57
(CB6)
2.50 + 0.02
248.6 + 14.3
NS
(4)
(4)
A62
(CB7)
2.78 + 0.04
291.8 + 7.8
NS
(5)
(5)
A61
(CB8)
2.79 +0.1
237.2 + 21.7
NS
(5)
(5)
EUROPEAN
YK
(CB1)
2.50 + 0.1
286.4 + 12.0
NS
(9)
(9)
YK
(CB2)
2.70 + 0.1
284.4 + 20.6
NS
(9)
(9)
YK
(CB4)
2.44 + 0.1
272.2 + 17.8
**
(11)
(11)
YK
(CB5)
2.57 + 0.1
282.6 + 12.0

(3)
(3)
WE
(CB3)
2.48 + 0.1
268.4 + 8.8
NS
(8)
(8)
YD
(CB6)
2.57 + 0.1
264.0 + 15.4
NS
(5)
(5)
N
(CB7)
2.81 + 0.1
264.3 + 14.0

(3)
(3)
GK
(CB8)
2.76 + 0.1
232.9 + 0.4

(2)
(2)
aSpearman's rank correlation coefficients, alpha = 0.05;
* = P<0.05; ** = P<0.01.
bCB = cell-producing colony number; refer to Table 5-1 for
explanation.

91
TABLE 5-5 Effect of cell-producing colony on queen cell length and
queen
pupal weight: mean
+ SD, (sample size).
AFRICANIZED QUEEN
GENOTYPE
(A26)
CBla
CB2
CB3
CB5
ANALYSES13
QUEEN CELL
LENGTH
2.58
+ 0.1
(9)
2.62
+ 0.2
(9)
2.54
+ 0.1
(3)
2.67
+ 0.1
(3)
NS
QUEEN PUPAL
WEIGHT
257.1
+ 7.6
(9)
255.8
+ 9.2
(9)
262.3
+ 6.4
(3)
243.0
+ 13.4
(3)
NS
AFRICANIZED QUEEN
GENOTYPE
(A57)
CB4
CB6
ANALYSES
c
QUEEN CELL
LENGTH
2.58
+ 0.1
(7)
2.50
+ 0.02
(4)
NS
QUEEN PUPAL
WEIGHT
260.0
+ 9.0
(11)
248.6
+ 14.3
(4)
NS
EUROPEAN QUEEN GENOTYPE (YK)
CB1
CB2
CB4
CB5
ANALYSES5
QUEEN CELL
LENGTH
2.50
+ 0.1
(9)
2.70
+ 0.1
(9)
2.44
+ 0.1
(11)
2.57
+ 0.1
(3)
***d
QUEEN PUPAL
WEIGHT
286.4
+ 12.0
(9)
284.4
+ 20.6
(9)
272.5
+ 16.5
(13)
282.6
+ 12.0
(3)
NS
aCB = cell-producing colony number; refer to Table 5-1 for explanation.
b0ne-way analysis of variance, alpha = 0.05; *** = P<0.001.
^Mann-Whitney U test, two-tailed, alpha = 0.05.
dWith CB2 removed, ANOVA is NS.

CHAPTER VI
egg'laying Arc BROOD production rates during the first brood cycle
Introduction
Africanized honey bees in South America are descendents from the
hybridization of African honey bees (Ad is me!1 ifera scutellata) and
European honey bees (primarily A. [n. 1 iaustica and A. m. mel 1 ifera)
(Goncalves 1982; Woyke 1969). In tropical and sub-tropical regions of
South America, Africanized bees have been more successful than European
bees as determined by their rapid rates of dispersal and high population
densities (Michener 1975; Taylor 1977, 1985). It is not surprising that
Africanized bees are more successful in these regions because they are
descendents of honey bees that evolved under similar tropical conditions
in Africa.
Rates of dispersal and population densities achieved by Africanized
honey bees require a high colony reproductive rate. Africanized honey
bees in South America have a reproductive rate that is four to five
times greater than the reproductive rate of European honey bees in North
America (Otis 1980, 1982a; Winston 1980a; Winston, Taylor and Otis
1983). However, based on this comparison, one cannot identify the
factors that account for differences in reproductive rates nor identify
the factors leading to the success of Africanized bees in South America.
Because the comparison of reproductive rates was not based on data
collected under similar environmental or experimental conditions, it is
92

93
not possible to determine to what extent reproductive differences
between the two populations are a result of differences in intrinsic
demographic parameters and/or environmental differences due to temperate
vs. tropical resources and climatic conditions. In addition,
experimental conditions were very different. For example, an important
variable affecting reproductive rates in honey bees is brood-nest
crowding (Baird and Seeley 1983; Simpson 1966, 1973; Simpson and Riedel
1963). Experimental Africanized colonies in South America were
maintained in 22-liter hives (Otis 1980; Winston 1979b), whereas,
experimental European colonies in North America, with which they were
compared, were maintained in 42-liter hives (Winston 1980a).
Reproductive rates in honey bees are a result of an interaction of
at least three factors: colony demography, resource availability, and
resource utilization efficiency. As part of a larger investigation
comparing intrinsic demographic factors between Africanized and European
honey bees to determine which aspects of demography, if any, are
responsible for the success of Africanized honey bees in South America,
this study evaluated one aspect of demographyqueen fecundity. Queen
egg laying rate is one of the primary demographic parameters that
affects colony growth rates (Brian 1965; Moeller 1961; Wilson 1971).
Although differences in egg laying rates between Africanized queens and
European queens have been reported (Fletcher 1978; Michener 1972, 1975;
Ribbands 1953), they cannot be compared because the data were collected
under different resource and experimental conditions. Therefore, the
present study was undertaken to compare egg laying and brood production
rates for both Africanized and European queens under identical, tropical
conditions in Venezuela. The experimental design allowed for a

94
comparison between the two honey bee populations during the first brood
cycle. Differences in initial colony growth rates between Africanized
and European honey bees may be an important factor in determining
differences in reproductive rates.
Egg laying and brood production rates for Africanized and European
honey bees were evaluated at both the queen and worker bee levels. The
interactions of both Africanized and European queens with both
Africanized and European worker bees were evaluated because of potential
behavioral and/or physiological differences between Africanized and
European nurse bees with respect to affecting brood production rates
and/or the queen's oviposition behavior. In order to compare egg laying
and brood production rates between Africanized and European honey bees,
four variables needed to be controlled.
First is colony size, because egg laying rates are positively
correlated with the number of worker bees in a colony (Moeller 1958).
In order to evaluate initial colony growth, a colony size was selected
that contained the number of worker bees within the range reported for
both Africanized and European swarms (Fell et. al. 1977; Otis 1980;
Rinderer, Collins, Bolten and Harbo 1981; Rinderer, Tucker and Collins
1982; Winston 1980a).
Second is hive cavity volume, which must be controlled in order to
avoid effects of differential brood-nest crowding on oviposition rates
(Brian 1965). A hive cavity volume was selected that represents natural
nest cavity volumes chosen by these two honey bee populations (Rinderer,
Collins, Bolten and Harbo 1981; Rinderer, Tucker and Collins 1982;
Seeley 1977; Seeley and Morse 1976; Winston, Taylor and Otis 1983).

95
A third variable, comb cell size, had to be controlled. If bees
were allowed to build their own comb, comb built by Africanized workers
would be smaller than comb built by European bees (Chapter III). The
larger, European comb was selected for these experiments for two
reasons: 1) European queens do not lay eggs in a uniform pattern in
Africanized comb; and 2) there is a higher brood mortality in colonies
with European nurse bees on Africanized combpossibly because the
larger European nurse bees have difficulty feeding the developing larvae
in the smaller, Africanized cells (Chapters II and III). On the other
hand, Africanized queens and worker bees appear to behave normally when
managed on European comb. Only Africanized bees that had been reared in
managed colonies with European combs were used in the experimental
colonies to avoid any delay in adjusting to larger comb cell size.
Finally, the fourth variable controlled was resource availability.
Africanized and European queens were compared simultaneously so that
floral resource conditions were identical. Surplus honey was also
provided for each experimental colony to reduce the effects of
differential foraging success between Africanized and European honey
bees in tropical resource conditions (Rinderer, Bolten, Collins and
Harbo 1984; Rinderer, Collins and Tucker 1985).
In addition to colony-level variables, conditions under which
experimental queens are produced may affect queen fecundity, e.g., age
of larvae used to produce queens, quantity and quality of food fed to
developing queen larvae, and temperature during development (Beetsma
1979; Laidlaw 1979; Weiss 1974; Woyke 1971). The age of a queen may
also affect her fecundity (Ribbands 1953). In this study, all

96
experimental queens were produced under identical conditions and were
the same age.
Estimates of daily egg laying rates derived from total brood
production may not be accurate (Merrill 1924) because mortality of
unsealed brood (eggs and larvae) may be quite high, up to 50% (Garofalo
1977; Merrill 1924; V/oyke 1977), particularly during the initial period
of colony growth (Winston, Dropkin and Taylor 1981). Therefore, both
daily egg laying rates and brood production during the first brood cycle
were analyzed.
Methods
D.fU.l.y, Egg_.Lay1ng_R.ates
Three different Africanized queen mothers (A57, A61 and A62) were
established from feral colonies that were found in an area of eastern
Venezuela with no known European honey bees. They were confirmed as
Africanized honey bees by their behavior and comb cell size (Chapter
III). Three different European queen mothers were shipped to Venezuela
from the U.S.A.; two (GK and YD) were from the U.S. Department of
Agriculture Bee Breeding and Stock Center Laboratory in Baton Rouge,
Louisiana, and the third (YK) was from a commercial queen producer in
southeastern U.S.A.
Experimental queens were produced from the queen mothers using
standard queen rearing techniques (Laidlaw 1979). Eggs from the queen
mothers were collected by restricting queens to a portion of comb for 4
to 6 hours under 8 x 8 cm push-in cages that had queen excluder material
soldered to the tops (Harbo, Bolten, Rinderer and Collins 1981). Combs
with the egg samples were then placed into a populous colony where the
eggs were incubated until the larvae hatched and were fed. Young

97
larvae, approximately 15 hours old, were transferred (grafted) into
beeswax queen-cell cups that had been primed with diluted royal jelly
and then introduced into cell-producing colonies for development. Three
days prior to adult emergence, sealed queen cells were put into an
incubator (35 +1 C.). Emerged, virgin queens were marked for
individual identification, using color-coded, plastic, numbered discs
glued to the queen's thorax (Smith 1972). Virgin queens were maintained
separately in small, two-frame colonies during the period of maturation
and after artificial insemination in order to maximize the number of
spermatozoa that migrate to the spermatheca (Woyke 1979).
Queens were artificially inseminated one week after emergence using
the apparatus designed by Harbo (1979) and Mackensen (Mackensen and
Roberts 1948; Mackensen and Tucker 1970). Queens were inseminated with
2.5 ul of wild-type semen on each of two occasions, 3 days apart, to
increase the percentage of spermatozoa entering the spermatheca (Bolten
and Harbo 1982; Mackensen 1964).
Four colony treatments were established:
1. Africanized queen with Africanized worker bees,
2. Africanized queen with European worker bees,
3. European queen with Africanized worker bees and
4. European queen with European worker bees.
Each experimental colony was in a five-frame hive (23 liters) that
contained three empty combs of European cell size. A different
geometric design was painted over the entrance of each hive in order to
aid in orientation and reduce drifting of foragers between colonies (von
Frisch 1967).

98
Two days prior to the beginning of the experiment young bees were
removed from brood frames of large colonies and put into screened cages
that measured 48 x 37 x 76 cm. One cage contained Africanized bees and
the other contained European bees. Africanized bees were removed only
from colonies that were being managed on combs of European cell size.
The cages were supplied with feeders containing 50% (volume:volume)
sugar syrup. Bees used to stock the experimental colonies were taken
from the appropriate cage, thereby insuring that all experimental
colonies were uniform in composition for a particular worker bee type,
Africanized or European. Each experimental colony was started with
approximately 775 grams of bees.
Test queens were introduced into each experimental colony using a
push-in cage (Laidlaw 1979). Queens were manually released from the
push-in cages after two days.
The experiment consisted of two separate trials. Trial 1 evaluated
colony treatments 2 and 4 only. Trial 2 evaluated all four colony
treatments simultaneously. Trial 2 was started with a new supply of
worker bees. Some of the queens used in trial 2 had also been evaluated
in trial 1.
Egg laying rates were determined by removing the frames from each
colony every 24 hours and counting the number of eggs. The removed
frames were immediately replaced with empty frames in order to minimize
disturbance. The frames with eggs were stored in a freezer until the
eggs were counted. Initial egg laying rates that were more than two
standard deviations from the mean were not used because these rates may
have occurred before queen maturation was complete. Experimental
colonies with Africanized worker bees (trial 2) began absconding

99
(abandoning the hive) after five to six days because of the disturbance
caused by the experimental procedure. The experiment was terminated at
that point and only egg laying rates prior to the beginning of
absconding were compared.
Brood Production Rates during First Brood Cycle
The same queens evaluated in the egg laying experiment were used
for this experiment. They were maintained in separate cages in a queen
storage colony (Laidlaw 1979; Reid 1975) for six days between
experiments. Four colony treatments were established as described
above, except that the hives were larger and were stocked with more
worker bees. Standard Langstroth hive bodies (48 liters) were used with
nine frames (all with European comb cell size): eight empty, drawn
combs and one filled with honey. All frames were weighed prior to being
put into the colonies in order that weight changes could be monitored.
Different geometric designs were placed above the entrances to
facilitate orientation and reduce drift between colonies.
As described for the first experiment, young worker bees were
collected and maintained in screened cages for two days prior to the
beginning of the experiment. Approximately 1200 grams of worker bees
were used to stock each colony. The number of bees put into each colony
was estimated by determining the mean individual bee weight from three
20-30 bee samples for each cage and then dividing the total weight of
introduced bees by the mean individual bee weight (Otis 1982b).
Queens were introduced into the experimental colonies using push-in
cages and manually released two days later in order to standardize the
starting day (day 0) for all experimental colonies. Queens that had
been in Africanized colonies for the daily egg laying rate experiment

100
were now introduced into European colonies; queens that had been in
European colonies during the first experiment were now introduced into
Africanized colonies.
After day 12 of oviposition, each experimental colony was inspected
and the amounts of unsealed brood (eggs and larvae) and sealed brood
(pre-pupae and pupae) for each colony were determined. To facilitate
measuring the amounts of unsealed and sealed brood, a 2.5 x 2.5 cm grid
was placed over each frame and the amount of brood within each square
was estimated. The number of developing brood cells was determined by
multiplying the brood area (in cm^) by 4.25 (the number of comb cells
O
per crrr). The number of worker bees present at day 12 was estimated by
assuming a constant rate of mortality of adults from day 0 to day 17.
At the end of day 17 of oviposition, colonies were closed and
killed with potassium cyanide. Unsealed and sealed brood, numbers of
adult bees, amount of pollen, and frame weights were determined.
The number of adult bees present at day 17 was estimated by
determining the mean individual bee weight from three samples of 150-200
bees taken from each colony. The total weight of bees in each colony
was then divided by the mean individual weight to get an estimate of the
total number of bees in each colony (Otis 1982b). The accuracy of this
technique was determined by comparing the estimates with the actual
counts (Table D1). A mean difference of only 1.5% was observed.
Data were analyzed using the Mann-Whitney U test (alpha = 0.05).
Correlations were evaluated using Spearman rank correlation coefficients
(alpha = 0.05).

101
Results
Daily Egg Laving Rates
There were no significant differences in daily egg laying rates
between Africanized and European queens (Tables 6-1 to 6-5). There
were, however, significant differences between individual queen lines
both between the two populations and within each population (Table 6-6).
There was no significant effect of worker bee type (Africanized vs.
European) on egg laying rates for either Africanized or European queens
(Table 6-5). Worker bee type also had no effect on daily egg laying
rates of sister queens (Table 6-7).
Brood Production Rates during First Brood Cycle
The amounts of unsealed, sealed and total brood at day 12 and at
day 17 for each of the experimental colonies are presented in Table 6-8.
There was no significant effect of worker bee population type on brood
production at either day 12 or day 17. When all colony treatments were
combined, total brood produced at day 12 was significantly correlated
with total brood produced at day 17 (P<0.05).
Changes in worker bee population for each colony are shown in Table
6-9. The estimated daily mortality rates for Africanized worker bees
were not significantly different than those for European worker bees.
The numbers of unsealed brood, sealed brood and total brood at day
12 and day 17 expressed as percentages of the adult population are
presented in Table 6-10. There was no significant difference between
Africanized and European worker bees.
Sister queens are compared in Table 6-11. The performance of the
European queen pair was similar with either Africanized and European
worker bees. One sister of Africanized queen pair (A62) performed

102
better with Africanized bees and one sister of Africanized queen pair
(A57) performed better with European bees.
Egg laying rates for queens evaluated in the daily egg laying
experiment were compared with estimated egg laying rates derived from
the brood production experiment (Table 6-12). There was no correlation
between daily egg laying rates with either the estimated daily egg
laying rates for the first 12 days or the estimated daily egg laying
rates for 17 days. Estimated daily egg laying rates at day 12 was
significantly correlated to the estimated overall egg laying rates at
day 17 (P<0.05).
Adequate pollen and nectar resources were available during the
experiment. Each colony stored pollen and had an overall weight gain.
There was no significant difference between colonies with Africanized or
European worker bees with respect to pollen stored or weight gained.
The amount of pollen stored by each colony was not significantly
correlated with colony weight gain, total brood produced, or mean
estimated daily mortality. Colony weight gain was positively correlated
(P<0.05) with total brood produced and negatively correlated (P<0.05)
with mean estimated daily mortality.
Discussion
The purpose of these experiments was to compare egg laying rates
between Africanized and European queen honey bees and to determine if
Africanized and European worker bees differentially affect brood
production and/or the queen's egg laying behavior. Results from daily
egg laying rates indicate that there was no significant difference
between Africanized and European queens during the initial colony growth
period under identical experimental conditions in Venezuela. There was

103
also no significant difference between Africanized and European worker
bees on brood production rates or egg laying rates of either Africanized
or European queens.
Differences in egg laying and brood production rates could be
evaluated by eliminating potential differences in foraging success
between Africanized and European bees by providing surplus honey to each
experimental colony and by controlling comb cell size. Differences in
brood production caused by differences in resource utilization
efficiency as a result of either smaller bee size or increased foraging
success were therefore, not evaluated. For a given amount of food, a
greater number of smaller, Africanized bees can be produced compared
with larger, European bees (Fletcher and Tribe 1977a; Tribe and Fletcher
1977). However, this advantage for Africanized bees with respect to
their smaller size, would only be present if resources were limited.
Therefore, surplus honey was provided for each colony to reduce the
effects of limited resources and the effects of differential foraging
success between the two honey bee populations. Controlling comb cell
size eliminated brood production differences based on bee size. For
example, a given number of nurse bees may be able to rear more smaller
bees than larger ones. In order that queen-worker bee interactions
could be evaluated, European comb size was selected because of the
difficulties both European queens and worker bees have with Africanized
comb as discussed earlier.
Egg laying rates observed during these experiments were within the
range reported for Africanized bees in French Guiana (Winston and Taylor
1980) but lower than the maximum reported for either population
(Fletcher 1978; Ribbands 1953). Several factors can account for this

104
difference. First, the experimental colonies used in this study may
have been only one-tenth the size of the managed, production colonies in
which the maximum rates were observed. Because egg laying rates are
correlated with the number of worker bees in the colony (Moeller 1958),
the lower egg laying rates may have been a result of smaller colonies.
Second, the intensity of colony disturbance, particularly during the
daily egg laying experiment, would have reduced egg laying rates and
increased egg and larval mortality. Third, egg laying rates for
artificially inseminated queens may be lower than for naturally mated
queens (Harbo and Szabo 1984). The purpose of this study was not to
determine absolute egg laying rates but to compare egg laying and brood
production rates for Africanized and European queens during the initial
colony growth phase under identical conditions.
There were no differences in egg laying and brood production rates
for Africanized and European bees during the initial colony growth
phase. When colonies increase in size and approach their maximum growth
phase and queens are maximally challenged, there may be a difference
between Africanized and European queens and colonies. However,
comparisons of queen pupal weights (as a correlate of egg laying rates)
suggest that there would be no difference between Africanized and
European queens with respect to potential egg laying capacity (Chapter
V).
Colony growth rates and therefore reproductive rates are affected
by two other demographic parameters: adult longevity and brood
mortality. Winston and Katz (1981) found that European worker bees were
longer lived than Africanized worker bees under identical conditions in
Venezuela (26.3 compared with 22.7 days). This difference would give

105
European colonies a growth rate advantage. Unfortunately, brood
mortality for both Africanized and European honey bees during the
initial colony growth phase has not been investigated under identical
conditions.
Based on the results from these studies and those evaluating other
colony demographic parameters, e.g., worker development times (Chapter
II) and queen maturation rates (Chapter IV), it must be concluded that
differences in reproductive rates between Africanized and European honey
bees in South America cannot be attributed to intrinsic demographic
factors. Reproductive rates in honey bees are a function of at least
two other factorsresource availability and resource utilization
efficiency. Chapter VIII presents a hypothesis to explain the success
of Africanized honey bees based on differences in resource utilization
efficiency. This hypothesis is based on differences between Africanized
and European bees with respect to foraging behavior, brood production
efficiency as a function of bee size, and resource-induced absconding.

106
TABLE 6-1. Daily egg laying rates of Africanized and European queens
with European nurse bees trial 1.
ONE-DAY HIGH
MEAN + SD (n)
AFRICANIZED QUEENS
A57 (W42)
550
473.2 + 68.5 (6)
A62 (W81)
857
825.0 + 47.0 (3)
EUROPEAN QUEENS
YK (Y4)
951
922.0 + 41.0 (2)
YD5 (Y42)
763
689.6 + 66.4 (5)
GK30 (Y63)
710
636.7 + 80.8 (3)
GK30 (Y64)
705
575.6 + 101.3 (5)
ANALYSIS3
NS
^ann-Wh itney U test, two-tailed, alpha = 0.05; evaluated for all
samples (n = 24).

107
TABLE 6-2. Daily egg laying rates of Africanized and European queens
with European nurse bees, trial 2.
ONE-DAY HIGH MEAN + SD (n)
AFRICANIZED QUEENS
A57 (W42)
A62 (W81)
EUROPEAN QUEENS
GK30 (Y63)
740 699.8 +39.5 (4)
953 915.8 + 30.7 (4)
779 736.8 + 32.2 (4)
ANALYSIS3
NS
^iann-Whitney U test, two-tailed, alpha = 0.05; evaluated for all
samples (n = 12).

TABLE 6-3. Daily egg laying rates of Africanized and European
queens with Africanized nurse bees, trial 2.
ONE-DAY HIGH MEAN + SD (n)
AFRICANIZED QUEENS
A61 (W61)
847
799.5
+ 67.2
(2)
A57 (W41)
725
694.0
t 39.1
(4)
A62 (W85)
883
798.7
+ 92.3
(3)
EUROPEAN QUEENS
GK30 (Y61)
850
823.5
+ 37.0
(4)
YD5 (Y52)
949
782.7
+ 231.7
(3)
ANALYSIS3
NS
^ann-Wh itney U test, two-tailed, alpha = 0.05; evaluated
for all samples (n = 16).

TABLE 6-4. Effect of nurse bee genotypes on the daily
egg laying rates of Africanized and European
queens, trial 2.
MEAN + SD (n)
AFRICANIZED NURSE BEES
AFRICANIZED QUEENS
A61 (W61)
799.5
+
67.2
(2)
AS7 (W41)
694.0
+
39.1
(4)
A62 (W85)
798.7
+
92.3
(3)
COMBINED
752.3
+
79.6
(9)
(A)
EUROPEAN QUEENS
GK30 (Y61)
823.5
+
37.0
(4)
YD5 (Y52)
782.7
+
231.7
(3)
COMBINED
806.0
+
138.0
(7)
(B)
COMBINED AFRICANIZED
AND EUROPEAN QUEENS
775.8
+
108.4
(16)
(C)
EUROPEAN NURSE BEES
AFRICANIZED QUEENS
A57 (W42)
699.8
+
39.5
(4)
A62 (W81)
915.8
+
30.7
(4)
COMBINED
807.8
+
120.0
(8)
(D)
EUROPEAN QUEEN
GK30 (Y63)
736.8
32.2
(4)
(E)
COMBINED AFRICANIZED
AND EUROPEAN QUEENS
784.1
+
103.3
(12)
(F)

110
TABLE 6-5. Analyses of the effect of queen genotypes and nurse bee
genotypes on the daily egg laying rates of Africanized and
European honey bees. Letters represent the different
treatments presented in Table 6-4. Mann-Whitney U test,
two-tailed, alpha = 0.05 (Siegel 1956).
HI: There is no difference in egg laying rates between Africanized
and European queens.
i.Africanized nurse bees
A x B NS
ii.European nurse bees
D x E NS
iii.Africanized and European
nurse bees combined
(A + D) x (B + E) NS
H2: There is no differential effect of Africanized and European
worker bees on egg laying rates.
i.Africanized queens
A x D NS
ii.European queens
B x E NS
iii.Africanized and European
queens combined
C x F
NS

TABLE 6-6. Daily egg laying rates of Africanized and European queens
comparison between genotypes within each population.
MEAN + SD (n) ANALYSES
AFRICANIZED QUEENS
1. TRIAL 1EUROPEAN NURSE BEES
A57 (W42)
473.2
+
68.5
(6)
A62 (W81)
825.0
+
47.0
(3)
P<0.05
2.
TRIAL 2EUROPEAN NURSE BEES
A57 (W42)
699.8
+
39.5
(4)
A62 (W81)
915.8
+
30.7
(4)
P 3.
TRIAL 2AFRICANIZED NURSE BEES
A61 (W61)
799.5
+
67.2
(2)
A57 (W41)
694.0
+
39.1
(4)
A62 (W85)
798.7
+
92.3
(3)
NSb
4.
COMBINING §2 AND #3
P. EUROPEAN QUEENS
1.
TRIAL 1EUROPEAN NURSE BEES
YK (Y4)
922.0
+
41.0
(2)
*c
YD5 (Y42)
689.6
+
66.4
(5)
GK30 (Y63)
636.7
+
80.8
(3)
GK30 (Y64)
575.6
+
101.3
(5)
NSb
2.
TRIAL 2EUROPEAN NURSE BEES
GK30 (Y63)
736.8
+
32.2
(4)
3.
TRIAL 2AFRICANIZED NURSE BEES
GK30 (Y61)
823.5
+
37.0
(4)
YD5 (Y52)
782.7
+
231.7
(3)
NSa
4.
COMBINING #2 AMD #3
NSb
Mann-Whitney U test, two-tailed, alpha = 0.05.
bKruskal-Wal1 is one-way analysis of variance by ranks, alpha = 0.05.
CYK (Y4) was significantly different, P<0.05, from other genotypes in
the group when evaluated by pairs using the Mann-Whitney U test,
two-tailed, alpha = 0.05.

112
TABLE 6-7. Comparison of daily egg laying rates for sister
Africanized and European nurse bees. Mean + SD
size).
queens with
(n = sample
AFRICANIZED
NURSE BEES
EUROPEAN
NURSE BEES
ANALYSES3
AFRICANIZED QUEENS
A57 (W41)
A57 (W42)
694.0 +39.1 (4)
699.8 + 39.5 (4)
NS
A62 (W85)
A62 (W81)
798.7 + 92.3 (3)
915.8 +30.7 (4)
NS
EUROPEAN QUEENS
GK30 (Y61)
GK30 (Y63)
823.5 +37.0 (4)
736.8 + 32.2 (4)
NS
^lann-Whitney U test, two-tailed, alpha = 0.05.

113
O
TABLE 6-8. Comparison of brood production in cm for Africanized and
European queens during two periods of the first brood cycle.
DAY 12
DAY 17
USB3 SBb TBC
USB SB TB
AFRICANIZED NURSE BEES
AFRICANIZED QUEENS
A62 (W81) 1053
A57 (W42) 416
EUROPEAN QUEEN
GK30 (Y63) 1775
EUROPEAN NURSE BEES
AFRICANIZED QUEENS
A62 (W85) 906
A57 (W41) 1481
EUROPEAN QUEEN
GK30 (Y61) 1614
962
666
2015
1082
1484
493
1301
818
2785
1311
819
2594
1330
1756
3086
823
487
1729
1968
434
1018
1354
1256
1788
2274
728
2342
1580
1519
3099
ANALYSES^
AFR WORKERS x EUR WORKERS NS NS NS NS NS NS
3USB = unsealed brood (eggs and larvae).
bSB = sealed brood (pre-pupae and pupae).
^TB = total brood.
dMann-Whitney U test, one-tailed, alpha = 0.05. AFR = Africanized;
EUR = European.

114
TABLE 6-9. Colony adult population changes during first brood cycle.
ESTIMATED DAILY
DAY 0a DAY 12b DAY 17c MORTALITY RATEd
AFRICANIZED NURSE BEES
AFRICANIZED QUEENS
A62 (W81)
A57 (W42)
EUROPEAN QUEEN
GK30 (Y63)
EUROPEAN NURSE BEES
AFRICANIZED QUEENS
A62 (W85)
AS7 (W41)
EUROPEAN QUEEN
GK30 (Y61)
10,169
10,169
4,850
4,408
10,169
6,511
10,435
10,435
5,375
6,102
10,435
6,321
2,634
2,007
443
480
4,987
305
MEAN = 409
3,267
4,297
422
361
4,607
343
MEAN = 375
ANALYSIS
NS
^Population at Day 0 was estimated as described in methods.
Population at Day 12 estimated by: Day 0 [(Day 0 Day 17)(12/17)].
^Population at Day 17 was estimated as described in methods.
(Day 0 Day 17)/17.
^iann-Whitney U test, one-tailed, alpha = 0.05.

115
TABLE 6-10. Brood production during first brood cycle expressed as a
percent of adult population.
DAY 12
DAY 17
USB3 SBb TBC USB SB TB
AFRICANIZED NURSE BEES
AFRICANIZED QUEENS
A62 (W81) 92.3
A57 (W42) 40.1
EUROPEAN QUEEN
GK30 (Y63) 115.9
EUROPEAN NURSE BEES
AFRICANIZED QUEENS
A62 (W85) 71.6
A57 (W41) 103.2
EUROPEAN QUEEN
GK30 (Y61) 108.5
84.3
64.2
176.6
104.3
239.4
104.4
209.9
173.2
449.3
274.6
53.4
169.3
113.3
149.6
262.9
65.1
33.9
136.7
137.1
56.4
100.7
176.1
124.2
232.5
224.9
48.9
157.4
145.8
140.1
285.9
ANALYSES^
AFR WORKERS x EUR WORKERS NS NS NS NS NS NS
aUnsealed brood (eggs and larvae); colony adult population estimated
for Day 12 and Day 17 as described in methods.
Sealed brood (pre-pupae and pupae).
^Total brood.
Mann-Whitney U test, one-tailed, alpha = 0.05. AFR = Africanized;
EUR = European.

TABLE 6-11. Comparison of overall egg laying rates
between Africanized and European sister
queens with Africanized and European
nurse bees during the first brood cycle.
AFRICANIZED EUROPEAN
NURSE BEESa NURSE BEES
AFRICANIZED QUEENS
A62 (W81)
696

A62 (W85)

447
A57 (W42)
328
A57 (W41)

568
EUROPEAN QUEENS
GK30 (Y63)
772
__
GK30 (Y61)
775
aOverall egg laying rate = (TBjj/17)(4.25).

117
Table 6-12. Comparison of egg laying rates during daily egg laying
rate experiment and brood production experiment.
AFRICANIZED EUROPEAN
NURSE BEES NURSE BEES
QUEENS
ELRa
BP12b
BP17C
ELR
BP12
BP17
AFRICANIZED
A62 (W81)

714
696
916


A62 (W85)
799



612
447
A57 (W42)

383
328
700
__
A57 (W41)
694



697
568
EUROPEAN
GK (Y63)

918
772
737
GK (Y61)
824

*
829
775
ANALYSES^
ELR
X
BP12
NS
ELR
X
BP 17
NS
BP12
X
BP 17
P<0.05
aDaily egg laying rate experiment, means.
bBrood production experiment; egg laying rate estimated for first
12 days of brood cycle by dividing total brood at day 12 by 12.
cBrood production experiment; egg laying rate estimated for first
17 days of brood cycle by dividing total brood at day 17 by 17.
dSpearman rank correlation, alpha = 0.05; Africanized and
European nurse bees combined.

CHAPTER VII
SUCCESSFUL HYBRIDIZATION BETWEEN AFRICANIZED AND EUROPEAN HONEY BEES
IN VENEZUELA WITH IMPLICATIONS FOR NORTH AMERICA
Introduction
In 1956 African honey bees, Apis mel1 ifera scutel1 ata, formerly
classified as A. m. adansonii (Ruttner 1976a, 1976b, 1981), were
imported into southeastern Brazil (Kerr 1967). Their hybridized
descendents, known as Africanized honey bees (Goncalves 1982), have
rapidly spread throughout tropical South and Central America as far
north as Honduras and El Salvador (Rinderer 1986). The dispersion from
their original importation site into new areas has been rapid200-500
km per year (Taylor 1977, 1985; Winston 1979a). As Africanized honey
bees have dispersed into new areas, they have rapidly increased in
number (Otis 1982a) and have attained dramatic population densities
(Michener 1975): 4-8 colonies/km^ (Taylor 1985), or as high as 107.5
colonies/km in the cerrado habitats in the Brazilian states of Goias
and Mato Grosso (Kerr 1971 cited in Michener 1975). There are now
probably more than ten million feral colonies of Africanized honey bees
in South and Central America (Winston, Taylor and Otis 1983). Their
success in these habitats, compared with European populations of honey
bees, may be attributed to their foraging behavior which is more suited
to the resource patterns of the tropics (Nunez 1973, 1979a, 1982;
Rinderer, Bolten, Collins and Harbo 1984; Rinderer, Collins and Tucker
1985; Winston and Katz 1982). As a result of both their foraging
118

119
success and length of time throughout the year that resources are
available in the tropics, Africanized honey bees have a high annual
reproductive rate, which is responsible for both their rate of dispersal
into new areas and high colony densities. Net reproductive rates for
Africanized bees have been estimated to be 16 colonies per colony per
year based on demographic data collected in French Guiana (Otis 1980,
1982a) compared with 0.92-0.96 (Seeley 1978) or 3-3.6 when afterswarms
are counted (Winston 1980a; Winston, Taylor and Otis 1983) for European
honey bees in North America.
Particularly in the region of their importation (southeastern
Brazil), there has been ample opportunity for hybridization with both
managed and feral European honey bees [primarily A, m. me!1ifera and A.
m. 1igustica which had been imported into Brazil by 1845 (Gerstaker
cited in Pellet 1938; Woyke 1969)]. However, despite opportunity for
hybridization, Africanized honey bees have maintained behavioral,
chemical and morphological characteristics similar to their African
parental population and distinguishable from European honey bees:
colony defense behavior (=stinging behavior) (Collins, Rinderer, Harbo
and Bolten 1982; Stort 1974, 1975a, 1975b, 1975c, 1976); reproductive
rates (Fletcher 1978; Fletcher and Tribe 1977a; Otis 1980, 1982a);
absconding behavior (reviewed by Fletcher 1978; Winston, Otis and Taylor
1979; Winston, Taylor,and Otis 1983); foraging and hoarding behavior
(Nunez 1973, 1979a, 1982; Rinderer, Bolten, Collins and Harbo 1984;
Rinderer, Bolten, Harbo and Collins 1982; Rinderer, Collins and Tucker
1985; Winston and Katz 1982); worker bee longevity (Winston and Katz
1981); development times (Chapters II and IV; Harbo, Bolten, Rinderer
and Collins 1981); selection preferences for nest cavity sizes

120
(Rinderer, Collins,. Bolten and Hanbo 1981; Rinderer, Tucker and Collins
1982); allozyme patterns (Nunamaker and Wilson 1981; Sylvester 1982);
cuticular hydrocarbon composition (Carlson and Bolten 1984, and
unpublished data); adult bee size and comb cell size (Chapter III;
Michener 1975); and morphometric relationships (Daly and Balling 1978).
This apparent lack of evidence for hybridization has been
attributed primarily to some degree of reproductive isolation between
the Africanized and European populations (Kerr and Bueno 1970; Taylor
1985). Three isolating mechanisms have been suggested: assortative
mating (Kerr and Bueno 1970); physiological incompatibility with respect
to the drone ejaculation response (Kerr and Bueno 1970); and differences
in drone and presumably the queen flight times between the Africanized
and European populations (Taylor 1985; Taylor, Kingsolver and Otis in
press). At best, these mechanisms may be only partially effective and
are not likely to account for the apparent lack of hybridization. For
example, Kerr and Bueno (1970) present data to support assortative
mating even though 35% and 42% of the matings evaluated were hybrid.
With respect to differences in queen and drone flight times, data from
Venezuela suggest that mean peak drone flight times for Africanized and
European populations are separated by only 23 minutes and that drones
from both populations are present in the mating areas at all times
during the approximately three hour flight period (Taylor, Kingsolver
and Otis in press).
A more probable argument for the maintenance of the African
characteristics is based on selection advantages for the African
genotype in tropical habitats of South America, which are characterized
by resource distribution patterns similar to the ones in Africa where

121
these bees evolved. Two lines of evidence support this selectionist
argument. First, the foraging behavior of the Africanized honey bees,
characterized by solitary foraging and less colony recruitment, would be
more adaptive in tropical areas with rich, but dispersed, resources
(Rinderer, Bolten, Collins and Harbo 1984; Rinderer, Collins and Tucker
1985). Second, there is much historic evidence that European honey bees
have not been successfully maintained (probably due to starvation) in
many areas of tropical South America that are now densely populated with
Africanized honey bees (Bolten, personal observation; Michener 1972;
Winston, Taylor and Otis 1983).
The selectionist argument allows for hybridization between
Africanized and European populations, with the African genotype being
selected for under the physical and biotic parameters characteristic of
tropical areas. Selection for the African genotype would then account
for the present population in South and Central America being
behaviorally, chemically and morphologically similar to the African
parental type.
These two alternative hypotheses for the maintenance of the African
parental characteristicsreproductive isolation between the Africanized
and European honey bee populations versus selection for the African
genotype in tropical regionssuggest different scenarios for the
potential impact of Africanized honey bees in North America. One
scenario resulting from reproductive isolation would limit Africanized
honey bees in their northern movement because of their inability to
overwinter (Taylor 1985; Taylor and Spivak 1984). This is based on the
observation that the parental African population as well as Africanized
bees do not have the thermoregulatory capabilities to survive the cold

122
temperatures of temperate winters (Nunez 1979b; Woyke 1973). The other
scenario based on successful hybridization and resultant genetic
introgression would have the stinging behavior of the Africanized honey
bees and the potential public health hazard widespread in North America
and not limited to the southern regions.
A comparison between hybrid and non-hybrid mating success is
lacking for the Africanized and European populations of honey bees.
Although reciprocal FI crosses (Africanized queen x European drone;
European queen x Africanized drone) can be successfully produced with
artificial insemination (Chapater III), natural matings have been
difficult to evaluate because of the inability to distinguish between
hybrid and non-hybrid progeny on the individual level. It is not
possible to determine which drones have mated a queen because honey bees
mate in the air away from the hive, and there are no available genetic
markers in the Africanized population. Identifying individual honey
bees as either Africanized or European is not presently possible
(Carlson and Bolten 1984; Page and Erickson 1985; Rinderer and Sylvester
1981). However, there is promise that DNA analyses will allow
individuals to be identified (Hall in press).
This paper evaluates an important parameter with respect to the
question of successful hybridization between Africanized and European
honey bees: the mating success of both Africanized and European queens
with Africanized drones. The mating success of European queens x
Africanized drones was compared with the mating success of Africanized
queens x Africanized drones in an isolated area in Venezuela with no
European honey bees but with a feral population of Africanized honey
bees. Because of present legal restrictions, Africanized bees cannot be

123
taken into areas with only European honey bees so the reciprocal cross
was not evaluated. However, evaluating the success of the European
queen x Africanized drone cross is important because it represents the
most probable initial hybridization that will occur when Africanized
honey bees invade North America (Mexico and U.S.A.).
Methods
The Africanized queen mother (A26) was removed from a feral colony
in eastern Venezuela where there were no known European honey bees. The
colony was identified as Africanized by its behavior and small comb cell
size characteristic of the Africanized population (4.5-5.0 mm between
opposite sides of the hexagon, Chapter III). The European queen mother
(L13) was produced by a commercial queen producer in the southeastern
U.S.A. and shipped to Venezuela.
Experimental queens were produced from each queen mother by the
standard queen rearing technique of transferring (grafting) 12 to 18-
hour-old larvae into artificial queen cells that were then introduced
into queen-cell producing colonies (Laidlaw 1979). After the queen
cells were sealed, each cell was protected by a wire mesh cylinder (mesh
size = 3.0 mm). Virgin queens were allowed to emerge in the cell-
producing colonies. Newly emerged virgins were marked for
identification and then stored in a strong, queenless colony. The
following day they were introduced into individual, five-frame colonies
with Africanized worker bees from which natural matings could occur.
Queens were released into these mating colonies using standard three-
hole mailing cages (Laidlaw 1979). Based on earlier calculations,
natural release from the mailing cages was estimated to take 2.5-3.0
days. Marked, virgin queens were introduced into mating colonies rather

124
than mature queen cells so that the identity of the experimental queens
would later be certain. The mating colonies were located in an area in
eastern Venezuela where there was no known European honey bees, but
which was densely populated with feral Africanized colonies.
Eighteen days after introduction into mating colonies, the queens
were collected, and the spermatazoa in their spermatheca were counted
using hemacytometers (Mackensen and Roberts 1948; Mackensen and Tucker
1970). The criterion for mating success was the number of spermatozoa
in the spermatheca. In addition, the age of the queen when oviposition
first started was calculated by determining the age of the oldest brood
in each colony.
Results
Numbers of spermatozoa counted in the spermatheca of Africanized
and European queens are summarized in Table 7-1. The mean number of
spermatozoa in Africanized queens (4.09 +0.50 million) was not
different from the mean number in European queens (4.12 +0.58 million;
t-test, two-tailed, alpha = 0.05). European queens began oviposition on
the 10th day post-emergence, one day sooner than the Africanized queens
(P<.001; Kolmogorov-Smirnov two-tailed test, chi-square distribution, df
= 2). The time from adult emergence to initiation of oviposition
reported here (Table 7-1) is longer than the maturation interval
reported in Chapter IV, which may be a result of having introduced
virgin queens into the mating colonies rather than mature queen cells.
There was no correlation between the time post-emergence to initiation
of oviposition and the number of spermatozoa in the spermatheca
(Spearman's rank correlation coefficient, two-tailed, alpha = 0.05).
The acceptance of the Africanized and the European virgin queens

125
introduced into the mating nuclei was 50% and 61% respectively. There
was no significant difference in acceptance (Fisher's exact probability
test, alpha = 0.05).
Discussion
Evidence for Hybridization
There appears to be no effective reproductive isolating mechanism
operating between the Africanized and European populations. The mating
of both Africanized and European queens with Africanized drones was
equally successful as judged by the number of spermatozoa in the
spermatheca. Offspring from the hybrid crosses were viable with no
apparent difference in mortality as determined by the uniformity of the
brood pattern. Kerr and Bueno (1970) report that there may be a
difference in ejaculation response between Africanized and European
drones that may provide a potential isolating mechanism. Even if this
exists, European queens were still able to successfully mate with
Africanized drones without any apparent problem, as determined by both
the numbers of spermatozoa in their spermatheca and the age when
oviposit ion began.
Although the same queen pheromone is produced by three sympatric
Asiatic species of Apis (Butler, Calam and Callow 1967; Shearer, Boch,
Morse and Laigo 1970), reproductive isolation occurs between the three
species because there is no overlap in times of drone flight (Koeniger
and Wijayagunasekera 1976). The situation between Africanized and
European populations of honey bees is quite different with respect to
the time of flight of the queens and drones. Data from Venezuela show
that flight times for Africanized and European drones completely overlap
during the approximately three hours of mating flight activity with only

126
23 minutes separating the mean times of peak flight activity for each
population (Taylor Kingsolver and Otis in press). This difference does
not provide a satisfactory mechanism for reproductive isolation between
the Africanized and European honey bee populationsparticularly because
any unfavorable climatic conditions e.g. high winds cloudiness high
humidity, or rain (Gary 1975), cause mating flights of reproductives
from both populations to more completely converge to times of favorable
weather conditions. The data on reproductive success (determined by the
number of spermatozoa in the spermatheca) of European queens mating with
Africanized drones presented in this study demonstrate that any
differences in mean peak flight times did not effectively prevent
hybridization of European queens with Africanized drones.
Evidence that extensive hybridization has already occurred between
the introduced African honey bees and the previously established
European honey bees can be demonstrated by the increase in genetic
diversity in the Africanized population. For example, Adams, Rothman,
Kerr and Paulino (1977) concluded that the large increase in number of
sex alleles in the population of honey bees in southeastern Brazil is a
result of hybridization between African and European honey bees. Page
and Erickson (1985) also suggest that hybridization has occurred based
on the variation in behavior and appearance of Africanized bees in
Venezuela.
Impact of Hybridization
Demonstration that successful hybridization can and does occur has
important implications with respect to the potential impact the
Africanized honey bee will have in North America. First is the negative
impact that hybridization would have. The parental African population

127
as well as Africanized bees may not have the thermoregulatory
capabilities to survive the cold temperatures of temperate winters
(Nunez 1979b; Woyke 1973). Based on the temperature limits of the
parental population, Taylor (1985) and Taylor and Spivak (1984)
predicted the northern limits of Africanized honey bees in North
America. However, Africanized honey bees may acquire, through
hybridization with European honey bees in Mexico and southern U.S.A.,
the ability to overwinter farther north than is presently expected.
That is, the overwintering genome of the European honey bees may become
introgressed into the Africanized genome. Or, the corollary, the
stinging behavior characteristic of the Africanized honey bees may
become introgressed into the overwintering European population.
Successful genetic introgression of these traits may not be a rapid
process because these traits are polygenic and/or may involve coadapted
genomes. However, because hybridization occurs, the potential for
successful genetic introgression exists and must be considered.
Hybridization may, therefore, result in the stinging behavior of the
Africanized honey bees becoming a potential public health hazard
throughout North America, not just in the warmer southern regions. That
this may be the unfortunate outcome of hybridization is supported by
recent investigations in Argentina, which have demonstrated that
Africanized honey bees are distributed farther south than predicted
based on temperature limits of the parental population (Dietz, Krell and
Eischen 1985; Krell, Dietz and Eischen 1985).
The U.S. Department of Agriculture Economic Research Service has
recently evaluated the potential impact of Africanized honey bees in the
U.S.A. (McDowell 1984). Unfortunately this report does not consider the

128
possibility that further hybridization between Africanized and European
honey bees might result in the stinging behavior of Africanized honey
bees becoming established throughout the northern regions of the U.S.A.
The economic consequences as well as public health hazards may be more
widespread throughout North America than previously thought. For any
solutions to the Africanized honey bee problem to be successful a
realistic assessment of the potential problem is necessary.
However, hybridization may also have a positive impact. Coupled
with selection favoring both the foraging and thermoregulatory behavior
of European honey bees in temperate regions, hybridization between
Africanized and European bees may have the positive effect of increasing
the rate at which African genes become rare in the population. There is
a large population of European honey bees, both managed and feral, in
North America (perhaps greater than 15 million colonies in the U.S.A.
alone), which is particularly dense in the south where the Africanized
bees will first enter the U.S.A. The invading Africanized population
would be quite small relative to the existing European population,
increasing the frequency of hybridization and resultant "swamping" (or
diluting) of African genes.
The problem of Africanized honey bees may be reduced prior to their
entry into the U.S.A. because of both the potential for hybridization
and competition for available floral resources with European honey bees
in Mexico. Mexico has a larger population of European honey bees than
any other country in Latin America. When Africanized honey bees enter
Mexico, they will be entering a region that already has an extensive,
established population of European honey bees, both managed and feral
[2.6 million managed colonies alone (Zozaya cited in Taylor 1985)].

129
Competition with an established population of honey bees for limited
floral nectar and pollen resources will be much greater than Africanized
bees have previously experienced in any areas in South America. This
competition will greatly slow their dispersal. In many areas of Mexico
and southern U.S.A., pollen and nectar resources are already close to
being saturated by the existing honey bee population. In addition,
under temperate resource conditions, the foraging behavior of
Africanized honey bees (which is more appropriate to tropical resource
patterns) will be at a disadvantage relative to the foraging behavior of
the European honey bee population, which is characterized by greater
colony recruitment (Rinderer, Bolten, Collins and Harbo 1984; Rinderer,
Collins and Tucker 1985; Visscher and Seeley 1982).
The selective advantage of the foraging and/or thermoregulatory
behavior of European honey bees has been demonstrated in temperate
regions. African honey bee queens were introduced into North America
during the late 1800's and early 1900's when the beekeeping industry in
the U.S.A. was developing (Morse et al. 1973) and more recently, into
Louisiana (Cantwell 1974; Morse et al. 1973; Taber 1961). However, due
to hybridization and selection against the African genotype, the impact
of these introductions of African bees is undetectable today. There
have also been unsuccessful introductions of African and Africanized
bees into Europe (Cantwell 1974; Morse et al. 1973; Woyke 1973).
Therefore, these factorsa large, established population of European
honey bees, and both a foraging and thermoregulatory behavior in
European bees better adapted to temperate conditionsprecludes using
South and Central America as a model for North America in predicting the
impact, as well as the rate of spread, of Africanized honey bees.

130
Kin recognition has been suggested as another mechanism that may
help preserve the African genotype in the hybridized honey bee
population in South and Central America (Hall in press). However, data
presented in Table 4-2 demonstrate that this is unlikely because both
Africanized and European worker bees reared both Africanized and
European queens with equal frequency.
There have been several suggestions that Africanized drones may
have a mating advantage over European drones (Kerr and Bueno 1970;
Michener 1975; Morse 1984; Rinderer 1986; Rinderer, Hellmich, Danka and
Collins 1985; Taylor 1985). Taylor (1985) suggests that this mating
advantage would reduce hybridization. On the contrary, a mating
advantage for Africanized drones would increase the rate of
hybridization between Africanized and European honey bee populations.
When Africanized honey bees begin invading North America (Mexico and
U.S.A.), they will be greatly outnumbered by the established European
honey bee population. With a mating advantage, the frequency of
European queen x Africanized drone matings would be greater than
expected based solely on the relative frequency of each population,
thereby resulting in greater hybridization.
Unfortunately, Africanized honey bee research has been
characterized by conceptualizing the Africanized honey bee as a distinct
entity that is reproductively isolated (Taylor 1977, 1985; Taylor and
Spivak 1984) rather than as a population within a species fully capable
of hybridization. Clearly, the Africanized honey bee is not a species
invading a new habitat (North America) that is free of competition from
conspecifics. Thus, the spread of Africanized bees (African genes) in
temperate North America will be: 1) farther north than predicted by the

131
geographic limits of the parental population because of hybridization
and resultant genetic introgression; 2) slowed considerably by
competition for available resources by an established population of
European honey bees; 3) swamped through hybridization with a more
numerous, established population of European honey bees; 4) at a
disadvantage with respect to foraging behavior; and 5) limited by
selection against those colonies that have not acquired through
hybridization the ability to overwinter.
There has been a general lack of support for the selectionist
argument for the maintenance of the African genotype in favor of the
hypothesis of reproductive isolation. With the demonstration that
hybridization is successful, coupled with the recent observations of the
distribution patterns of Africanized honey bees in Argentina (Dietz,
Krell and Eischen 1985; Krell, Dietz and Eischen 1985), further
consideration of the selectionist argument for the maintenance of the
African characteristics in the Africanized honey bee in South and
Central America is necessary.

132
TABLE 7-1. Mating success of Africanized and European honey bee queens.
TIME TO
NO. SPERM
OVIPOSITION3
(x 106)b
CORRELATIONS'
AFRICANIZED
11
4.09 + 0.50
NS
GENOTYPE (A26)
(8)
(8)
EUROPEAN
10
4.12 + 0.58
NS
GENOTYPE (L13)
(11)
(11)
ANALYSES
P<0.001d
NSe
Median days post-emergence to initiation of oviposition (sample size).
One-day-old virgins were introduced into mating colonies.
Mean + SD (sample size) of spermathecal spermatozoa number.
^Spearmans rank correlation coefficient, two-tailed, alpha = 0.05.
Kolmogorov-Smirnov two-tailed test, chi-square distribution, df = 2.
et-test, two-tailed, alpha = 0.05.

CHAPTER VIII
DISCUSSION: FACTORS CONTRIBUTING TO THE SELECTION ADVANTAGE OF
AFRICANIZED HONEY BEES IN SOUTH AMERICA--
THE RESOURCE UTILIZATION EFFICIENCY HYPOTHESIS
Success of Introduced Populations of Honey Bees
Thirty years ago African honey bees, Apis me!1ifera scute!1 ata
[formerly classified as A. m. adansonii (Ruttner 1976a, 1976b, 1981)],
were introduced into southeastern Brazil (Kerr 1967). Offspring, known
as Africanized honey bees because of hybridization with European honey
bees (Goncalves 1982), have rapidly dispersed throughout South America,
sometimes achieving dramatically high population densities (Michener
1975; Taylor 1977, 1985). In 1982 Africanized honey bees entered Panama
(Buchmann 1982) and by 1986 were as far north as Honduras and El
Salvador (Rinderer 1986). The success and biological impact of
Africanized honey bees in these tropical and sub-tropical regions,
compared with the lack of success of European honey bees in these same
regions, is a result of a selection advantage for the Africanized (=
African) genotype in tropical resource and climatic conditions. The
difference in success between Africanized and European honey bees is
evidenced by the fact that
European bees in Brazil were never commonly found living wild
in the forests and countryside. This was especially true in
tropical forest regions, where honey bees were virtually
restricted to a few apiaries...Everyone questioned on the
matter emphasized the increase in bees away from apiaries that
occurred with the arrival of the Brazilian [Africanized] bees.
(Michener 1972, p. 15).
133

134
The selection advantage for Africanized bees may be a result of
behavioral and/or physiological characteristics that may include
differences in resource utilization and/or colony demography. It is not
surprising that Africanized honey bees are better adapted to tropical
conditions than are European honey bees, considering the former are
derived from imported African bees that evolved under similar tropical
and sub-tropical conditions in Africa. Fletcher (1978) has reviewed the
biological characteristics of the parental population of African honey
bees in Africa.
The spread and impact of Africanized honey bees in South America
must, however, be kept in perspective. European honey bees introduced
into North America early in the 17th century (Pellett 1938) dispersed
throughout North America, also achieving high population densities. In
general, honey bees are very successful not only in their native
habitats but in almost every region where they have been introduced.
Their success is based on a highly developed social system that allows
honey bees to: 1) develop large, perennial colonies that are able to
buffer climatic changes; 2) efficiently utilize resources because of
advanced communication and recruitment systems; and 3) defend against
both vertebrate and invertebrate predators because of their very
effective colony defense behavior.
The question with which we are concerned in these studies is not
what makes A. me!1 ifera more successful than other species nor what
impact introduced honey bees have on native pollinator communities (see
Roubik 1978, 1979, 1980, 1982, 1983; Roubik and Buchmann 1984). Nor is
it a question of comparing Africanized honey bees in tropical regions
with European honey bees in temperate regions (see Winston, Dropkin and

135
Taylor 1981 and Winston, Taylor and Otis 1983). Rather, the question
is: what are the differences between Africanized and European
populations of A. mel1ifera that make Africanized honey bees more
successful in tropical regions not only in South America but also in
Africa? European honey bees have not been successfully introduced into
tropical areas of Africa despite numerous attempts (Fletcher 1977b,
1978).
Factors Affecting Honey Bee Reproductive Rates
The success of Africanized honey bees in South Americaas judged
by their rate of dispersal and their population densities (Michener
1975; Taylor 1977, 1985)must surely be a result of a high reproductive
rate. What are the differences between Africanized and European honey
bees that allow for high reproductive rates in Africanized bees, and can
these differences account for the impact of Africanized honey bees?
This question does not involve identification or analysis of the
proximal factors that are responsible for initiating reproduction, but
does involve analysis of the components that affect the rate of
reproduction.
Reproductive rates in honey bees are a result of an interaction of
at least three factors, all of which affect colony growth rates:
resource availability, resource utilization efficiency (foraging
success, brood production efficiency, and bee size), and colony
demographic parameters (primarily queen fecundity and adult worker bee
longevity). Therefore, in order to evaluate reproductive differences
between Africanized and European honey bee populations, all three
factors need to be considered. Unfortunately, early research evaluated
reproductive rates of Africanized honey bees by comparing data for

136
Africanized honey bees from South America with data from studies of
European honey bees from North America, which not only were collected
under different resource conditions but also different experimental
conditions (Otis 1980, 1982a; Winston 1979b, 1980a; Winston, Dropkin and
Taylor 1981).
Results from these earlier studies characterized the Africanized
population as one with a dramatically high annual colony reproductive
ratefour to five times greater than European honey bees in temperate
regions (Otis 1980, 1982a; Winston 1980a; Winston, Taylor and Otis
1983). However, because these comparisons were not based on data
collected under similar environmental or experimental conditions, they
are inappropriate comparisons and cannot be used to identify either the
factors responsible for the difference in reproductive rates between the
two populations or the factors responsible for the success of
Africanized honey bees. Because the experimental conditions were
different (e.g., hive volume), these comparisons were also inappropriate
for comparing temperate and tropical honey bee populations. Were the
apparent differences in colony reproduction between the two honey bee
populations the result of differences in: 1) colony demography; 2)
environmental and climatic factors; 3) experimental design; 4) resource
utilization efficiencies; or 5) some combination of factors? Are there
intrinsic differences between the two honey bee populations with respect
to colony demographic parameters that allow for a more rapid colony
growth rate and result in a greater reproductive rate for the
Africanized honey bee population? Or, are the differences in
reproductive rates a result of climatic conditions and/or resource
availabilities and utilization in the tropics compared with temperate

137
regions? Finally, were the relatively high reproductive rates observed
for Africanized honey bees in these studies (Otis 1980, 1982a) simply an
artifact of experimental conditions, particularly with respect to brood-
nest crowding?
Brood-nest crowding is a primary stimulus for reproductive swarming
in honey bees (Baird and Seeley 1983; Simpson 1966, 1973; Simpson and
Riedel 1963). However, the experimental conditions affecting brood-nest
crowding for Africanized colonies in South America were significantly
different from the experimental conditions for European colonies in
North America: the nest cavity volume for Africanized colonies was 22
liters (Otis 1980; Winston 1979b) compared with 42 liters for European
colonies in North America (Winston 1980a). Despite these problems with
respect to making valid comparisons, the earlier studies (particularly
Winston 1979b) leave one with the impression that the apparent
differences in reproductive rates between the two populations were
primarily due to differences in demographic parameters and not to
differences in environmental and experimental conditions, resource
utilization, or some combination of factors.
What is the consequence of comparing reproductive rates of
Africanized honey bees in South America with those of European honey
bees in North America without considering differences in environmental
conditions? Certainly, environmental conditions in temperate regions
impose strict limits on the length of the reproductive (= swarming)
season for honey bees because of a reduced growing season when floral
resources (nectar and pollen) are available. The honey bee reproductive
season is significantly shorter than the growing season, because
colonies first have to grow to reproductive size before swarming can

138
occur. In addition, offspring (swarms) need a rather long period of
time to grow and to hoard necessary surplus honey, while there are still
floral resources available, in order to prepare for winter.
Most mortality of honey bee colonies in temperate areas occurs
primarily due to starvation during winter: 11% for first-year colonies
and 90% for established colonies (Seeley 1978, 1983). There is a high
energetic cost of maintaining proper brood nest or cluster temperature
during the cold winter. In addition, the high energetic cost of winter
survival in temperate regions may greatly reduce the survivorship of
small, secondary swarms or afterswarms, which would greatly reduce the
net reproductive rate of honey bees in temperate regions. In contrast,
periods of resource dearth in the tropics are not only shorter, but
require less stored honey (per unit time) to enable the colonies to
survive because of reduced energetic costs for maintaining brood nest
temperature.
The reproductive season in French Guiana, South America, was nine
months (Winston 1980b) compared with two to four months for North
America (references cited in Winston 1980b and Winston, Dropkin and
Taylor 1981). Is it a coincidence that the difference in the annual
reproductive rate of Africanized bees in South America compared with
European bees in North America, 16 vs. 3.0-3.6, respectively (Otis
1982a; Winston 1980a; Winston, Taylor and Otis 1983), is approximately
of the same order (factor of 4-5) as the difference in the length of the
reproductive season between South America and many areas of North
America?
Another factor affecting differences in reproductive rates between
honey bee populations in South America with those in North America is

139
the extent to which brood rearing ceases during resource dearths.
European honey bee populations in temperate areas have a distinct
seasonal decline in brood production and may stop brood rearing
altogether for a variable period during winter (Bodenheimer 1937;
Bodenheimer and Ben-Nerya 1937; Jeffree 1955; McLellan 1978; Nolan 1925,
1928). On the other hand, Winston reports that many Africanized
colonies in French Guiana
persist during the relative dearth season (March to June)
without the cessation of brood rearing characteristic for
temperate conditions, and are strong enough (i.e., have a
relatively high worker population and sufficient young
workers) to grow rapidly to swarming strength when resources
improve. (Winston 1980b, p. 164).
This difference between tropical and temperate conditions allows
tropical honey bee colonies to grow rapidly when resources become
available and thereby increase their potential reproductive rates
compared with temperate honey bee colonies.
More recently, investigations of both Africanized and European
honey bee populations under identical experimental conditions in
Venezuela have been undertaken. These studies have evaluated both
demographic parameters as well as resource utilization behaviors. These
investigations include the research presented in Chapters II-VII;
studies by Winston and Katz (1981, 1982); and the research by the U.S.
Department of Agriculture Bee Breeding and Stock Center Laboratory
(Collins, Rinderer, Harbo and Bolten 1982; Harbo, Bolten, Rinderer and
Collins 1981; Rinderer, Bolten, Collins and Harbo 1984; Rinderer,
Bolten, Harbo and Collins 1982; Rinderer, Collins, Bolten and Harbo
1981; Rinderer, Collins and Tucker 1985; Rinderer, Tucker and Collins
1982). Results of these investigations present quite a different

140
picture as to the factors leading to the success and resulting impact of
the Africanized honey bee in South America.
Factors Contributing to the Selective Advantage of
Africanized Honev Bees in South America
Colony Demography
Table 8-1 summarizes the factors affecting colony survival and
reproductive success for both Africanized and European honey bees under
tropical conditions in Venezuela. Studies comparing parameters of
colony demography for Africanized and European honey bees under
identical conditions in Venezuela have produced surprising results (see
Chapters IIVI). These studies were based on the assumption that the
life history of the Africanized honey bee population in South America
(as well as the parental population in Africa) was characterized by a
high reproductive rate. Demographic features that were expected to be
correlated with this high rate of colony reproduction, or short swarm to
swarm interval, were shorter worker bee development time, smaller worker
bee size, more rapid queen development and maturation, increased egg
laying and brood production, reduced brood mortality, and increased
adult worker bee longevity.
Colony demographic characteristics can be divided into two groups:
those affecting the rate of colony growth and those affecting the time
interval from swarming to the beginning of adult population increase. A
rapid colony growth rate is most important for a high colony
reproductive rate and is primarily a function of queen fecundity, adult
worker longevity, and brood mortality (Brian 1965; Moeller 1961; Wilson
1971).

141
Although Africanized queens have been reported to have greater egg
laying rates than European queens (Fletcher 1978; Michener 1972, 1975;
Ribbands 1953), under identical experimental conditions in Venezuela,
there was no significant difference in queen fecundity during the
initial colony growth period (Chapters V and VI). Also, European honey
bee workers live longer (Winston and Katz 1981), giving European honey
bee colonies a growth rate advantage with respect to this demographic
parameter.
Because of the relationship of worker longevity to colony growth
rates, initial colony growth rates may be affected by the age structure
of bees in a swarm. Colonies established from swarms with older bees
will have a more rapid decline in population, which will adversely
affect colony growth because egg laying rates are a function of the
number of bees in a colony (Moeller 1958). The age structure of
Africanized swarms has been evaluated (Winston and Otis 1978) but there
are no data for both Africanized and European swarms under similar
conditions.
Another parameter affecting colony growth rate is brood mortality,
but there are no data available that simultaneously compare Africanized
and European honey bees under identical conditions. Experimental and
environmental conditions are particularly important with respect to this
parameter. Rates of brood mortality can be as high as 50% and are
affected by season, resource availability and colony adult population
(Garofalo 1977; Merrill 1924; Woyke 1977). These high rates of brood
mortality and/or brood cannibalism may function to regulate protein
balance in honey bee colonies during protein (pollen) shortages (Weiss
1984). Therefore, earlier studies comparing brood mortalities in

142
Africanized and European colonies (Winston, Dropkin and Taylor 1981),
which were observed under very different conditions, need to be re
evaluated, and new studies should be undertaken.
Also affecting colony growth rates is the extent to which brood
rearing ceases during periods of resource dearth. As discussed earlier,
Winston (1980b) reports that Africanized bees do not cease brood rearing
to the extent observed for European bees and are therefore capable of
rapid colony growth when conditions improve. European honey bees, under
some tropical conditions, may have a sharp decline in brood rearing
during resource shortages (Otis and Taylor 1979). However, these
differences in brood rearing were not apparent when both Africanized and
European honey bees were managed under identical conditions in Venezuela
(Bolten, personal observation). Therefore, this behavior needs to be
analyzed with both honey bee populations under a variety of tropical
resource conditions to determine if there are differences, and whether
the differences are a function of foraging behavior (see below) and/or
intrinsic demographic parameters.
The two most important factors affecting the interval from swarming
to the beginning of adult population increase are worker development
time and queen maturation (Chapters II and IV). Worker development time
has previously been considered an important factor affecting the rate of
colony growth (Fletcher 1977a, 1978; Fletcher and Tribe 1977a; Tribe and
Fletcher 1977; Winston 1979b; Winston, Dropkin and Taylor 1981; Winston
and Katz 1982; Winston, Taylor and Otis 1983). As discussed in Chapter
II, this is incorrect and is probably a result of confusing models for
colony growth (increase in the number of bees in the colony) with models
for population growth (increase in the number of colonies). Population

143
growth models are designed for other species in which all individuals
are potential reproductives. For honey bees, individual (or worker bee)
development time is not equal to generation time. Worker bee
development time only affects the interval between^a given change in egg
laying rate and its resulting change in rate of adult emergence. The
difference in worker development time between Africanized and European
honey bees is only one day (Chapter II) and is trivial with respect to
other factors affecting reproductive rates.
Time from virgin queen emergence to initiation of oviposition also
only affects the interval from swarming until adult population increase
begins and not the rate of colony growth (Chapter IV). The results
presented in Chapter IV for maturation rates of Africanized and European
queens were unexpected. European honey bee queens began oviposition at
an earlier age post-emergence than did Africanized queens.
Reproductive Output
Reproductive output is mainly determined by the swarm to swarm
interval and the number of swarms produced per swarming cycle. The
swarm to swarm interval is a function of all the colony demographic
parameters discussed in the previous section and resource utilization
parameters discussed in the next section. Although there are no data on
the swarm to swarm intervals for Africanized and European honey bees
under identical conditions, results from investigations of demographic
parameters reported above suggest that if differences in swarm to swarm
intervals exist, they would not be a result of demographic differences.
Rather, if differences exist, they are hypothesized to be a result of
differences in resource utilization between the two populations (see
below).

144
Although Africanized and European honey bees have not been compared
under identical conditions, European colonies in North America (Kansas)
produced the same number of small, secondary swarms, or afterswarms, per
swarming cycle as did Africanized colonies in South America (French
Guiana) (Otis 1980; Winston 1980a; Winston, Dropkin and Taylor 1981).
These results are not directly comparable, but they do demonstrate that,
at least under certain conditions, European honey bees can produce as
many afterswarms as Africanized honey bees. Whether the number of
afterswarms for European bees would be similar to Africanized bees under
identical conditions needs to be analyzed. As discussed above,
survivorship of small afterswarms would be much lower in temperate
regions than in tropical regions, because of the energetic demands of
temperate winters on honey bee colonies.
Resource Utilization
The most important factors leading to the success of Africanized
honey bees in South America are associated with resource utilization:
foraging behavior, brood production efficiency, worker bee size and
absconding behavior. Although evaluated under different conditions, the
foraging range of the parental African population is similar to that of
European bees (Smith 1958b). However, under resource conditions typical
of tropical regions, the foraging behavior of Africanized honey bees is
significantly more successful than that of European honey bees
(Rinderer, Bolten, Collins and Harbo 1984; Rinderer, Collins and Tucker
1985), Their success is a result of more frequent solitary foraging and
reduced recruitment when resources are dispersed and limited, as is
characteristic of most tropical habitats (Rinderer, Bolten, Collins and
Harbo 1984; Rinderer, Collins and Tucker 1985). Using artificial

145
flowers, Nunez (1973, 1979a, 1982) analyzed foraging behaviors of
Africanized and European honey bees. Differences between the two
populations were observed that were appropriate to having evolved under
either temperate or tropical resource availability patterns. Hoarding
cage studies also demonstrated differences in response between
Africanized and European honey bees, suggesting differences in resource
utilization behavior (Rinderer, Bolten, Harbo and Collins 1982).
Increased numbers of bees in a colony are important for successful
foraging, colony defense and reproduction (see Wilson 1971). When
floral resources (nectar and pollen) are limited, a greater number of
individual bees can be produced from a given amount of food if brood
production is more efficient and/or if bees are smaller. There are no
data available that were collected under identical conditions that allow
comparison of brood production efficiency between Africanized and
European honey bees, as measured by the ratio of developing brood to
adult population for a range of different adult populations (Michener
1964; Moeller 1961). However, their smaller size may result in
increased brood production efficiency in Africanized bees, i.e., less
food is necessary to produce smaller bees (Chapter III; Fletcher and
Tribe 1977a; Tribe and Fletcher 1977). Therefore, with a limited food
supply, Africanized honey bees could increase their population at a
greater rate than European honey bees. With more successful foraging
behavior and smaller bee size, Africanized colonies can grow rapidly
under conditions where European colonies may not be able to survive.
Another important difference between Africanized and European honey
bees is the strategy used to survive during periods of food shortage.
Honey bee colonies may either hoard sufficient quantities of food

146
(primarily honey) to sustain them during periods of resource dearth or
the colonies can abscond (relocate or migrate) to another area where
conditions may be better. Hoarding large surpluses of honey is
characteristic of European honey bees in temperate regions. However,
hoarding behavior may be disadvantageous in the tropics because colonies
with large food surpluses may be more easily discovered by predators and
less easily protected. Because predation has been a major evolutionary
force for tropical honey bee populations (Seeley 1983; Seeley, Seeley
and Akratanakul 1982), resource-induced absconding may be a better
evolutionary alternative to hoarding.
Resource-induced absconding occurs when an entire colony abandons a
nest and is quite common in tropical species of Apis (A. florea, A.
dorsata, and A. cerana) and tropical populations of A. mel1ifera during
periods of resource dearth (Winston, Otis and Taylor 1979; Winston,
Taylor and Otis 1983; Woyke 1976). Africanized honey bees have a high
rate of resource-induced absconding in South America (Winston, Otis and
Taylor 1979). Resource-induced absconding behavior may not be
advantageous in temperate regions because colonies may not have enough
time once they settle in a new area to store sufficient food to
successfully overwinter (Butler 1974). European honey bees generally do
not abscond in either temperate or tropical regions (Butler 1974;
Fletcher 1978; Winston, Otis and Taylor 1979; Winston, Taylor and Otis
1983).
Whether resource-induced absconding behavior is a more successful
strategy in the tropics than is hoarding behavior requires further study
to determine the advantages and disadvantages of each strategy under
tropical conditions. Both may be viable strategies and may not be

147
mutually exclusive (Winston, Otis and Taylor 1979). Fletcher suggests
that resource-induced absconding may not always be appropriate,
considering
the distance that bees can fly in relation to the general
distribution of their food plants. The maximum flight range
is unlikely to exceed about 16 kilometres...and yet huge areas
of Africa inhabited by honey-bees consist of more or less
uniform grasslands and savannah. With certain exceptions,
therefore, such as movements up and down mountain slopes and
in and out of river valleys, there would appear to be little
advantage in absconding in such areas, for within their flight
range the bees would very often find only more of the same
type of country they had left. (Fletcher 1975, p. 13).
However, based on measurements of engorgement and estimates of metabolic
rates, the maximum flight range of absconding colonies of honey bees has
been calculated to be as great as 131 km (Otis, Winston and Taylor
1981). In addition, periodic foraging while in-transit could extend the
potential distance even further. Until comparative studies demonstrate
the advantages of either resource-induced absconding or hoarding,
resource-induced absconding behavior, which is common in tropical honey
bee populations, is concluded to be an advantageous behavior under some
tropical conditions.
Predation and Colony Defense
Ability to defend the nest from predators affects colony survival
and therefore reproduction. The colony defense behavior characteristic
of Africanized honey bees (Collins, Rinderer, Harbo and Bolten 1982;
Stort 1974, 1975a, 1975b, 1975c, 1976) may be more effective than that
of European honey bees. One aspect of colony defense behavior of
Africanized honey bees, their stinging behavior, is so extreme that
Africanized bees are a public health hazard for both humans and domestic
animals (Taylor 1986). This colony defense behavior is particularly
effective against vertebrate predators. Africanized honey bees in

148
Venezuela also reduce the size of their nest entrances to a greater
extent than European honey bees, which helps to protect against
invertebrate predators, particularly ants (Bolten, personal
observation).
Besides colony defense, another response to predation is to abscond
(relocate). Because predation by both vertebrates and invertebrates and
infestation by wax moths (Galleria me!lone!1 a and Achroia arise!la) on
honey bee colonies is extensive in tropical regions, disturbance-induced
absconding would be an advantageous behavior and is frequently observed
in tropical honey bees (Fletcher 1976; Seeley 1983; Seeley, Seeley and
Akratanakul 1982; Winston, Taylor and Otis 1983). Disturbance-induced
absconding was more frequently observed in Africanized honey bees than
in European honey bees under similar conditions in Venezuela-,
particularly with respect to attacks by ants (Bolten, personal
observation).
In addition to predation on the colony-level, Africanized worker
bees may have behaviors that are better adapted to avoiding predators
and parasites while foraging. The rapid, zig-zag flight of worker bees
in the African parental population may be more advantageous in avoiding
predators (invertebrate as well as vertebrate) compared with the slower,
less erratic flight of European honey bees (Fletcher 1977b). Also,
queen honey bees on mating flights are susceptible to predators.
Fletcher (1977b) suggests that queens from the African parental
population have shorter mating flights than European queens which may
reduce predation. The differences in flight patterns and behaviors of
Africanized and European honey bees need to be investigated under
similar conditions.

149
Nest Sites and Cavity Volume
Fletcher (1976) suggested another adaptive advantage that
Africanized bees have is their ability to utilize a greater variety of
nest sites. Fletcher may be confusing cause with effect when he
suggests that it is this ability that "has enabled them to establish
themselves in areas not previously inhabited by honey-bees at all"
(1976, p. 6). A more likely explanation for the success of Africanized
bees in those areas would be their ability to utilize the particular
nectar and pollen resources available. That is, without a more
efficient utilization of resources, Africanized honey bees would not be
able to exploit these other habitats irrespective of their ability to
utilize a greater variety of nest sites. As discussed above,
Africanized honey bees are more successful foragers than are European
honey bees under resource conditions typical of tropical habitats.
Nest cavity volume is another factor affecting reproduction in
honey bees. One of the stimuli for reproductive swarming is brood-nest
crowding (Baird and Seeley 1983; Simpson 1966, 1973; Simpson and Riedel
1963; Winston and Taylor 1980). Colonies inhabiting smaller cavities
become crowded more rapidly and have a higher tendency to swarm.
Colonies established in large cavities would be less crowded and have a
lower rate of swarming. In temperate regions, small cavities would be
selected against because there would be less volume available for
storing surplus honey to enable the colony to overwinter. Therefore,
Seeley proposed that nest-cavity volume may "regulate mature colony size
at an optimum between small colonies with low survivorship and large
colonies with low fertility" (Seeley 1977, p. 226). Jaycox and Parise
(1980, 1981) found that honey bees from northern Europe selected larger

150
nest cavities than did honey bees from southern Europe. Southern
European winters would be much less severe than those in northern
Europe, and therefore the need for larger nest cavities to store large
food surpluses is less important.
For tropical honey bee populations, there may be a selective
advantage for smaller nest cavity volumes, e.g., to facilitate
protection against infestation from wax moths (Galleria mellonella and
Achroia grisella) (Fletcher 1976). Africanized honey bees utilize a
wider variety of nest sites than do European honey bees, including
smaller cavity volumes (Fletcher 1976). The negative factors associated
with smaller nest cavities may be absent in the tropics because there is
less need to store large surpluses to survive periods of resource
dearthperiods are generally shorter and less costly with respect to
energetic demands for maintaining proper brood nest temperature. In
addition, honey bees that evolved in the tropics commonly abscond during
periods of resource dearth as opposed to hoarding surplus food.
Although nest cavity choice for Africanized and European honey bees
has not been studied under identical conditions, nest cavities selected
by Africanized honey bees in Venezuela were not smaller than cavities
selected by European honey bees in temperate regions (Rinderer, Collins,
Bolten and Harbo 1981; Rinderer, Tucker and Collins 1982). Because of
the importance of nest cavity volume to reproductive rates, nest cavity
volume for both populations needs to be investigated under identical
conditions.
Density-Dependent Factors Regulating Queen Rearing
Other parameters that might account for differences in reproductive
rates between Africanized and European honey bees may be certain

151
density-dependent factors that are responsible for regulating queen
rearing in colonies preparing to reproduce. In European honey bees,
initiation of queen rearing prior to reproduction is not a result of a
decrease in queen pheromone production (Seeley and Fell 1981). Two
other possibilities are suggested by Seeley and Fell (1981). First,
there may be failure to adequately disperse queen pheromone in crowded
colonies prior to swarming. And, second, worker bee response to queen
pheromone may change prior to swarming.
Threshold levels for queen pheromone that inhibit queen rearing in
worker bees may be different for Africanized and European honey bees.
Also, dispersal of queen pheromone by "messenger bees (Seeley 1979) may
be different for Africanized bees compared with European bees. Baird
and Seeley (1983) developed an equilibrium model to explain the
regulation of queen rearing in colonies preparing to reproduce. Their
model postulated that "there is a balance between nurse bees becoming
inhibited from queen rearing and nurses losing their inhibition, and
that whether a colony does or does not rear queens reflects the
equilibrium percentage of inhibited nurses" (Baird and Seeley 1983, p.
221). Therefore, differences between Africanized and European honey
bees with respect to density-dependent factors regulating queen rearing
may result in differences in reproductive rates by affecting: 1) adult
population size when colonies reproduce; 2) prime swarm size; and 3)
number of afterswarms. Some of these density-dependent factors have
been compared for Africanized bees in South America with European bees
in North America under different environmental and experimental
conditions (Winston, Dropkin and Taylor 1981). Unfortunately, there are
no data collected under identical conditions that allow valid

152
comparisons to be made between Africanized and European honey bees that
enable any density-dependent factors responsible for the reproductive
rates and success of Africanized honey bees in South America to be
identified.

Conclusion
In tropical regions, the success of Africanized honey bees compared
with European honey bees is not a function of any intrinsic differences
in colony demography. Rather, it must be concluded that the success of
Africanized honey bees is due primarily to their ability to efficiently
utilize tropical resources, enabling them to survive and reproduce under
conditions where European honey bees are frequently not able to survive.
If European honey bee colonies are not able to survive and/or grow under
some of the tropical resource conditions of South America, they
obviously cannot reproduce. It is precisely because the European honey
bees were not successful foragers (= honey producers) in most tropical
regions of Brazil that African honey bees were imported into Brazil
(Goncalves 1974, 1975, 1982; Woyke 1969).
The efficient utilization of tropical resources by Africanized
honey bees is a result of a set of adaptive behaviors involving solitary
foraging, reduced recruitment, increased brood production efficiency
because of smaller worker bee size, and both resource-induced and
disturbance-induced absconding. These characteristics, combined with an
effective colony defense behavior, give Africanized honey bees a
selective advantage that results in increased survivorship, increased
colony growth rates and ultimately increased reproduction, which is
responsible for their rapid dispersal and high population densities.

153
The Africanized honey bees studied in Venezuela are only a small
sample of the total Africanized honey bee population in South and
Central America and represent only a fraction of the variation within
the population, particularly if we consider that Africanized honey bees
are a result of hybridization. Nevertheless, the results presented in
the foregoing chapters demonstrate that at least some portion of the
Africanized honey bee population is similar to the European honey bee
population with respect to the demographic parameters analyzed.
Potential Impact of Africanized Honev Bees in North America
The selective advantage of Africanized honey bees in South America
will be lost as they disperse north into temperate regions. European
honey bees will have the selective advantage in temperate regions
because of their particular behavioral repertoire which is better
adapted to temperate conditions. However, because the populations can
interbreed successfully, negative characteristics of the Africanized
population, e.g., their stinging behavior, may become genetically
introgressed into the European population of North America and therefore
widespread wherever honey bees can survive (Chapter VII). A more
optimistic scenario is that the large population of European honey bees
in Mexico will slow the spread of African genes because of competition
for available resources as well as through hybridization. Therefore,
through selection, hybridization, and competition, the impact of
Africanized honey bees may be minimized in North America (Chapter VII).

154
TABLE 8-1. Factors affecting colony survival and reproductive success
for Africanized and European honey bees in Venezuela.
POPULATION
FACTOR WITH ADVANTAGE REFERENCES
COLONY DEMOGRAPHY
Growth Rate
Egg Laying Rate No Difference Chapter VI; Chapter V
Worker Longevity European Winston & Katz 1981
Swarm Age Structure No Data
Brood Mortality No Data
Brood Production No Data
during Resource Dearth
Interval3
Worker Development Time Africanized
Queen Maturation European
REPRODUCTIVE OUTPUT
Number of Afterswarms No Data
per Swarming Cycle
RESOURCE UTILIZATION
Foraging Behavior Africanized
Brood Production
Efficiency
Bee Size
Resource-Induced
Absconding
PREDATION
Colony Defense
No Data
Africanized
Africanized
Africanized
Disturbance-Induced Africanized
Absconding
Flight Behavior No Data
NEST CAVITY VOLUME No Data
DENSITY-DEPENDENT FACTORS No Data
Chapter II
Chapter IV
Rinderer, Bolten, Collins
& Harbo 1984; Rinderer,
Bolten, Harbo & Collins
1982; Rinderer, Collins &
Tucker 1985; Nunez 1979,
1982; Winston & Katz 1982
Chapter III
Winston, Otis & Taylor 1979
Winston, Taylor & Otis 1983
Collins, Rinderer, Harbo &
Bolten 1982
Bolten, pers. observation;
Winston, Taylor & Otis 1983
aInterval from swarming to beginning of population increase.

APPENDIX A
WORKER BEE DEVELOPMENT TIMES AND MORTALITY
DURING DEVELOPMENT

TABLE A-l
. Comparison of worker bee development time (in days) for
Africanized and European honey bees: median, (range), mean
+ SD, (n = sample size). All development measured in
European comb cell size with European nurse bees.
EGG
GENOTYPES UNSEALED BRtfOD SEALED BROOD
TOTAL
DEVELOPMENT3
AFRICANIZED
A53
(n = 25)
5.0
(4-5)
4.8 + 0.4
11.0
(11-12)
11.4 + 0.5
19.0
(19-20)
19.2 + 0.4
A26
(n = 9)
5.0
(5)
5.0 + 0
11.0
(11-12)
11.3 + 0.5
19.0
(19-20)
19.3 + 0.5
A25
(n = 19)
5.0
(4-5)
4.6 + 0.5
11.0
(11-12)
11.4 + 0.5
19.0
(19)
19.0 + 0
COMBINED
(n = 53)
5.0
(4-5)
4.8 + 0.4
11.0
(11-12)
11.4 + 0.5
19.0
(19-20)
19.2 + 0.4
EUROPEAN
W18
(n = 28)
5.0
(4-5)
4.8 + 0.4
12.0
(11-13)
12.0 + 0.3
20.0
(19-20)
19.8 + 0.4
HI
(n = 19)
5.0
(5-6)
5.4 + 0.5
12.0
(11-13)
12.1 + 0.4
20.0
(20-21)
20.5 + 0.5
Y(A5)
(n = 26)
5.0
(4-5)
4.8 + 0.4
12.0
(11-12)
12.0 + 0.2
20.0
(19-20)
19.8 + 0.4
COMBINED
(n = 73)
5.0
(4-6)
5.0 + 0.5
12.0
(11-13)
12.0 + 0.3
20.0
(19-21)
20.0 + 0.5
ANALYSES5
NS
P<0.001
P<0.001
3Total development = time from oviposit ion to adult emergence.
bKolmogorov-Smirnov one-tailed test, chi-square distribution, df = 2,
alpha = 0.05 (Siegel 1956). Combined samples used for analyses.
156

157
TABLE A-2. Mortality during different developmental stages. Mortality
was measured in European comb cell size with European nurse
bees.
El3 E2b L1C L2d Sb8 Nf
AFRICANIZED EGG GENOTYPES
30
28
29
30
30
30
fMortality during first 24 hours in test colony (acceptance).
^Mortality between 24-72 hours (before hatching).
'-Mortality between 72-96 hours (at time of hatching).
Mortality during older larval stages, before sealing.
^Mortality during the pupal stage.
fN = total eggs monitored.
9Not distinguished between and L2.
A53
A26
A25
EUROPEAN EGG GENOTYPES
W18
HI
Y(A5)
0
0
1
0
0
0
0 5
0
0 9
0 0
0 3
0 4
0 0
199 o
0 0
2 0
8 0
0 0

APPENDIX B
HONEY BEE SIZE, COMB CELL SIZE AND SIZE VARIATION

TABLE B-l. Coefficients of variation (CV) of worker bees in honey
bee and bumble bee colonies calculated from data
presented in the references.
CV
REFERENCES
Honey bees (Ap is mel1 ifera)
Weights
Adult (fresh)
0.4-0.6
Abdellatif 1965
4.0-4.53
Bolten (Table B-2)
5.4-7.0b
Bolten (Table B-2)
10.7-11.2
Kerr and Hebling 1964
Adult (dry)
4.8-8.1
Grout 1937
4.1-4.53
Bolten (Table B-2)
4.1-6.7
Bolten (Table B-2)
Linear Measurements
Length forewing
1.5-1.6
Grout 1937
Width forewing
2.2-2.5
Grout 1937
Length proboscis
1.6-1.9
Grout 1937
Bumble bees (Bombus)
Weights
Adult (fresh) 31.0-36.7 Brian 1952
(Bombus agrorum)
Linear Measurements0*
Length radial cell 7.4-13.8 Medler 1965
(Bombms f.erv.idiig)
^European genotypes.
Africanized genotypes in South America.
cCV may be high as a result of variable engorgement during 4 hour delay
from emergence to weighing.
Different linear measurements for Bombus are significantly correlated
(P<0.01, Medler 1962).
159

TABLE B-2
160
TABLE B-2.
Africanized
mean + SD,
and European adult honey bee weights (mg):
coefficient of variation (CV), (sample size).
GENOTYPE
COMB CELL
TYPE3
FRESHLY
EMERGED
DRIEDb
CORRELATIONS'
AFRICANIZED
A26
EUR
94.9 + 5.1
CV = 5.4
(28)
13.5 + 0.6
CV = 4.4
(28)
#**
A57
EUR
88.6 + 6.2
CV = 7.0
(30)
11.9 + 0.8
CV = 6.7
(30)
***
B39
EUR
87.4 + 4.7
CV = 5.4
(16)
12.3 + 0.5
CV = 4.1
(16)
NS
A60
AFR
95.2 +5.1
CV = 5.4
(30)
13.2 + 0.6
CV = 4.5
(30)
***
EUROPEAN
WE2
EUR
107.1 + 4.8
CV = 4.5
(29)
14.6 + 0.6
CV = 4.1
(29)
**
Y (K)
EUR
116.1 + 4.7
CV = 4.0
(27)
15.5 + 0.7
CV = 4.5
(27)
***
aEUR = European comb cell diameter = 5.4 mm.
AFR = Africanized comb cell diameter = 4.8 mm.
Dried at 50C for 48 hrs.
Pearson's correlation coefficient alpha = 0.05;
** = PcO.Ol; *** = P<0.001.

161
TABLE B-3. Comparison of Africanized and European comb cell diameter
and comb cell volume: mean + SD, range, CV, (sample size).
DIAMETER3 VOLUME5
(mm) (ml x 10-3) CORRELATIONS0
AFRICANIZED COMB CELLS 4.8 +0.1
4.6 4.9
CV =2.1
(50)
184.6 + 15.8
160 215
CV = 8.6
(50)
EUROPEAN COMB CELLS
5.4 + 0.05
5.4 5.5
CV = 1.0
(30)
264.3 + 23.5
225 300
CV = 8.9
(30)
NS
NSd
ANALYSES
P<0.001 P<0.001
determined by measuring 10 horizontal, consecutive cells; cell-wall
thickness not considered.
5Cell volume determined by filling cells with water with a pipette.
cSpearman's rank correlation coefficient, alpha = 0.05.
dNegative correlation, P<0.01.
et-test, one-tailed.

TABLE B-4. Comparison of Africanized and European comb cell diameter
and comb cell depth: mean + SD, range (sample size).
DIAMETER3
DEPTH
(mm)
(mm)
CORRELATIONS
AFRICANIZED COMB CELLS
4.8 + 0.1
11.8 + 0.2
NS
4.7 4.9
11.4 12.3
(17)
(17)
EUROPEAN COMB CELLS
5.4 + 0.05
12.2 + 0.4
NS
5.3 5.4
11.5 12.8
(26)
(26)
ANALYSES0
P<0.001 PcO.OOl
determined by measuring 10 horizontal, consecutive cells; cell-wall
thickness not considered.
bSpearman,s rank correlation coefficient, alpha = 0.05.
t-test, one-tailed.

163
TABLE B-5. Changes in European worker bee pupal weight (mg) with
changes in pupal age: mean + SD, (sample size).
Fresh weights were measured in Gainesville, Florida.
AGE (DAYS POST-OVIPOSITION)
11.5 12.5 13.5 14.5 15.5 16.5 17.5
145.8
145.5
141.9
140.9
141.1
139.1
138.0
+ 4.8
+ 3.8
+ 3.8
+ 4.2
+ 4.6
+ 4.0
+ 4.8
(10)
(45)
(31)
(38)
(31)
(32)
(34)
A
B
C
D
E
F
G
ANALYSES
CDEF
NS
A x B
NS
B x C
NS
C x D
NS
D x E
NS
E x F
NS
F x G
NS
^One-way analysis of variance, alpha = 0.05.
bt-test, two-tailed, alpha = 0.05.

APPENDIX C
CHANGES IN QUEEN PUPAL WEIGHT WITH AGE

TABLE C-l. Changes in European queen pupal weights (mg) with
changes in age: mean + SD, (sample size). Queen pupal
weights were measured in Baton Rouge, Louisiana.
DAYS POST-OVIPOSITION
9
10
11
12
13
14
311.6
+ 12.4
(10)
291.6
+ 8.3
(10)
294.6
+ 9.7
(10)
293.4
+ 8.4
(10)
287.4
+ 11.2
(10)
274.1
+ 16.9
(5)
A
B
C
D
E
F
ANALYSES3
ABCDEF
BCDE
P<0.001
NS
a0ne-way analysis of variance, alpha = 0.05.
165

APPENDIX D
ACCURACY OF TECHNIQUE USED TO ESTIMATE NUMBERS OF BEES IN A COLONY

TABLE D-l. Accuracy of technique used to estimate number of bees in
colony.
SAMPLE NO.
COUNTED
ESTIMATED3
DIFFERENCE
% DIFFERENCE
1
2284
2324
40
1.8
2
2906
2918
12
0.4
3
3085
3080
5
0.2
4
3079
3124
45
1.5
5
2657
2717
60
2.2
6
2046
2102
56
2.7
aThe number of
adult bees was estimated by determining the
mean
individual bee weight from three, 150-200
bee samples.
The total
weight of bees in each sample was then divided by the mear
i individual
bee weight to get an estimate of the total
number of bees
in each
sample.
167

LITERATURE CITED
Abdellatif, M.A. 1965. Comb cell size and Its effect on the body
weight of the worker bee Apis mel11fera L. Am. Bee J. 105:86-87.
Adams, J. E.D. Rothman, W.E. Kerr and Z.L. Paulino. 1977. Estimation
of the number of sex alleles and queen matings from diploid male
frequencies in a population of Apis mel1ifera. Genetics 86:
583-596.
Alies, P. 1961. Role of queen and worker bees 1n the hereditary
transmission of certain characteristics. Apic. Absts. 12:167.
Alpatov, W.W. 1929. Biometrical studies on variation and races of the
honey bee. Quart. Rev. Biol. 4:1-58.
Alpatov, W.W. 1933. South African bees biometrically investigated.
Bee World 14:62-64.
Anderson, R.H., B. Buys and M.F. Johannsmeier. No date. Beekeeping in
South Africa. Technical Services Bulletin 394. South African
Dept, of Agriculture, Pretoria.
Baird, D.H., and T.D. Seeley. 1983. An equilibrium theory of queen
production in honeybee colonies preparing to swarm. Behav. Ecol.
Sociobiol. 13:221-228.
Baudoux, U. 1933. The influence of cell size. Bee World 14:37-41.
Beetsma, J. 1979. The process of queen-worker differentiation in the
honeybee. Bee World 60:24-39.
Boch, R. 1957. Rassenmabige Unterschiede bei den Tanzen der
Honigbiene. Z. Vergl. Physiol. 40:289-320.
Boch, R., and C.A. Jamieson. 1960. Relation of body weight to
fecundity in queen honeybees. Can. Ent. 92:700-701.
Boch R. D.A. Shearer and J.C. Young. 1975. Honeybee pheromones:
Field tests of natural and artificial queen substance. J. Chem.
Ecol. 1:133-148.
Bodenheimer, F.S. 1937. Studies in animal populations, II. Seasonal
population trends of the honey-bee. Quart. Rev. Biol. 12:406-425.
168

169
Bodenheimer, F.S., and A. Ben-Nerya. 1937. One-year studies on the
biology of the honey-bee in Palestine. Ann. Appl. Biol. 24:
385-403.
Bolten, A.B., and J.R. Harbo. 1982. Numbers of spermatozoa in the
spermatheca of the queen honeybee after multiple inseminations with
small volumes of semen. J. Apic. Res. 21:7-10.
Brian A.D. 1952. Division of labour and foraging in Bombus agrorum
Fabricius. J. Anim. Ecol. 21:223-240.
Brian, M.V. 1965. Social insect populations. Academic Press, London.
Brother Adam. 1966. In search of the best strains of bees.
Ehrenwirth, Munich.
Buchmann, S.L. 1982. Africanized bees confirmed in Panama. Am. Bee J.
122:322.
Buchner, R. 1955. Effect on the size of workers of restricted space
and nutrition during larval development. Ap1c. Absts. 6:15.
Butler, C.G. 1971. The mating behaviour of the honeybee (Apis
mellifera L.). J. Ent. (A) 46:1-11.
Butler, C.G. 1974. The world of the honeybee. Collins, London.
Butler, C.G., D.H. Calam and R.K. Callow. 1967. Attraction of Apis
mellifera drones by the odours of the queens of two other species
of honeybees. Nature 213:423-424.
Cantwell, G.E. 1974. The African (Brazilian) bee problem. Am. Bee J.
114:368-372.
Carlson, D.A., and A.B. Bolten. 1984. Identification of Africanized
and European honey bees, using extracted hydrocarbons. Bull. Ent.
Soc. Am. 30:32-35.
Collins, A.M., T.E. Rinderer, J.R. Harbo and A.B. Bolten. 1982. Colony
defense by Africanized and European honey bees. Science 218:72-74.
Crewe, R.M., and H. Hastings. 1976. Production of pheromones by
workers of Apis mellifera adansoni1. J. Apic. Res. 15:149-154.
Daly, H.V., and S.S. Balling. 1978. Identification of Africanized
honeybees in the Western Hemisphere by discriminant analysis. J.
Kans. Ent. Soc. 51:857-869.
Daly, H.V., K. Hoelmer, P. Norman and T. Allen. 1982. Computer-
assisted measurement and identification of honey bees (Hymenoptera:
Apidae). Ann. Ent. Soc. Am. 75:591-594.
Darwin, C. 1958. The origin of species. The New American Library,
Inc. New York. (Original work published 1859).

170
Dietz, A. 1978. An anatomical character suitable for separating drone
honey bees of Apis mellifera 1 iqustica from Apis me11 ifera
adansoni1. Pages 102-106 in Apicultura em clima quente.
Apimondia, Florianopolis, Brazil.
Dietz, A., R. Krell and F.A. Eischen. 1985. Preliminary investigation
on the distribution of Africanized honey bees in Argentina.
Apidologie 16:99-108.
DuPraw, E.J. 1965. Non-Linnean taxonomy and the systematics of honey
bees. Syst. Zool. 14:1-24.
Eckert, J.E. 1934. Studies on the number of ovarioles in queen
honeybees in relation to body size. J. Econ. Ent. 27:629-635.
Ewel, J.J., and A. Madriz. 1968. Zonas de vida de Venezuela.
Ministerio de Agricultura y Cria, Caracas.
Fell, R.D., J.T. Ambrose, D.M. Burgett, D. De Jong, R.A. Morse and T.D.
Seeley. 1977. The seasonal cycle of swarming in honeybees. J.
Apic. Res. 16:170-173.
Fletcher, D.J.C. 1975. New perspectives in the causes of absconding in
the African bee (Apis mel1 ifera adansonii L.), Part I. S. Afr. Bee
J. 47:11-14.
Fletcher, D.J.C. 1976. New perspectives in the causes of absconding in
the African bee (Apis mellifera adansoni i L.), Part II. S. Afr.
Bee J. 48:6-9.
Fletcher, D.J.C. 1977a. A preliminary analysis of rapid colony
development in Apis mellifera adansonii. Pages 144-145 in Proc.
Eighth Int. Congress, Int. Union for Study of Social Insects.
Wageningen, Netherlands.
Fletcher, D.J.C. 1977b. Evaluation of introductions of European honey
bees into southern and eastern Africa. Pages 146-147 in Proc.
Eighth Int. Congress, Int. Union for Study of Social Insects.
Wageningen, Netherlands.
Fletcher, D.J.C. 1978. The African bee, Apis mellifera adansonii, in
Africa. Ann. Rev. Ent. 23:151-171.
Fletcher, D.J.C., and G.D. Tribe. 1977a. Swarming potential of the
African bee, Apis mellifera adansonii L. Pages 25-34 in D.J.C.
Fletcher (ed.) African bees: Taxonomy, biology and economic use.
Apimondia, Pretoria.
Fletcher, D.J.C., and G.D. Tribe. 1977b. Natural emergency queen
rearing by the African bee gl. adansonii and its relevance for
successful queen production by beekeepers, II. Pages 161-168 in
D.J.C. Fletcher (ed.), African bees: Taxonomy, biology and
economic use. Apimondia, Pretoria.

171
Fletcher D.J.C., and G.D. Tribe. 1977c. Natural emergency queen
rearing by the African bee A. ul. adansonii and its relevance for
successful queen production by beekeepers I. Pages 132-140 in
D.J.C. Fletcher (ed.) African bees: Taxonomy biology and
economic use. Apimondia Pretoria.
Free, J.B. 1965. The allocation of duties among worker honeybees.
Symp. Zool. Soc. (London) 14:39-59.
Fyg, W. 1959. Normal and abnormal development in the honeybee. Bee
World 40:57-66, 85-96.
Garofalo, C.A. 1977, Brood viability in normal colonies of Apis
mellifera. J. Apic. Res. 16:3-13.
Gary, N.E. 1975. Activities and behavior of honey bees. Pages 185-264
in Dadant and Sons (eds.), The hive and the honey bee. Dadant and
Sons, Inc., Hamilton, Illinois.
Glushkov, N.M. 1958. Experiments on combs with enlarged cells. Apic.
Absts. 9:102.
Goncalves, L.S. 1974. Comments on the aggressiveness of the
Africanized bees in Brazil. Am. Bee J. 114:448-450.
Goncalves, L.S. 1975. Do the Africanized bees of Brazil only sting?
Am. Bee J. 115:8-10.
Goncalves, L.S. 1982. The economic impact of the Africanized honey bee
in South America. Pages 134-137 in M.D. Breed, C.D. Michener and
H.E. Evans (eds.), The biology of social insects. Westview Press,
Boulder, Colorado.
Gould, J.L. 1982. Why do honey bees have dialects? Behav. Ecol.
Sociobiol. 10:53-56.
Grout, R.A. 1937. The influence of size of brood cell upon the size
and variability of the honeybee. Research Bulletin 218,
Agricultural Experiment Station, Iowa State College of Agriculture
and Mechanic Arts, Ames.
Hall, H.G. In press. DNA differences found between Africanized and
European honeybees. Proc. Natl. Acad. Sci., USA.
Harbo, J.R. 1979. Storage of honeybee spermatozoa at -196C. J. Apic.
Res. 18:57-63.
Harbo J.R., and A.B. Bolten. 1981. Development times of male and
female eggs of the honey bee. Ann. Ent. Soc. Am. 74:504-506.
Harbo, J.R., A.B. Bolten, T.E. Rinderer and A.M. Collins. 1981.
Development periods for eggs of Africanized and European honeybees.
J. Apic. Res. 20:156-159.

Harbo, J.R., and T.I. Szabo. 1984. A comparison of instrumentany
inseminated and naturally mated queens. J. Apic. Res. 23:31-36.
172
Harp, E.R. 1973. A specialized system for multiple rearing of quality
honeybee queens. Am. Bee J. 113:256-258, 261.
Heinrich, B. 1979a. Bumblebee economics. Harvard Univ. Press,
Cambridge, Mass.
Heinrich, B. 1979b. Thermoregulation of African and European honeybees
during foraging, attack, and hive exits and returns. J. Exp. Biol.
80:217-229.
Holdridge, L.R. 1964. Life zone ecology. Tropical Science Center, San
Jose, Costa Rica.
Hoopingarner, R., and C.L. Farrar. 1959. Genetic control of size of
queen honey bees. J. Econ. Ent. 52:547-548.
Jay, S.C. 1963. The development of honeybees in their cells. J. Apic.
Res. 2:117-134.
Jaycox, E.R., and S.G. Parise. 1980. Homesite selection by Italian
honey bee swarms, Apis mellifera 1iqustica (Hymenoptera: Apidae).
J. Kans. Ent. Soc. 53:171-178.
Jaycox, E.R., and S.G. Parise. 1981. Homesite selection by swarms of
black-bodied honey bees, Apis mellifera caucasica and A. m. carnica
(Hymenoptera: Apidae). J. Kans. Ent. Soc. 54:697-703.
Jeffree, E.P. 1955. Observations on the decline and growth of honey
bee colonies. J. Econ. Ent. 48:723-726.
Johansson, T.S.K., and M.P. Johansson. 1973. Methods for rearing
queens. Bee World 54:149-175.
Kerr, W.E. 1967. The history of the introduction of African bees to
Brazil. S. Afr. Bee J. 39:3-5.
Kerr, W.E., and D. Bueno. 1970. Natural crossing between Apis
roe.11 if era adansonii and Apis mell if era 1 igus.lica. Evolution
24:145-155.
Kerr, W.E., L.S. Goncalves, L.F. Blotta and H.B. Made!. 1972.
Biologa comprada entre as abelhas italianas (Apis mel1 ifera
1iqustica) Africana (Apis mellifera adansonii) e suas hibridas.
Pages 151-185 in Anais do Io Congresso Brasileiro de Apicultura.
Congresso Brasileiro de Apicultura, Florianopolis, Brazil.
Kerr, W.E., and N.J. Hebling. 1964. Influence of the weight of worker
bees on division of labor. Evolution 18:267-270.
Koeniger, N., and H.N.P. Wljayagunasekera. 1976. Time of drone flight
in the three Asiatic honeybee species. J. Apic. Res. 15:67-71.

173
Krell, R., A. Dietz and F.A. Eischen. 1985. A preliminary study on
winter survival of Africanized and European honey bees in Cordoba*
Argentina. Apidologie 16:109-118.
Kulzh inskaya, K.P. 1956. The role of the food factor in the growth of
bees. Apic. Absts. 7:177.
Laidlaw, H.H., Jr. 1979. Contemporary queen rearing. Dadant and Sons,
Inc., Hamilton, Illinois.
Laidlaw, H.H., Jr., and J.E. Eckert. 1962. Queen rearing. Univ. of
California Press, Berkeley.
Lindauer, M. 1953. Division of labour in the honeybee colony. Bee
World 34:63-73, 85-90.
Mackensen, 0. 1964. Relation of semen volume to success in artificial
inseminations of queen honey bees. J. Econ. Ent. 57:581-583.
Mackensen, 0., and W.C. Roberts. 1948. A manual for the artificial
insemination of queen bees. USDA-ARA Bur. Ent. and Plant Quar.
ET-250.
Mackensen, 0., and K. Tucker. 1970. Instrumental insemination of queen
bees. Agrie. Hdbk. 390. USDA, Washington, D.C.
McDowell, R. 1984. The Africanized bee in the United States: What
will happen to the U.S. beekeeping industry? Agrie. Econ. Rpt.
519. USDA, Washington, D.C.
McGregor, S.E. 1938. Environmental factors and size variations in
honeybee appendages. J. Econ. Ent. 31:570-573.
McLellan, A.R. 1978. Growth and decline of honeybee colonies and
inter-relationships of adult bees, brood, honey and pollen. J.
Appl. Ecol. 15:155-161.
Medler, J.T. 1962. Morphometric studies in bumble bees. Ann. Ent.
Soc. Am. 55:212-218.
Medler, J.T. 1965. Variation in size in the worker caste of Bombus
fervidus (Fab.). Pages 388-389 in P. Freeman (ed.), Proc. Xllth
International Congress of Entomology. Royal Entomological Society
of London, London.
Melampy, R.M., and E.R. Willis. 1939. Respiratory metabolism during
larval and pupal development of the female honeybee (Apis mellifera
L.). Physiol. Zool. 13:283-293.
Mel'nichenko, A.N. 1962. Experiments in directed alteration of the
characteristics of female and male honeybees in rearing in the hive
of a different variety. Biol. Abs. 39:661.

174
Merrill, J.H. 1924. Observations on brood-rearing. Am. Bee J. 64:
337-338.
Michener, C.D. 1964. Reproductive efficiency in relation to colony
size in hymenopterous societies. Insectes Sociaux 11:317-341.
Michener, C.D. 1972. Final report of the Committee on the African
Honey Bee. Nat. Res. Counc., Nat. Acad. Sci., Washington, D.C.
Michener, C.D. 1974. The social behavior of the bees. Belknap Press,
Cambridge, Mass.
Michener, C.D. 1975. The Brazilian bee problem. Ann. Rev. Ent.
20:399-416.
Michener, C.D. 1979. Biogeography of the bees. Ann. Missouri Bot.
Gard. 66:277-347.
Milum, V.G. 1930. Variations in time of development of the honeybee.
J. Econ. Ent. 23:441-447.
Moeller, F.E. 1958. Relation between egg-laying capacity of queen bee
and populations and honey production of their colonies. Am. Bee J.
98:401-402.
Moeller, F.E. 1961. The relationship between colony populations and
honey production as affected by honey bee stock lines. Production
Research Report 55. USDA, Washington, D.C.
Morse, R.A. 1984. The mating behavior of African queens. Glean. Bee
Cult. 112:125.
Morse, R.A., D.M. Burgett, J.T. Ambrose, W.E. Conner and R.D. Fell.
1973. Early introductions of African bees into Europe and the New
World. Bee World 54:57-60.
Nelson, J.A., and A.P. Sturtevant. 1924. The rate of growth of the
honeybee larvae. Department Bulletin 1222. USDA, Washington, D.C.
Nolan, W.J. 1925. The brood-rearing cycle of the honeybee. Department
Bulletin 1349. USDA, Washinton, D.C.
Nolan, W.J. 1928. Seasonal brood-rearing activity of the Cyprian
honeybee. J. Econ. Ent. 21:392-403.
Nunamaker, R.A., and W.T. Wilson. 1981. Comparison of MDH allozyme
patterns 1n the African honey bee (Apis mel1 ifera adanson i i L.) and
the Africanized populations of Brazil. J. Kans. Ent. Soc. 54:
704-710.
Nunez, J.A. 1973. Quantitative investigation of the behaviour of Apis
mellifera 1 iqustica Spinola and Apis mellifera adansonii Latreille:
Energy factors, forager recruitment and foraging activity. Apiacta
8:151-154.

175
Nunez J.A 1979a. Times spent on various components of foraging
activity: Comparison between European and Africanized honeybees in
Brazil. J. Apic. Res. 18:110-115.
Nunez J.A. 1979b. Comparative study of thermoregulation between
European and Africanized Apis me!1 ifera in Brazil. J. Apic. Res.
18:116-121.
Nunez J.A. 1982. Comparacin del comportamiento recolector de abejas
Africanizadas y abejas Europeas. Pages 221-231 in P. Jaisson
(ed.) Social insects in the tropics. Universite Paris-Nord
Paris.
Oertel E. 1940. Mating flights of queen bees. Glean. Bee Cult.
68:292-293.
Otis G.W. 1980. The swarming biology and population dynamics of the
Africanized honeybee. Ph.D. Dissertation, Univ. of Kansas,
Lawrence.
Otis, G.W. 1982a. Population biology of the Africanized honey bee.
Pages 209-219 in P. Jaisson (ed.), Social insects in the tropics.
Universite Paris-Nord, Paris.
Otis, G.W. 1982b. Weights of worker honeybees in swarms. J. Apic.
Res. 21:88-92.
Otis, G.W., and O.R. Taylor. 1979. Beekeeping in the Guianas. Pages
145-154 in Beekeeping in rural development: Unexploited beekeeping
potential in the tropics. Commonwealth Secretariat, London.
Otis, G.W., M.L. Winston and O.R. Taylor. 1981. Engorgement and
dispersal of Africanized honeybee swarms. J. Apic. Res. 20:3-12.
Page, R.E. Jr. and E.H. Erickson, Jr. 1984. Selective rearing of
queens by worker honey bees: Kin or nestmate recognition. Ann.
Ent. Soc. Am. 77:578-580.
Page, R.E. Jr. and E.H. Erickson, Jr. 1985. Identification and
certification of Africanized honey bees. Ann. Ent. Soc. Am.
78:149-158.
Page, R.E. Jr. and R.A. Metcalf. 1982. Multiple mating, sperm
utilization and social evolution. Am. Nat. 119:263-281.
Peer, D.F. 1956. Multiple mating of queen honey bees. J. Econ. Ent.
49:741-743.
Pellett F.C. 1938. History of American beekeeping. Collegiate Press,
Inc., Ames, Iowa.
Reid, M. 1975. Storage of queen honeybees. Bee World 56:21-31.

176
Ribbands C.R. 1953. The behaviour and social life of honeybees. Bee
Research Assoc., Ltd. England.
Rinderer T.E. 1986. Africanized bees: An overview. Am. Bee J.
126:98-100, 128-129.
Rinderer T.E. A.B. Bolten^A.M. Collins and J.R. Harbo. 1984.
Nectar-foraging characteristics of Africanized and European
honeybees in the neotropics. J. Apic. Res. 23:70-79.
Rinderer T.E., A.B. Bolten, J.R. Harbo and A.M. Collins. 1982.
Hoarding behavior of European and Africanized honey bees
(Hymenoptera: Apidae). J. Econ. Ent. 75:714-715.
Rinderer, T.E. A.M. Collins, A.B. Bolten and J.R. Harbo. 1981. Size
of nest cavities selected by swarms of Africanized honeybees in
Venezuela. J. Apic. Res. 20:160-164.
Rinderer T.E. A.M. Collins and K.W. Tucker. 1985. Honey production
and underlying nectar harvesting activities of Africanized and
European honeybees. J. Apic. Res. 24:161-167.
Rinderer T.E., R.L. Hellmich II R.G. Danka and A.M Collins. 1985.
Male reproductive parasitism: A factor in the Africanization of
European honey-bee populations. Science 228:1119-1121.
Rinderer, T.E., and H.A. Sylvester. 1981. Identification of
Africanized bees. Am. Bee J. 121:512-516.
Rinderer, T.E., K.W. Tucker and A.M. Collins. 1982. Nest cavity
selection by swarms of European and Africanized honeybees. J.
Apic. Res. 21:98-103.
Roberts, W.C. 1944. Multiple mating of queen bees proved by progeny
and flight tests. Glean. Bee Cult. 72:255-259, 303.
Roberts, W.C. and S. Taber. 1965. Egg-weight variance in honey bees.
Ann. Ent. Soc. Am. 58:303-306.
Root, A.I. 1947. ABC and XYZ of bee culture. Root Co. Medina, Ohio.
Roubik, D.W. 1978. Competitive interactions between neotropical
pollinators and Africanized honey bees. Science 201:1030-1032.
Roubik, D.W. 1979. Africanized honeybees, stingless bees and the
structure of tropical plant-pollinator communities. Pages 403-417
in D. Caron (ed.), Proc. IVth Inti. Symp. on Pollination. Mise.
Publ. no. 1, Maryland Agrie. Exp. Sta. Univ. of Maryland, College
Park.
Roubik, D.W. 1980. Foraging behavior of competing Africanized
honeybees and stingless bees. Ecology 61:836-845.

177
Roubik, D.W. 1982. Ecological Impact of Africanized honeybees on
native neotropical pollinators. Pages 233-247 in P. Jaisson (ed.),
Social insects in the tropics. Universite Paris-Nord, Paris.
Roubik D.W. 1983. Experimental community studies: Time-series tests
of competition between African and neotropical bees. Ecology
64:971-978.
Roubik, D.W., and S.L. Buchmann. 1984. Nectar selection by Mel 1ipona
and Apis me!1ifera (Hymenoptera: Apidae) and the ecology of nectar
intake by bee colonies in a tropical forest. Oecologia 61:1-10.
Ruttner, F. 1968. Les races dabeilles. Pages 27-44 in R. Chauvin
(ed.) Traite de biologie de labeilles, 1. Masson et Cie Paris.
Ruttner, F. 1975. Races of bees. Pages 19-38 in Dadant and Sons
(eds.) The hive and the honey bee. Dadant and Sons, Inc.
Hamilton, Illinois.
Ruttner, F. 1976a. The races of bees of Africa. Proc. 25th Int.
Beekeap. Congr. 1975, Grenoble. Apimondia, Bucharest, Rumania.
Ruttner, F. 1976b. Honeybees of the tropics: Their variety and
characteristics of importance for apiculture. Pages 41-46 in E.
Crane (ed.), Apiculture in tropical climates. International Bee
Research Assoc. London.
Ruttner, F. 1981. On the taxonomy of honey bees of tropical Africa.
Pages 278-287 in Proc. 28th Int. Congr. Apimondia, Acapulco,
Mexico.
Seeley, T.D. 1977. Measurement of nest cavity volume by the honey bee
(Apis mellifera). Behav. Ecol. Sociobiol. 2:201-227.
Seeley, T.D. 1978. Life history strategy of the honey bee, Apis
mellifera. Oecologia 32:109-118.
Seeley, T.D. 1979. Queen substance dispersal by messenger workers in
honeybee colonies. Behav. Ecol. Sociobiol. 5:391-415.
Seeley, T.D. 1982. Adaptive significance of the age polyethism
schedule in honeybee colonies. Behav. Ecol. Sociobiol. 11:287-293.
Seeley, T.D. 1983. The ecology of temperate and tropical honeybee
societies. Am. Sci. 71:264-272.
Seeley, T.D., and R.D. Fell. 1981. Queen substance production in honey
bee (Apis me!1ifera) colonies preparing to swarm (Hymenoptera:
Apidae). J. Kans. Ent. Soc. 54:192-196.
Seeley, T.D. and R.A. Morse. 1976. The nest of the honey bee (Apis
mellifera L.). Insectes Sociaux 23:495-512.

178
Seeley, T.D., RH. Seeley and P. Akratanakul. 1982. Colony defense
strategies of the honeybees in Thailand. Ecol. Monogr. 52:43-63.
Shearer, D.A., R. Boch, R.A. Morse and F.M. Laigo. 1970. Occurrence of
9-oxodec-trans-2-enoic acid in queens of Apis dorsata, Apis cerana
and Apis mellifera. J. Insect Physiol. 16:1437-1441.
Siegel, S. 1956. Nonparametric statistics for the behavioral sciences.
McGraw-Hill Book Co., New York.
Simpson, J. 1966. Congestion of adult honeybees with and without
adequate hive space. J. Apic. Res. 5:59-61.
Simpson, J. 1973. Influence of hive-space restriction on the tendency
of honeybee colonies to rear queens. J. Apic. Res. 12:183-186.
Simpson, J., and I.B.M. Riedel. 1963. The factor that causes swarming
by honeybee colonies in small hives. J. Apic. Res. 2:50-54.
Smith, F.G. 1958a. Beekeeping observations in Tanganyika 1949-1957.
Bee World 39:29-36.
Smith, F.G. 1958b. Communication and foraging range of African bees
compared with that of European and Asian bees. Bee World 39:
249-252.
Smith, F.G. 1961. The races of honeybees in Africa. Bee World 42:
255-260.
Smith, M.V. 1972. Marking bees and queens. Bee World 53:9-13.
Sokol, R.R., and F.J. Rohlf. 1969. Biometry. W.H. Freeman and Co.,
San Francisco.
Stort, A.C. 1974. Genetic study of aggressiveness of two subspecies of
Apis mellifera in Brazil, 1. Some tests to measure aggressiveness.
J. Apic. Res. 13:33-38.
Stort, A.C. 1975a. Genetic study of aggressiveness of two subspecies
of Apis mellifera in Brazil, 2. Time at which the first sting
reached a leather ball. J. Apic. Res. 14:171-175.
Stort, A.C. 1975b. Genetic study of the aggressiveness of two
subspecies of Apis mellifera in Brazil, IV. Number of stings in the
gloves of the observer. Behavior Genetics 5:269-274.
Stort, A.C. 1975c. Gentica! study of the aggressiveness of two
subspecies of Apis mel1ifera in Brasil, V. Number of stings in the
leather ball. J. Kans. Ent. Soc. 48:381-387.
Stort, A.C. 1976. Genetic study of the aggressiveness of two
subspecies of Apis mellifera in Brazil, III. Time taken for the
colony to become aggressive. Ciencia e Cultura 28:1182-1185.

179
Stort, A.C. and N. Bareli 1. 1981. Genetic study of olfactory
structures in the antennae of two Apis me!1ifera subspecies. J.
Kans. Ent. Soc. 54:352-358.
Sylvester? H.A. 1982. Electrophoretic identification of Africanized
honeybees. J. Apic. Res. 21:93-97.
Taber, S. III. 1954. The frequency of multiple mating of queen honey
bees. J. Econ. Ent. 47:995-998.
Taber, S. III. 1961. Successful shipments of honeybee semen. Bee
World 42:173-176.
Taber, S., Ill, and W.C. Roberts. 1963. Egg weight variability and its
inheritance in the honey bee. Ann. Ent. Soc. Am. 56:473-476.
Taber, S., Ill, and J. Wendel. 1958. Concerning the number of times
queen bees mate. J. Econ. Ent. 51:786-789.
Taylor, O.R. 1977. The past and possible future spread of Africanized
honeybees in the Americas. Bee World 58:19-30.
Taylor, O.R. 1985. African bees: Potential impact in the United
States. Bull. Ent. Soc. Am. 31:14-24.
Taylor, O.R. 1986. Health problems associated with African bees. Ann.
Int. Med. 104:267-268.
Taylor, O.R., R.W. Kingsolver and G.W. Otis. In press. A neutral
mating model for honey bees (Apis me!1ifera L.). J. Apic. Res.
Taylor, O.R., and M. Spivak. 1984. Climatic limits of tropical African
honeybees in the Americas. Bee World 65:38-47.
Taylor, O.R., and G.B. Williamson. 1975. Current status of the
Africanized honey bee in northern South America. Am. Bee J.
115:92-93, 98-99.
Tribe, G.D., and D.J.C. Fletcher. 1977. Rate of development of the
workers of Apis mellifera adansonii L. Pages 115-119 in D.J.C.
Fletcher (ed.), African bees: Taxonomy, biology and economic use.
Apimondia, Pretoria.
Tuenin, T.A. 1927. Variation of bees and their organs. Am. Bee J.
67:19,
Visscher, P.K., and T.D. Seeley. 1982. Foraging strategy of honeybee
colonies in a temperate deciduous forest. Ecology 63:1790-1801.
von Frisch, K. 1967. The dance language and orientation of bees.
Belknap Press, Cambridge, Mass.
Waddington, K.D. 1981. Patterns of size variation in bees and
evolution of communication systems. Evolution 35:813-814.

180
Waddington, K.D., L.H. Herbst and D.W. Roubik. 1986. Relationship
between recruitment systems of stingless bees and with in-nest
worker size variation. J. Kans. Ent. Soc. 59:95-102.
Wafa, A.K., S.E. Rashad and M.M. Mazeed. 1965. Biometrical studies on
the Egyptian honeybee. J. Apic. Res. 4:161-166.
Weiss, K. 1974. Zur frage des koniginnengewichtes in abhangigkeit von
umlarvalter und larvenversorgung. Apidologie 5:127-147.
Weiss, K. 1984. Regulierung des proteinhaushaltes im bienenvolk (Apis
mellifica L.) durch brutkannibalismus. Apidologie 15:339-354.
Wenner, A.M. 1962. Sound production during the waggle dance of the
honey bee. Anim. Behav. 10:79-95.
Wiese, H. 1972 Abel has africanas, suas caractersticas e tecnologa
de manejo. Pages 95-108 in Anais do Io Congresso Brasileiro de
Apicultura. Congresso Brasileiro de Apicultura, Florianopolis,
Brazil.
Wilson, E.O. 1971. The insect societies. Belknap Press, Cambridge,
Mass.
Winston, M.L. 1979a. The potential Impact of the Africanized honey bee
on apiculture in Mexico and Central America. Am. Bee J. 119:584-
586, 642-645.
Winston, M.L. 1979b. Intra-colony demography and reproductive rate of
the Africanized honeybee in South America. Behav. Ecol. Sociobiol.
4:279-292.
Winston, M.L. 1979c. Events following queen removal 1n colonies of
Africanized honey bees in South America. Insectes Sociaux 26:
373-381.
Winston, M.L. 1980a. Swarming, afterswarming and reproductive rate of
unmanaged honeybee colonies (Apis mellifera). Insectes Sociaux
27:391-398.
Winston, M.L. 1980b. Seasonal patterns of brood rearing and worker
longevity in colonies of the Africanized honey bee (Hymenoptera:
Apidae) in South America. J. Kans. Ent. Soc. 53:157-165.
Winston, M.L., J. Dropkin and O.R. Taylor. 1981. Demography and life
history characteristics of two honey bee races (Apis me!1ifera).
Oecologla 48:407-413.
Winston, M.L., and S.J. Katz. 1981. Longevity of cross-fostered honey
bee workers (Apis mellifera) of European and Africanized races.
Can. J. Zool. 59:1571-1575.

181
Winston, M.L. and S.J. Katz. 1982. Foraging differences between
cross-fostered honeybee workers (Apis mel1 ifera) of European and
Africanized races. Behav. Ecol. Sociobiol. 10:125-129.
Winston, M.L., and G.W. Otis. 1978. Ages of bees in swarms and
afterswarms of the Africanized honeybee. J. Apic. Res. 17:123-129.
Winston, M.L., G.W. Otis and O.R. Taylor. 1979. Absconding behaviour
of the Africanized honeybee in South America. J. Apic. Res. 18:85-
94.
Winston, M.L., and O.R. Taylor. 1980. Factors preceding queen rearing
in the Africanized honeybee (Apis mellifera) in South America.
Insectes Sociaux 27:289-304.
Winston, M.L., O.R. Taylor and G.W. Otis. 1983. Some differences
between temperate European and tropical African and South American
honeybees. Bee World 64:12-21.
Woyke, J. 1969. African honey bees in Brazil. Am. Bee J. 109:342-344.
Woyke, J. 1971. Correlations between the age at which honeybee brood
was grafted, characteristics of the resultant queens, and results
of insemination. J. Apic. Res. 10:45-55.
Woyke, J. 1973. Experiences with Apis mellifera adansonii in Brazil
and in Poland. Apiacta 8:115-116.
Woyke, J. 1976. Brood-rearing efficiency and absconding in Indian
honeybees. J. Apic. Res. 15:133-143.
Woyke, J. 1977. Cannibalism and brood-rearing efficiency in the
honeybee. J. Apic. Res. 16:84-94.
Woyke, J. 1979. Effects of the access of worker honeybees to the queen
on the results of the instrumental insemination. J. Apic. Res.
19:136-143.
Woyke, J. 1984. Correlations and interactions between population,
length of worker life and honey production by honeybees in a
temperate region. J. Apic. Res. 23:148-156.
Zeuner, F.E., and F.J. Manning. 1976. A monograph on fossil bees
(Hymenoptera: Apoidea). Bull. Brit. Mus. (Nat. Hist.), Geol.
27:151-268.
Zmarlicki, C. and R.A. Morse. 1963. Drone congregation areas. J.
Apic. Res. 2:64-66.

BIOGRAPHICAL SKETCH
Alan Bolten was born on May 27, 1945 In Newark, New Jersey. In
1959, he was graduated from Maple Avenue Grammar School. Four years
later, he completed h1s secondary education at Weequahic High School in
Newark. He received his undergraduate education from Union College in
Schenectady, New York, where he was graduated with honors in biology in
1967. Alan began graduate studies in the Department of Zoology at the
University of Florida in 1977. He is married to Karen Bjorndal.
182

I certify that I have read this study and that in my opinion it
conforms to acceptable standards of scholarly presentation and is fully
adequate, in scope and quality, as a dissertation for the degree of
Doctor of Philosophy.
1
Thomas C. Emmel, Chairman
Professor of Zoology
I certify that I have read this study and that in my opinion it
conforms to acceptable standards of scholarly presentation and is fully
adequate, in scope and quality, as a dissertation for the degree of
Doctor of Philosophy.
latlran Reiskind
Associate Professor of Zoology
I certify that I
conforms to acceptable
adequate, in scope and
Doctor of Philosophy.
have read this study and that in my opinion it
standards of scholarly presentation and is fully
quality, as a dissertation for the degree of
Malcolm T. Sanford
Associate Professoryof Entomology
and Nematology
This dissertation was submitted to the Graduate Faculty of the
Department of Zoology in the College of Liberal Arts and Sciences and to
the Graduate School and was accepted as partial fulfillment of the
requirements for the degree of Doctor of Philosophy.
August 1986
Dean, Graduate School

o r
zb 0
/;



122
temperatures of temperate winters (Nunez 1979b; Woyke 1973). The other
scenario based on successful hybridization and resultant genetic
introgression would have the stinging behavior of the Africanized honey
bees and the potential public health hazard widespread in North America
and not limited to the southern regions.
A comparison between hybrid and non-hybrid mating success is
lacking for the Africanized and European populations of honey bees.
Although reciprocal FI crosses (Africanized queen x European drone;
European queen x Africanized drone) can be successfully produced with
artificial insemination (Chapater III), natural matings have been
difficult to evaluate because of the inability to distinguish between
hybrid and non-hybrid progeny on the individual level. It is not
possible to determine which drones have mated a queen because honey bees
mate in the air away from the hive, and there are no available genetic
markers in the Africanized population. Identifying individual honey
bees as either Africanized or European is not presently possible
(Carlson and Bolten 1984; Page and Erickson 1985; Rinderer and Sylvester
1981). However, there is promise that DNA analyses will allow
individuals to be identified (Hall in press).
This paper evaluates an important parameter with respect to the
question of successful hybridization between Africanized and European
honey bees: the mating success of both Africanized and European queens
with Africanized drones. The mating success of European queens x
Africanized drones was compared with the mating success of Africanized
queens x Africanized drones in an isolated area in Venezuela with no
European honey bees but with a feral population of Africanized honey
bees. Because of present legal restrictions, Africanized bees cannot be


130
Kin recognition has been suggested as another mechanism that may
help preserve the African genotype in the hybridized honey bee
population in South and Central America (Hall in press). However, data
presented in Table 4-2 demonstrate that this is unlikely because both
Africanized and European worker bees reared both Africanized and
European queens with equal frequency.
There have been several suggestions that Africanized drones may
have a mating advantage over European drones (Kerr and Bueno 1970;
Michener 1975; Morse 1984; Rinderer 1986; Rinderer, Hellmich, Danka and
Collins 1985; Taylor 1985). Taylor (1985) suggests that this mating
advantage would reduce hybridization. On the contrary, a mating
advantage for Africanized drones would increase the rate of
hybridization between Africanized and European honey bee populations.
When Africanized honey bees begin invading North America (Mexico and
U.S.A.), they will be greatly outnumbered by the established European
honey bee population. With a mating advantage, the frequency of
European queen x Africanized drone matings would be greater than
expected based solely on the relative frequency of each population,
thereby resulting in greater hybridization.
Unfortunately, Africanized honey bee research has been
characterized by conceptualizing the Africanized honey bee as a distinct
entity that is reproductively isolated (Taylor 1977, 1985; Taylor and
Spivak 1984) rather than as a population within a species fully capable
of hybridization. Clearly, the Africanized honey bee is not a species
invading a new habitat (North America) that is free of competition from
conspecifics. Thus, the spread of Africanized bees (African genes) in
temperate North America will be: 1) farther north than predicted by the


138
occur. In addition, offspring (swarms) need a rather long period of
time to grow and to hoard necessary surplus honey, while there are still
floral resources available, in order to prepare for winter.
Most mortality of honey bee colonies in temperate areas occurs
primarily due to starvation during winter: 11% for first-year colonies
and 90% for established colonies (Seeley 1978, 1983). There is a high
energetic cost of maintaining proper brood nest or cluster temperature
during the cold winter. In addition, the high energetic cost of winter
survival in temperate regions may greatly reduce the survivorship of
small, secondary swarms or afterswarms, which would greatly reduce the
net reproductive rate of honey bees in temperate regions. In contrast,
periods of resource dearth in the tropics are not only shorter, but
require less stored honey (per unit time) to enable the colonies to
survive because of reduced energetic costs for maintaining brood nest
temperature.
The reproductive season in French Guiana, South America, was nine
months (Winston 1980b) compared with two to four months for North
America (references cited in Winston 1980b and Winston, Dropkin and
Taylor 1981). Is it a coincidence that the difference in the annual
reproductive rate of Africanized bees in South America compared with
European bees in North America, 16 vs. 3.0-3.6, respectively (Otis
1982a; Winston 1980a; Winston, Taylor and Otis 1983), is approximately
of the same order (factor of 4-5) as the difference in the length of the
reproductive season between South America and many areas of North
America?
Another factor affecting differences in reproductive rates between
honey bee populations in South America with those in North America is


60
x 1 mm, and each bag was approximately 5 x 10 cm. Bags were
individually suspended on monofilament line about 6.5 meters above the
ground, centered between two poles 20 meters apart. Queens could be
rapidly raised and lowered by a pulley system. Queens were put into the
mesh bags just prior to testing in order to avoid any pheromone
accumulation.
The testing location was in an open field in a drone congregation
area (Zmarlicki and Morse 1963), which was located by walking with a
helium-filled weather balloon with mature queens suspended 10-20 meters
above the ground. The drone congregation area was identified when
hundreds of drones oriented to the tethered queens. Boundaries appeared
to be quite distinct and stable through time. Both Africanized and
European drones were probably present, but the identity of each drone
responding to specific queens during the experiment was not known
because there are no reliable techniques to identify individual
Africanized and European honey bees. How drone congregation areas
become established is not understood, but these areas are probably where
most mating occurs.
Individual queens were tested for drone response on consecutive
days, beginning on the day of emergence. Only one queen at a time was
tested so that the relative attraction of each queen would not be
influenced by other queens being tested simultaneously. Testing lasted
for a maximum of 3 minutes for each queen, even if no drone response was
observed. Periodically, empty bags (blanks) were tested to insure that
drones were responding only to the queens and not orienting to the
experimental set-up and responding to the mesh bags. At no time did
drones respond to the blanks. A random sequence for testing individual


62
These ranking categories were easily discriminated and were not affected
by the absolute numbers of drones flying. No estimates of the drone
population were made.
Time Post-Emergence to the Initiation of Oviposition
Queens were produced as described above from one Africanized egg
source (A26) and one European egg source (We). Twenty-seven Africanized
and twenty-five European mature queen cells (two days prior to
emergence) were each introduced into a four-frame queenless mating
colony. Any natural queen cells in the mating colonies were destroyed
before introducing the experimental queen cells. This insured that the
only queen in the mating colony would be the experimental queen. When
only one queen cell is present worker bees usually do not confine her
to her cell and the problem of calculating maturation time is avoided.
Because Africanized and European queens did not develop at the same
rate the day of queen emergence was determined by the mean time of
emergence for a sample of sister queens from the same graft that were
left to emerge in an incubator at 35 + 1C. On the eleventh day after
the queens emerged the colonies were inspected and the age of the brood
was evaluated to determine the age post-emergence when the queens had
begun ovipositing. Those colonies in which there were no larvae were
inspected three and five days later.
This experiment took place during the dry season. Clear weather
prevailed so that mating flights were not affected by weather
conditions. Both Africanized and European drones were in the area.


143
growth models are designed for other species in which all individuals
are potential reproductives. For honey bees, individual (or worker bee)
development time is not equal to generation time. Worker bee
development time only affects the interval between^a given change in egg
laying rate and its resulting change in rate of adult emergence. The
difference in worker development time between Africanized and European
honey bees is only one day (Chapter II) and is trivial with respect to
other factors affecting reproductive rates.
Time from virgin queen emergence to initiation of oviposition also
only affects the interval from swarming until adult population increase
begins and not the rate of colony growth (Chapter IV). The results
presented in Chapter IV for maturation rates of Africanized and European
queens were unexpected. European honey bee queens began oviposition at
an earlier age post-emergence than did Africanized queens.
Reproductive Output
Reproductive output is mainly determined by the swarm to swarm
interval and the number of swarms produced per swarming cycle. The
swarm to swarm interval is a function of all the colony demographic
parameters discussed in the previous section and resource utilization
parameters discussed in the next section. Although there are no data on
the swarm to swarm intervals for Africanized and European honey bees
under identical conditions, results from investigations of demographic
parameters reported above suggest that if differences in swarm to swarm
intervals exist, they would not be a result of demographic differences.
Rather, if differences exist, they are hypothesized to be a result of
differences in resource utilization between the two populations (see
below).


logistic support. Laboratory facilities at the Universidad de Oriente
in Jusepin were made available by Professor Dick Pulido.
The research presented in this dissertation and the commitment to
complete the writing could not have been accomplished without the
collaboration, companionship, encouragement and insights of my wife,
Karen Bjorndal, who shared not only the excitement and successes but
also the frustrations and discomforts of Africanized honey bee research.
Finally, I would like to thank my parents, who have always
supported and encouraged my work.
iv


59
The queen-cell-producing colonies were inspected only after the
queen cells had been sealed in order to avoid disturbance which could
affect development times. Once the cells are sealed, cell-producing
colonies only maintain the appropriate temperature for the pupae to
develop normally. On the sixth day after grafting, each sealed cell was
protected by placing a 3 mm wire mesh tube around it to avoid any
problems associated with queens being confined to their cells by worker
bees. In addition, this also prevented any emerged virgin queens from
destroying sealed cells that had not yet emerged. Beginning 24 hours
before any expected queen emergence, the cell-producing colonies were
inspected daily at 0630, 1200 and 1730 hours to record queen emergence.
In two trials, cell-producing colonies were inspected daily at 0630,
1200 and 1730 hours, beginning 24 hours prior to estimated sealing time,
in order to determine unsealed development times.
Development of Attractiveness of Virgin Africanized and European Honey
Bee Queens to Drones
Two Africanized queen mothers (A26 and A57) were removed from feral
colonies in eastern Venezuela. The two European queen mothers (We and
Yk) were shipped to Venezuela from different commercial queen breeders
in southeastern USA.
Queens from the four queen mothers (A26, A57, We and Yk) were
produced as described above. Sealed queen cells were removed from the
cell-producing colony and placed in an incubator (35 + 1C) 48 hours
prior to emergence. After emergence, the queens were marked for
individual identification and maintained in separate cages in a queen
storage colony (Laidlaw 1979).
In order to test for the degree of attractiveness to drones, each
queen was tethered in a clean, plastic screen bag. The mesh size was 1


56
demographic characteristics of Africanized honey bees that account for
high reproductive rates.
The reproductive rate of Africanized honey bees results in a swarm-
to-swarm interval of approximately 90 days (Winston 1979b). During that
period a virgin queen emerges, develops pheromones necessary to attract
drones, and mates; ovarian follicles mature; oviposition is initiated;
and the colony population growth period begins prior to the next
swarming. One expected demographic feature for a population with a high
reproductive rate would be a short queen maturation interval (Fletcher
1977a). For the Africanized queens in French Guiana, the maturation
interval from pupal eclosin to initiation of oviposition was 9.7 days
(Otis 1980), over 10% of their swarm-to-swarm interval (calculated from
Winston 1979b). Fletcher and Tribe (1977b) report that in the parental
African population, oviposition begins on the 8th to 9th day after queen
emergence. European queens begin ovipositing between the 6th and 17th
day after emergence (Laidlaw and Eckert 1962; Oertel 1940; Root 1947).
Otis (1980) calculated that the mean interval from pupal eclosin to
oviposition for European queens (10.7 days) was not significantly
different from that of Africanized queens (9.7 days). However,
comparisons between reported values for both Africanized and European
honey bees are inappropriate because the data were collected under very
different experimental conditions. Therefore, this study was undertaken
to determine if the queen maturation interval for Africanized honey bees
is significantly different than that for European honey bees under
identical conditions. Three aspects of queen maturation were evaluated:
1) larval, pupal and total development time from egg to adult emergence;


27
TABLE 2-3. Summary of hypotheses and tests for evaluating development
times; letters represent treatments (see Table 2-1).
HI: Worker bee development is faster for Africanized genotypes than
for European genotypes.
A x B
C x D
E x F
G x H
A x H
AC x BD
EG x FH
AE x BF
CG x DH
ACEG x BDFH
African'ized comb cell size; Africanized nurse bees
Africanized comb cell size; European nurse bees
European comb cell size; Africanized nurse bees
European comb cell size; European nurse bees
Africanized comb cell size and nurse bees compared with
European comb cell size and nurse bees
Africanized comb cell size; both nurse bee genotypes
combined
European comb cell size; both nurse bee genotypes combined
Africanized nurse bees; both comb cell sizes combined
European nurse bees; both comb cell sizes combined
Both comb cell size and both nurse bee genotype variables
combined
H2: Worker bee development is more rapid in Africanized comb cells
than in European comb cells.
A x E Africanized egg genotype; Africanized nurse bees
C x G Africanized egg genotype; European nurse bees
B x F European egg genotype; Africanized nurse bees
D x H European egg genotype; European nurse bees
AC x EG Africanized egg genotype; both nurse bee genotypes
combined
BD x FH European egg genotype; both nurse bee genotypes combined
H3: Worker bee development is more rapid with Africanized nurse bees
than with European nurse bees.
A x C
E x G
B x D
F x H
AE x CG
BF x DH
Africanized egg genotype; Africanized comb cell size
Africanized egg genotype; European comb cell size
European egg genotype; Africanized comb cell size
European egg genotype; European comb cell size
Africanized egg genotype; both comb cell sizes combined
European egg genotype; both comb cell sizes combined


13
the way In which nurse bees from the two populations interact with the
developing larvae. Mel'nichenko (1962) suggested that differences
between nurse bee genotypes might affect developmental rates as well as
size of developing larvae. For European honey bees, Lindauer (1953)
calculated that each developing larva requires over 2785 adult bee
visits taking a total of 10.3 hours. This appears to provide sufficient
opportunity for possible genotype differences, either quantitative or
qualitative, to affect development rates. In addition to potential
qualitative or quantitative differences in feeding of larvae, nurse bees
of different genotypes may also maintain different brood nest
temperatures. Therefore, development times for Africanized and European
worker bees were evaluated in both Africanized and European colonies.
The third colony-level factor affecting worker development is
colony size (number of worker bees in a colony). Colony size affects
both brood nest temperature and larval feeding rates, which, as already
discussed, are two major factors affecting development times.
In addition to these colony-level parameters, resource conditions
also affect development time. Nelson and Sturtevant (1924) reported
that development of European bees was more rapid with increased larval
feeding associated with a nectar flow. Ribbands (1953) and Jay (1963)
both summarized evidence of the effect of food on larval development
rates. Therefore, all comparisons of worker development times were
conducted simultaneously to avoid any differences due to resource
conditions.
This paper reports the results from a comparison of the development
times of Africanized and European honey bees under identical conditions
in Venezuela. The experimental design allowed for the discrimination


TABLE 6-4. Effect of nurse bee genotypes on the daily
egg laying rates of Africanized and European
queens, trial 2.
MEAN + SD (n)
AFRICANIZED NURSE BEES
AFRICANIZED QUEENS
A61 (W61)
799.5
+
67.2
(2)
AS7 (W41)
694.0
+
39.1
(4)
A62 (W85)
798.7
+
92.3
(3)
COMBINED
752.3
+
79.6
(9)
(A)
EUROPEAN QUEENS
GK30 (Y61)
823.5
+
37.0
(4)
YD5 (Y52)
782.7
+
231.7
(3)
COMBINED
806.0
+
138.0
(7)
(B)
COMBINED AFRICANIZED
AND EUROPEAN QUEENS
775.8
+
108.4
(16)
(C)
EUROPEAN NURSE BEES
AFRICANIZED QUEENS
A57 (W42)
699.8
+
39.5
(4)
A62 (W81)
915.8
+
30.7
(4)
COMBINED
807.8
+
120.0
(8)
(D)
EUROPEAN QUEEN
GK30 (Y63)
736.8
32.2
(4)
(E)
COMBINED AFRICANIZED
AND EUROPEAN QUEENS
784.1
+
103.3
(12)
(F)


TABLE 6-3. Daily egg laying rates of Africanized and European
queens with Africanized nurse bees, trial 2.
ONE-DAY HIGH MEAN + SD (n)
AFRICANIZED QUEENS
A61 (W61)
847
799.5
+ 67.2
(2)
A57 (W41)
725
694.0
t 39.1
(4)
A62 (W85)
883
798.7
+ 92.3
(3)
EUROPEAN QUEENS
GK30 (Y61)
850
823.5
+ 37.0
(4)
YD5 (Y52)
949
782.7
+ 231.7
(3)
ANALYSIS3
NS
^ann-Wh itney U test, two-tailed, alpha = 0.05; evaluated
for all samples (n = 16).


45
Further evidence of the effectiveness of maternal inheritance
reducing size variation can be seen by comparing the size variation
within a colony to the size variation within a population. Alpatov
(1929) found that within honey bee colonies, worker size variation
(e.g., for tongue length) was less than the variation for the local
population of a managed apiary. For seven different apiaries in Russia,
each apiary had an average 22.3% (range 5-42%) increased variation over
the mean colony variation within the apiary. Although the genetic
homogeneity of the apiaries is artificially high as a result of
management practices of the beekeepers compared with the variation of
natural populations of animals (Alpatov 1929), with in-colony variation
was still noticeably reduced.
Evolution of complex communication systems in highly eusocial
species may be responsible for selection for reduced size variation
(Waddington 1981; Waddington, Herbst and Roubik 1986). Foragers within
honey bee colonies have the ability to communicate information to nest
mates about the direction, distance and "profitability" of new resources
(von Frisch 1967) which may be interpreted correctly only if worker bees
within the colony are the same size (Waddington 1981). Profitability of
the resource may be size-dependent as Waddington (1981) suggested. That
is, a high quality resource for a small bee may not be a high quality
resource for a larger bee.
In addition to the profitablity component, correct interpretation
of the distance component of the honey bee waggle dance (von Frisch
1967; Wenner 1962) may also be size-dependent. Different distance
dialects occur not only between subspecies (Boch 1957; Gould 1982) but
also between colonies (Esch 1978 cited in Gould 1982). There is greater


TABLE 2-8. Comparison of differences in development times (in days)
for Africanized and European honey bees for different
developmental stages.
DEVELOPMENTAL STAGES
EGG HRS
(DAYS)3
UNSEALED
BROODb
SEALED
BROOD0
UNSEALED
& SEALED
TOTAL
DEVELOPMENT
AFRICANIZED EGG
GENOTYPE (A26)
69.6
(2.90)
4.3
11.6
15.9
18.80
EUROPEAN EGG
GENOTYPE (Y5)
73.6
(3.07)
4.9
11.9
16.8
19.87
% DIFFERENCEd
5.7
14.0
2.6
5.7
5.7
aFrom Harbo, Bolten, Rinderer and Collins (1981); data used are
their Africanized #3 = A26 and their European #5 = Y5.
Unsealed larval period only.
^Pre-pupae and pupae.
d% Difference = C(Y5)-(A26)/(A26)] x 100.


104
difference. First, the experimental colonies used in this study may
have been only one-tenth the size of the managed, production colonies in
which the maximum rates were observed. Because egg laying rates are
correlated with the number of worker bees in the colony (Moeller 1958),
the lower egg laying rates may have been a result of smaller colonies.
Second, the intensity of colony disturbance, particularly during the
daily egg laying experiment, would have reduced egg laying rates and
increased egg and larval mortality. Third, egg laying rates for
artificially inseminated queens may be lower than for naturally mated
queens (Harbo and Szabo 1984). The purpose of this study was not to
determine absolute egg laying rates but to compare egg laying and brood
production rates for Africanized and European queens during the initial
colony growth phase under identical conditions.
There were no differences in egg laying and brood production rates
for Africanized and European bees during the initial colony growth
phase. When colonies increase in size and approach their maximum growth
phase and queens are maximally challenged, there may be a difference
between Africanized and European queens and colonies. However,
comparisons of queen pupal weights (as a correlate of egg laying rates)
suggest that there would be no difference between Africanized and
European queens with respect to potential egg laying capacity (Chapter
V).
Colony growth rates and therefore reproductive rates are affected
by two other demographic parameters: adult longevity and brood
mortality. Winston and Katz (1981) found that European worker bees were
longer lived than Africanized worker bees under identical conditions in
Venezuela (26.3 compared with 22.7 days). This difference would give


46
variation in individual dialects in colonies that are genetically
heterogeneous compared with colonies that are genetically homogeneous
(Gould 1982). Variation in bee size within a colony may accentuate
differences in distance dialects and increase the possibility of
miscommunication. Therefore, worker size variation within a colony of
honey bees needs to be reduced in order for a communication system that
recruits foragers to a particular floral resource to function correctly
and efficiently with respect to either the profitability (Waddington
1981; Waddington, Herbst and Roubik 1986) or distance component.
Maternal effects operate to reduce bee size variation within a colony of
honey bees, thereby allowing their communication system to function
effectively.
Africanized and European Honev Bee Size Difference
Several hypotheses have been suggested to explain the smaller
worker bee size of the Africanized population. One advantage suggested
for smaller size is more rapid development times, permitting more rapid
colony growth resulting in increased reproductive swarming (Fletcher
1977a; Fletcher and Tribe 1977a; Tribe and Fletcher 1977). However,
cell size and bee size do not affect development times, and, in
addition, worker development times do not affect colony growth rates
(Chapter II).
Fletcher and Tribe (1977a) and Tribe and Fletcher (1977) suggested
that smaller bee size would permit greater numbers of worker bees to be
reared on the same amount of food compared with larger bees. Advantages
of increased worker numbers include frequency of reproductive swarming,
colony defense and foraging success (Wilson 1971). Thus, smaller,
individual bee size maximizes the use of the limited food that


TABLE 5-2. Queen pupal weights for the nine lines
analyzed.
POPULATION QUEEN LINE MEAN PUPAL WEIGHT(MG)
European
GK
233
Africanized
A61
237
Africanized
A26
256
European
N
257
Africanized
A57
257
European
YD
264
European
WE
267
European
YK
280
Africanized
A62
292
ANALYSIS3
NS
^ann-Whitney U test one-tailed, alpha = 0.05


152
comparisons to be made between Africanized and European honey bees that
enable any density-dependent factors responsible for the reproductive
rates and success of Africanized honey bees in South America to be
identified.

Conclusion
In tropical regions, the success of Africanized honey bees compared
with European honey bees is not a function of any intrinsic differences
in colony demography. Rather, it must be concluded that the success of
Africanized honey bees is due primarily to their ability to efficiently
utilize tropical resources, enabling them to survive and reproduce under
conditions where European honey bees are frequently not able to survive.
If European honey bee colonies are not able to survive and/or grow under
some of the tropical resource conditions of South America, they
obviously cannot reproduce. It is precisely because the European honey
bees were not successful foragers (= honey producers) in most tropical
regions of Brazil that African honey bees were imported into Brazil
(Goncalves 1974, 1975, 1982; Woyke 1969).
The efficient utilization of tropical resources by Africanized
honey bees is a result of a set of adaptive behaviors involving solitary
foraging, reduced recruitment, increased brood production efficiency
because of smaller worker bee size, and both resource-induced and
disturbance-induced absconding. These characteristics, combined with an
effective colony defense behavior, give Africanized honey bees a
selective advantage that results in increased survivorship, increased
colony growth rates and ultimately increased reproduction, which is
responsible for their rapid dispersal and high population densities.


180
Waddington, K.D., L.H. Herbst and D.W. Roubik. 1986. Relationship
between recruitment systems of stingless bees and with in-nest
worker size variation. J. Kans. Ent. Soc. 59:95-102.
Wafa, A.K., S.E. Rashad and M.M. Mazeed. 1965. Biometrical studies on
the Egyptian honeybee. J. Apic. Res. 4:161-166.
Weiss, K. 1974. Zur frage des koniginnengewichtes in abhangigkeit von
umlarvalter und larvenversorgung. Apidologie 5:127-147.
Weiss, K. 1984. Regulierung des proteinhaushaltes im bienenvolk (Apis
mellifica L.) durch brutkannibalismus. Apidologie 15:339-354.
Wenner, A.M. 1962. Sound production during the waggle dance of the
honey bee. Anim. Behav. 10:79-95.
Wiese, H. 1972 Abel has africanas, suas caractersticas e tecnologa
de manejo. Pages 95-108 in Anais do Io Congresso Brasileiro de
Apicultura. Congresso Brasileiro de Apicultura, Florianopolis,
Brazil.
Wilson, E.O. 1971. The insect societies. Belknap Press, Cambridge,
Mass.
Winston, M.L. 1979a. The potential Impact of the Africanized honey bee
on apiculture in Mexico and Central America. Am. Bee J. 119:584-
586, 642-645.
Winston, M.L. 1979b. Intra-colony demography and reproductive rate of
the Africanized honeybee in South America. Behav. Ecol. Sociobiol.
4:279-292.
Winston, M.L. 1979c. Events following queen removal 1n colonies of
Africanized honey bees in South America. Insectes Sociaux 26:
373-381.
Winston, M.L. 1980a. Swarming, afterswarming and reproductive rate of
unmanaged honeybee colonies (Apis mellifera). Insectes Sociaux
27:391-398.
Winston, M.L. 1980b. Seasonal patterns of brood rearing and worker
longevity in colonies of the Africanized honey bee (Hymenoptera:
Apidae) in South America. J. Kans. Ent. Soc. 53:157-165.
Winston, M.L., J. Dropkin and O.R. Taylor. 1981. Demography and life
history characteristics of two honey bee races (Apis me!1ifera).
Oecologla 48:407-413.
Winston, M.L., and S.J. Katz. 1981. Longevity of cross-fostered honey
bee workers (Apis mellifera) of European and Africanized races.
Can. J. Zool. 59:1571-1575.


22
contributing to either differences in rate of colony population increase
or to differences in reproductive rates between the two honey bee
populations.
The importance attributed to worker development time on the rate of
colony growth may be a result of confusing colony population increase
(increase in the number of bees in the colony) with general population
growth models designed for other species in which all individuals are
potential reproductives. For honey bees, individual (or worker bee)
development time is not equal to generation time. Organism growth
models must be used to evaluate colony growth even though the number of
individual worker bees within the hive increases. The hive is the
organism. Worker bee development time does not affect the rate of
colony growth. Worker development time affects only the length of time
between a given change in egg laying rate and its resulting change in
population increase or decrease. Africanized bees develop in 19 days
and begin their population increase (^growth) on the 19th day of the
colony cycle, compared with the 20th day for European bees. This
difference is trivial compared to potential differences from other
demographic factors that do affect rates of colony growth. Egg laying
and brood production rates, worker bee longevity, brood mortality, and
resource availability are factors that do affect the rate of colony
population increase and, therefore, affect the reproductive rates.
Tribe and Fletcher (1977) have suggested that African worker bees
have a shorter unsealed development stage because they do not grow as
large as European honey bees. They compare their data for African bees
with data for European bees in the literature and conclude that African
bees have a 20-30% shorter unsealed larval stage. There are four


34
TABLE 2-9. Mortality during different developmental stages.
AFRICANIZED EGG GENOTYPE (A26) EUROPEAN EGG GENOTYPE (Y5)
E1 E2 l1 L2 SB N
AFRICANIZED COMB
CELL SIZE
AFRICANIZED
NURSE BEES
A55 1 5 0 3 1 40 1 0 0 2 0 40
EUROPEAN
NURSE BEES
H2 1 0 3 7 0 40 1 0 0 13 0 36
EUROPEAN COMB
CELL SIZE
AFRICANIZED .
NURSE BEES
A41 0 0 0 0 0 30 8 2 0 0 0 29
EUROPEAN
NURSE BEES
IBR877 00220 30 13 6100 27
^Mortality during first 24 hours in test colony (acceptance).
^Mortality between 24-72 hours (before hatching).
^Mortality between 72-96 hours (at time of hatching).
Mortality during older larval stages before sealing.
^Mortality during the pupal stage.
fN = total eggs monitored.


131
geographic limits of the parental population because of hybridization
and resultant genetic introgression; 2) slowed considerably by
competition for available resources by an established population of
European honey bees; 3) swamped through hybridization with a more
numerous, established population of European honey bees; 4) at a
disadvantage with respect to foraging behavior; and 5) limited by
selection against those colonies that have not acquired through
hybridization the ability to overwinter.
There has been a general lack of support for the selectionist
argument for the maintenance of the African genotype in favor of the
hypothesis of reproductive isolation. With the demonstration that
hybridization is successful, coupled with the recent observations of the
distribution patterns of Africanized honey bees in Argentina (Dietz,
Krell and Eischen 1985; Krell, Dietz and Eischen 1985), further
consideration of the selectionist argument for the maintenance of the
African characteristics in the Africanized honey bee in South and
Central America is necessary.


51
TABLE 3-3. Worker bee size: hypotheses and analyses (Mann-
Whitney U test, one-tailed, alpha = 0.05).
Letters refer to experimental treatments, see
Table 3-1.
HI: Africanized bee pupae are smaller than European bee
pupae independent of comb cell size.
ACE
X
MOQ
***a
BDF
X
NPR
***
ACE
X
N3 R
***
BDF
X
MOQ
NS
H2: For a given egg genotype, pupae that develop in
Africanized comb cell size are smaller than pupae
that develop in European comb cell size.
A x
B
***
C x
D
***
E x
F
***
G x
H
***
I X
J
***
K x
L
***
M x
N
***
0 x
P
***
Q x
R
***
ACE
x BDF
***
MOQ
x rPR
***
a*** = p

TABLE 2-3continued.
H4: Worker bee development is more rapid with Africanized comb cells
and Africanized nurse bees than with European comb cells and
European nurse bees.
A x G
B x H
Africanized egg genotype
European egg genotype


135
Taylor 1981 and Winston, Taylor and Otis 1983). Rather, the question
is: what are the differences between Africanized and European
populations of A. mel1ifera that make Africanized honey bees more
successful in tropical regions not only in South America but also in
Africa? European honey bees have not been successfully introduced into
tropical areas of Africa despite numerous attempts (Fletcher 1977b,
1978).
Factors Affecting Honey Bee Reproductive Rates
The success of Africanized honey bees in South Americaas judged
by their rate of dispersal and their population densities (Michener
1975; Taylor 1977, 1985)must surely be a result of a high reproductive
rate. What are the differences between Africanized and European honey
bees that allow for high reproductive rates in Africanized bees, and can
these differences account for the impact of Africanized honey bees?
This question does not involve identification or analysis of the
proximal factors that are responsible for initiating reproduction, but
does involve analysis of the components that affect the rate of
reproduction.
Reproductive rates in honey bees are a result of an interaction of
at least three factors, all of which affect colony growth rates:
resource availability, resource utilization efficiency (foraging
success, brood production efficiency, and bee size), and colony
demographic parameters (primarily queen fecundity and adult worker bee
longevity). Therefore, in order to evaluate reproductive differences
between Africanized and European honey bee populations, all three
factors need to be considered. Unfortunately, early research evaluated
reproductive rates of Africanized honey bees by comparing data for


10
parameter that has been compared between Africanized and European bees
under identical conditions. The greater longevity of European honey
bees (Winston and Katz 1981) gives European bees a colony growth rate
advantage. Other demographic characteristics that affect reproductive
rates of Africanized and European honey bees (for example, worker bee
development times, brood mortality, queen development and maturation
periods, queen fecundity and brood production rates) have not been
evaluated for Africanized and European bees under similar conditions.
As part of a larger study evaluating these demographic parameters, this
study compares worker bee development periods for Africanized and
European honey bees in Venezuela.
Smith (1958a) and Tribe and Fletcher (1977) reported that the total
development period (from oviposition to adult emergence) for worker bees
of Apis mellifera adansonii (now classified as &. m, scutellata: Ruttner
1976a, 1976b, 1981) from South Africa was between 18.6-20 days. Similar
development times for the Africanized honey bee populations (descendents
of A. m. scutellata) in Brazil have been presented (Kerr, Goncalves,
Blotta and Maciel 1972; Wiese 1972). Worker bee development times for
European populations (primarily A. m. mel1ifera, 1ioustica, carnica and
caucasica) from Europe and North America range from 20-24 days (Jay
1963).
The differences in development times between African (and
Africanized) and European genotypes, which range from 1.4 to 5.4 days,
are difficult to evaluate because they are based on data collected under
very different experimental conditions. Jay (1963) summarized a number
of factors that affect development times: seasonal variation in
temperature; temperature differences in different areas within the brood


175
Nunez J.A 1979a. Times spent on various components of foraging
activity: Comparison between European and Africanized honeybees in
Brazil. J. Apic. Res. 18:110-115.
Nunez J.A. 1979b. Comparative study of thermoregulation between
European and Africanized Apis me!1 ifera in Brazil. J. Apic. Res.
18:116-121.
Nunez J.A. 1982. Comparacin del comportamiento recolector de abejas
Africanizadas y abejas Europeas. Pages 221-231 in P. Jaisson
(ed.) Social insects in the tropics. Universite Paris-Nord
Paris.
Oertel E. 1940. Mating flights of queen bees. Glean. Bee Cult.
68:292-293.
Otis G.W. 1980. The swarming biology and population dynamics of the
Africanized honeybee. Ph.D. Dissertation, Univ. of Kansas,
Lawrence.
Otis, G.W. 1982a. Population biology of the Africanized honey bee.
Pages 209-219 in P. Jaisson (ed.), Social insects in the tropics.
Universite Paris-Nord, Paris.
Otis, G.W. 1982b. Weights of worker honeybees in swarms. J. Apic.
Res. 21:88-92.
Otis, G.W., and O.R. Taylor. 1979. Beekeeping in the Guianas. Pages
145-154 in Beekeeping in rural development: Unexploited beekeeping
potential in the tropics. Commonwealth Secretariat, London.
Otis, G.W., M.L. Winston and O.R. Taylor. 1981. Engorgement and
dispersal of Africanized honeybee swarms. J. Apic. Res. 20:3-12.
Page, R.E. Jr. and E.H. Erickson, Jr. 1984. Selective rearing of
queens by worker honey bees: Kin or nestmate recognition. Ann.
Ent. Soc. Am. 77:578-580.
Page, R.E. Jr. and E.H. Erickson, Jr. 1985. Identification and
certification of Africanized honey bees. Ann. Ent. Soc. Am.
78:149-158.
Page, R.E. Jr. and R.A. Metcalf. 1982. Multiple mating, sperm
utilization and social evolution. Am. Nat. 119:263-281.
Peer, D.F. 1956. Multiple mating of queen honey bees. J. Econ. Ent.
49:741-743.
Pellett F.C. 1938. History of American beekeeping. Collegiate Press,
Inc., Ames, Iowa.
Reid, M. 1975. Storage of queen honeybees. Bee World 56:21-31.


173
Krell, R., A. Dietz and F.A. Eischen. 1985. A preliminary study on
winter survival of Africanized and European honey bees in Cordoba*
Argentina. Apidologie 16:109-118.
Kulzh inskaya, K.P. 1956. The role of the food factor in the growth of
bees. Apic. Absts. 7:177.
Laidlaw, H.H., Jr. 1979. Contemporary queen rearing. Dadant and Sons,
Inc., Hamilton, Illinois.
Laidlaw, H.H., Jr., and J.E. Eckert. 1962. Queen rearing. Univ. of
California Press, Berkeley.
Lindauer, M. 1953. Division of labour in the honeybee colony. Bee
World 34:63-73, 85-90.
Mackensen, 0. 1964. Relation of semen volume to success in artificial
inseminations of queen honey bees. J. Econ. Ent. 57:581-583.
Mackensen, 0., and W.C. Roberts. 1948. A manual for the artificial
insemination of queen bees. USDA-ARA Bur. Ent. and Plant Quar.
ET-250.
Mackensen, 0., and K. Tucker. 1970. Instrumental insemination of queen
bees. Agrie. Hdbk. 390. USDA, Washington, D.C.
McDowell, R. 1984. The Africanized bee in the United States: What
will happen to the U.S. beekeeping industry? Agrie. Econ. Rpt.
519. USDA, Washington, D.C.
McGregor, S.E. 1938. Environmental factors and size variations in
honeybee appendages. J. Econ. Ent. 31:570-573.
McLellan, A.R. 1978. Growth and decline of honeybee colonies and
inter-relationships of adult bees, brood, honey and pollen. J.
Appl. Ecol. 15:155-161.
Medler, J.T. 1962. Morphometric studies in bumble bees. Ann. Ent.
Soc. Am. 55:212-218.
Medler, J.T. 1965. Variation in size in the worker caste of Bombus
fervidus (Fab.). Pages 388-389 in P. Freeman (ed.), Proc. Xllth
International Congress of Entomology. Royal Entomological Society
of London, London.
Melampy, R.M., and E.R. Willis. 1939. Respiratory metabolism during
larval and pupal development of the female honeybee (Apis mellifera
L.). Physiol. Zool. 13:283-293.
Mel'nichenko, A.N. 1962. Experiments in directed alteration of the
characteristics of female and male honeybees in rearing in the hive
of a different variety. Biol. Abs. 39:661.


140
picture as to the factors leading to the success and resulting impact of
the Africanized honey bee in South America.
Factors Contributing to the Selective Advantage of
Africanized Honev Bees in South America
Colony Demography
Table 8-1 summarizes the factors affecting colony survival and
reproductive success for both Africanized and European honey bees under
tropical conditions in Venezuela. Studies comparing parameters of
colony demography for Africanized and European honey bees under
identical conditions in Venezuela have produced surprising results (see
Chapters IIVI). These studies were based on the assumption that the
life history of the Africanized honey bee population in South America
(as well as the parental population in Africa) was characterized by a
high reproductive rate. Demographic features that were expected to be
correlated with this high rate of colony reproduction, or short swarm to
swarm interval, were shorter worker bee development time, smaller worker
bee size, more rapid queen development and maturation, increased egg
laying and brood production, reduced brood mortality, and increased
adult worker bee longevity.
Colony demographic characteristics can be divided into two groups:
those affecting the rate of colony growth and those affecting the time
interval from swarming to the beginning of adult population increase. A
rapid colony growth rate is most important for a high colony
reproductive rate and is primarily a function of queen fecundity, adult
worker longevity, and brood mortality (Brian 1965; Moeller 1961; Wilson
1971).


136
Africanized honey bees from South America with data from studies of
European honey bees from North America, which not only were collected
under different resource conditions but also different experimental
conditions (Otis 1980, 1982a; Winston 1979b, 1980a; Winston, Dropkin and
Taylor 1981).
Results from these earlier studies characterized the Africanized
population as one with a dramatically high annual colony reproductive
ratefour to five times greater than European honey bees in temperate
regions (Otis 1980, 1982a; Winston 1980a; Winston, Taylor and Otis
1983). However, because these comparisons were not based on data
collected under similar environmental or experimental conditions, they
are inappropriate comparisons and cannot be used to identify either the
factors responsible for the difference in reproductive rates between the
two populations or the factors responsible for the success of
Africanized honey bees. Because the experimental conditions were
different (e.g., hive volume), these comparisons were also inappropriate
for comparing temperate and tropical honey bee populations. Were the
apparent differences in colony reproduction between the two honey bee
populations the result of differences in: 1) colony demography; 2)
environmental and climatic factors; 3) experimental design; 4) resource
utilization efficiencies; or 5) some combination of factors? Are there
intrinsic differences between the two honey bee populations with respect
to colony demographic parameters that allow for a more rapid colony
growth rate and result in a greater reproductive rate for the
Africanized honey bee population? Or, are the differences in
reproductive rates a result of climatic conditions and/or resource
availabilities and utilization in the tropics compared with temperate


7
hybridization or reproductive isolation between Africanized and European
honey bee populations could result in very different scenarios for the
potential impact of Africanized honey bees on North America,
particularly the U.S.A.
Identification of Honey Bees Used in My Research
For the experiments presented here, Africanized honey bee colonies
were established from queens removed from feral colonies in an area in
eastern Venezuela where there were no known European honey bees. They
were identified as Africanized bees primarily by their distinctly
smaller comb cell size as compared with European honey bees.
European honey bees used in the experiments were from commercially
produced queens from three different queen breeders in the U.S.A.
Additional European lines were obtained from the U.S. Department of
Agriculture Bee Research Laboratories in Madison, Wisconsin, and Baton
Rouge, Louisiana. All of these European queens were either naturally
mated or instrumentally inseminated in the U.S.A. and then shipped to
Venezuela.
When and Where Research Was Conducted
All field research with Africanized and European honey bees was
conducted from December 1978 through February 1980 at the Ministerio de
Agricultura y Cria de Venezuela Africanized Honey Bee Research
facilities near Maturin, Monagas. The area originally was a Tropical
Dry Forest [sensu Holdridge Life Zone System (Holdridge 1964; Ewel and
Madriz 1968)]. The forest had been partially cleared, and the area was
grazed by cattle.


TABLE 3-1. Effect of comb cell size and egg genotype
on bee pupal weights. Experimental design
matrix (code letters A-R used in tables of
statistical analyses).
COMB CELL SIZE
EGG GENOTYPES AFRICANIZED EUROPEAN
AFRICANIZED QUEEN X
AFRICANIZED DRONE
A26a
A57a
B39a
A
C
E
B
D
F
AFRICANIZED QUEEN X
EUROPEAN DRONE
SDA12b G H
EUROPEAN QUEEN X
AFRICANIZED DRONE
SDY10b I J
SDYllb K L
EUROPEAN QUEEN X
EUROPEAN DRONE
YD28b
WEla
SDYlb
M
0
Q
N
P
R
aNatural matings, multiple inseminations
bSingle drone insemination.


CHAPTER VIII
DISCUSSION: FACTORS CONTRIBUTING TO THE SELECTION ADVANTAGE OF
AFRICANIZED HONEY BEES IN SOUTH AMERICA--
THE RESOURCE UTILIZATION EFFICIENCY HYPOTHESIS
Success of Introduced Populations of Honey Bees
Thirty years ago African honey bees, Apis me!1ifera scute!1 ata
[formerly classified as A. m. adansonii (Ruttner 1976a, 1976b, 1981)],
were introduced into southeastern Brazil (Kerr 1967). Offspring, known
as Africanized honey bees because of hybridization with European honey
bees (Goncalves 1982), have rapidly dispersed throughout South America,
sometimes achieving dramatically high population densities (Michener
1975; Taylor 1977, 1985). In 1982 Africanized honey bees entered Panama
(Buchmann 1982) and by 1986 were as far north as Honduras and El
Salvador (Rinderer 1986). The success and biological impact of
Africanized honey bees in these tropical and sub-tropical regions,
compared with the lack of success of European honey bees in these same
regions, is a result of a selection advantage for the Africanized (=
African) genotype in tropical resource and climatic conditions. The
difference in success between Africanized and European honey bees is
evidenced by the fact that
European bees in Brazil were never commonly found living wild
in the forests and countryside. This was especially true in
tropical forest regions, where honey bees were virtually
restricted to a few apiaries...Everyone questioned on the
matter emphasized the increase in bees away from apiaries that
occurred with the arrival of the Brazilian [Africanized] bees.
(Michener 1972, p. 15).
133


32
TABLE 2-7. Comb cell size for worker development time experiment: comb
measurements = mm for 10 consecutive, horizontal cells, mean
+ SD, (sample size).
COMB CELL SIZE
NURSE BEE AFRICANIZED EGG EUROPEAN EGG
COLONY GENOTYPE GENOTYPE
AFRICANIZED COMB CELL SIZE3
AFRICANIZED NURSE
BEES (A55)
47.5 + 0.58
(4)
49.8 + 0.50
(4)
EUROPEAN NURSE
BEES (H2)
EUROPEAN COMB CELL SIZE5
48.2 + 0.96
(4)
48.5 + 0.58
(4)
AFRICANIZED NURSE
BEES (A41)
54.0 + 0.0
(3)
54.0 + 0.0
(3)
EUROPEAN NURSE
BEES (IBR877)
53.3 + 0.58
(3)
53.3 + 0.58
(3)
45.8 + 0.50
(4)
48.5 + 0.58
(4)
54.0 + 0.0
(3)
53.7 + 0.58
(3)
3Natural comb built without foundation.
bBuilt from foundation.


52
TABLE 3-4. Reciprocal F-, cross. Pupal weights (mg)> mean + SD,
(sample size). Data are from European comb cell size only.
AFRa QUEEN
AFR QUEEN
EUR'
a QUEEN
EUR QUEEN
X
X
X
X
AFR DRONE0
EUR DRONE0
AFR
DRONE0
EUR DRONE*
(A26)
(SDA12)
(SDY10)
(SDY11)
(SDY1)
.21.7 +5.5
122.4 + 4.4
133.7 +3.3
138.9 + 4.9
135.0 + 6.<
(80)
(40)
(23)
(30)
(19)
B
H
J
L
R
aAFR = Africanized; EUR = European.
Naturally mated.
Single-drone, artificial insemination.


144
Although Africanized and European honey bees have not been compared
under identical conditions, European colonies in North America (Kansas)
produced the same number of small, secondary swarms, or afterswarms, per
swarming cycle as did Africanized colonies in South America (French
Guiana) (Otis 1980; Winston 1980a; Winston, Dropkin and Taylor 1981).
These results are not directly comparable, but they do demonstrate that,
at least under certain conditions, European honey bees can produce as
many afterswarms as Africanized honey bees. Whether the number of
afterswarms for European bees would be similar to Africanized bees under
identical conditions needs to be analyzed. As discussed above,
survivorship of small afterswarms would be much lower in temperate
regions than in tropical regions, because of the energetic demands of
temperate winters on honey bee colonies.
Resource Utilization
The most important factors leading to the success of Africanized
honey bees in South America are associated with resource utilization:
foraging behavior, brood production efficiency, worker bee size and
absconding behavior. Although evaluated under different conditions, the
foraging range of the parental African population is similar to that of
European bees (Smith 1958b). However, under resource conditions typical
of tropical regions, the foraging behavior of Africanized honey bees is
significantly more successful than that of European honey bees
(Rinderer, Bolten, Collins and Harbo 1984; Rinderer, Collins and Tucker
1985), Their success is a result of more frequent solitary foraging and
reduced recruitment when resources are dispersed and limited, as is
characteristic of most tropical habitats (Rinderer, Bolten, Collins and
Harbo 1984; Rinderer, Collins and Tucker 1985). Using artificial


TABLE 5-3. Comparison of Africanized and European
lengths (mm): mean + SD, (sample size)
(genotypes).
queen cell
9
AFRICANIZED
CELL BUILDER3 GENOTYPES
EUROPEAN
GENOTYPES
ANALYSES13
1 2.58 + 0.1
(9) (A26)
2.50 + 0.1
(9) (YK)
NS
2 2.62 + 0.2
(9) (A26)
2.70 + 0.1
(9) (YK)
*
3 2.54 + 0.1
(3) (A26)
2.48 + 0.1
(9) (WE)
NS
4 2.58 + 0.1
(7) (A57)
2.44 + 0.1
(11) (YK)
c
5 2.67 + 0.1
(3) (A26)
2.57 + 0.1
(3) (YK)
NS
6 2.50 + 0.02
(4) (A57)
2.57 + 0.1
(5) (YD)
NS
7 2.78 + 0.04
(5) (A62)
2.81 + 0.1
(3) (N)
NS
8 2.79 + 0.1
(5) (A61)
2.76 + 0.1
(2) (GK)
NS
aCen-producing colonies; European nurse bees and European comb
cell size.
bMann-Whitney U test, one-tailed, alpha = 0.05; = P<0.05.
difference in wrong direction for one-tailed test; two-tailed
test results in a P<0.02.


98
Two days prior to the beginning of the experiment young bees were
removed from brood frames of large colonies and put into screened cages
that measured 48 x 37 x 76 cm. One cage contained Africanized bees and
the other contained European bees. Africanized bees were removed only
from colonies that were being managed on combs of European cell size.
The cages were supplied with feeders containing 50% (volume:volume)
sugar syrup. Bees used to stock the experimental colonies were taken
from the appropriate cage, thereby insuring that all experimental
colonies were uniform in composition for a particular worker bee type,
Africanized or European. Each experimental colony was started with
approximately 775 grams of bees.
Test queens were introduced into each experimental colony using a
push-in cage (Laidlaw 1979). Queens were manually released from the
push-in cages after two days.
The experiment consisted of two separate trials. Trial 1 evaluated
colony treatments 2 and 4 only. Trial 2 evaluated all four colony
treatments simultaneously. Trial 2 was started with a new supply of
worker bees. Some of the queens used in trial 2 had also been evaluated
in trial 1.
Egg laying rates were determined by removing the frames from each
colony every 24 hours and counting the number of eggs. The removed
frames were immediately replaced with empty frames in order to minimize
disturbance. The frames with eggs were stored in a freezer until the
eggs were counted. Initial egg laying rates that were more than two
standard deviations from the mean were not used because these rates may
have occurred before queen maturation was complete. Experimental
colonies with Africanized worker bees (trial 2) began absconding


CHAPTER V
QUEEN PUPAL WEIGHTS
Introduction
Africanized honey bees in South America are hybridized descendents
of African honey bees (Apis me!1 ifera scute!lata) and European honey
bees (primarily A. ¡n. 1 igustica and A. m.. me! 1 if era! (Goncalves 1982;
Woyke 1969). The annual net reproductive rate of Africanized honey bees
in South America is four to five times greater than that of European
honey bees in temperate regions: 16 colonies per colony per year
compared with 3-3.6 (Otis 1980 1982a; Winston 1980a; Winston, Taylor
and Otis 1983). Differences in reproductive rates between these two
honey bee populations may be a result of: 1) colony demography;
2) temperate vs. tropical climate and floral resources; 3) resource
utilization behaviors; or 4) a combination of factors. Because
Africanized and European honey bees have not been compared under
identical experimental conditions, it is not possible to determine to
what extent reproductive differences are a result of genetic or
environmental parameters.
One demographic parameter associated with rapid colony growth and a
high rate of colony reproduction would be a high oviposition rate (Brian
1965; Moeller 1961; Wilson 1971). In the evolution of social insects,
queen oviposition rates have increased primarily due to one of the
following: increased number of ovarioles, increased length of the
79


TABLE 6-6. Daily egg laying rates of Africanized and European queens
comparison between genotypes within each population.
MEAN + SD (n) ANALYSES
AFRICANIZED QUEENS
1. TRIAL 1EUROPEAN NURSE BEES
A57 (W42)
473.2
+
68.5
(6)
A62 (W81)
825.0
+
47.0
(3)
P<0.05
2.
TRIAL 2EUROPEAN NURSE BEES
A57 (W42)
699.8
+
39.5
(4)
A62 (W81)
915.8
+
30.7
(4)
P 3.
TRIAL 2AFRICANIZED NURSE BEES
A61 (W61)
799.5
+
67.2
(2)
A57 (W41)
694.0
+
39.1
(4)
A62 (W85)
798.7
+
92.3
(3)
NSb
4.
COMBINING §2 AND #3
P. EUROPEAN QUEENS
1.
TRIAL 1EUROPEAN NURSE BEES
YK (Y4)
922.0
+
41.0
(2)
*c
YD5 (Y42)
689.6
+
66.4
(5)
GK30 (Y63)
636.7
+
80.8
(3)
GK30 (Y64)
575.6
+
101.3
(5)
NSb
2.
TRIAL 2EUROPEAN NURSE BEES
GK30 (Y63)
736.8
+
32.2
(4)
3.
TRIAL 2AFRICANIZED NURSE BEES
GK30 (Y61)
823.5
+
37.0
(4)
YD5 (Y52)
782.7
+
231.7
(3)
NSa
4.
COMBINING #2 AMD #3
NSb
Mann-Whitney U test, two-tailed, alpha = 0.05.
bKruskal-Wal1 is one-way analysis of variance by ranks, alpha = 0.05.
CYK (Y4) was significantly different, P<0.05, from other genotypes in
the group when evaluated by pairs using the Mann-Whitney U test,
two-tailed, alpha = 0.05.


TABLE C-l. Changes in European queen pupal weights (mg) with
changes in age: mean + SD, (sample size). Queen pupal
weights were measured in Baton Rouge, Louisiana.
DAYS POST-OVIPOSITION
9
10
11
12
13
14
311.6
+ 12.4
(10)
291.6
+ 8.3
(10)
294.6
+ 9.7
(10)
293.4
+ 8.4
(10)
287.4
+ 11.2
(10)
274.1
+ 16.9
(5)
A
B
C
D
E
F
ANALYSES3
ABCDEF
BCDE
P<0.001
NS
a0ne-way analysis of variance, alpha = 0.05.
165


ages when queens attracted drones and the ages when oviposit ion was
initiated.
Daily egg laying rates and brood production during initial colony
growth were not significantly different for Africanized and European
queens. Africanized and European worker bees did not differentially
affect egg laying and brood production rates.
Differences in reproductive rates between Africanized and European
honey bees in South America cannot be attributed to differences in
intrinsic demographic factors. A hypothesis based on differences in
resource utilization efficiency is presented to explain the success of
Africanized bees compared with European bees in South America.
Results from reciprocal crosses indicate that bee size is a
function of egg genotype, comb cell size and maternal genotype. The
importance of maternal inheritance for reducing worker bee size
variation within a colony is discussed. Advantages of smaller worker
bee size are evaluated for Africanized bees.
There are no effective reproductive isolating mechanisms operating
between Africanized and European honey bee populations. Both
Africanized and European queens mated with equal success with
Africanized drones as measured by the numbers of spermatozoa in the
spermatheca. The potential impact of Africanized bees on North America
is analyzed with respect to hybridization and genetic introgression,
resource competition, and selection advantages for European bees in
temperate regions.
v i i i


142
Africanized and European colonies (Winston, Dropkin and Taylor 1981),
which were observed under very different conditions, need to be re
evaluated, and new studies should be undertaken.
Also affecting colony growth rates is the extent to which brood
rearing ceases during periods of resource dearth. As discussed earlier,
Winston (1980b) reports that Africanized bees do not cease brood rearing
to the extent observed for European bees and are therefore capable of
rapid colony growth when conditions improve. European honey bees, under
some tropical conditions, may have a sharp decline in brood rearing
during resource shortages (Otis and Taylor 1979). However, these
differences in brood rearing were not apparent when both Africanized and
European honey bees were managed under identical conditions in Venezuela
(Bolten, personal observation). Therefore, this behavior needs to be
analyzed with both honey bee populations under a variety of tropical
resource conditions to determine if there are differences, and whether
the differences are a function of foraging behavior (see below) and/or
intrinsic demographic parameters.
The two most important factors affecting the interval from swarming
to the beginning of adult population increase are worker development
time and queen maturation (Chapters II and IV). Worker development time
has previously been considered an important factor affecting the rate of
colony growth (Fletcher 1977a, 1978; Fletcher and Tribe 1977a; Tribe and
Fletcher 1977; Winston 1979b; Winston, Dropkin and Taylor 1981; Winston
and Katz 1982; Winston, Taylor and Otis 1983). As discussed in Chapter
II, this is incorrect and is probably a result of confusing models for
colony growth (increase in the number of bees in the colony) with models
for population growth (increase in the number of colonies). Population


99
(abandoning the hive) after five to six days because of the disturbance
caused by the experimental procedure. The experiment was terminated at
that point and only egg laying rates prior to the beginning of
absconding were compared.
Brood Production Rates during First Brood Cycle
The same queens evaluated in the egg laying experiment were used
for this experiment. They were maintained in separate cages in a queen
storage colony (Laidlaw 1979; Reid 1975) for six days between
experiments. Four colony treatments were established as described
above, except that the hives were larger and were stocked with more
worker bees. Standard Langstroth hive bodies (48 liters) were used with
nine frames (all with European comb cell size): eight empty, drawn
combs and one filled with honey. All frames were weighed prior to being
put into the colonies in order that weight changes could be monitored.
Different geometric designs were placed above the entrances to
facilitate orientation and reduce drift between colonies.
As described for the first experiment, young worker bees were
collected and maintained in screened cages for two days prior to the
beginning of the experiment. Approximately 1200 grams of worker bees
were used to stock each colony. The number of bees put into each colony
was estimated by determining the mean individual bee weight from three
20-30 bee samples for each cage and then dividing the total weight of
introduced bees by the mean individual bee weight (Otis 1982b).
Queens were introduced into the experimental colonies using push-in
cages and manually released two days later in order to standardize the
starting day (day 0) for all experimental colonies. Queens that had
been in Africanized colonies for the daily egg laying rate experiment


119
success and length of time throughout the year that resources are
available in the tropics, Africanized honey bees have a high annual
reproductive rate, which is responsible for both their rate of dispersal
into new areas and high colony densities. Net reproductive rates for
Africanized bees have been estimated to be 16 colonies per colony per
year based on demographic data collected in French Guiana (Otis 1980,
1982a) compared with 0.92-0.96 (Seeley 1978) or 3-3.6 when afterswarms
are counted (Winston 1980a; Winston, Taylor and Otis 1983) for European
honey bees in North America.
Particularly in the region of their importation (southeastern
Brazil), there has been ample opportunity for hybridization with both
managed and feral European honey bees [primarily A, m. me!1ifera and A.
m. 1igustica which had been imported into Brazil by 1845 (Gerstaker
cited in Pellet 1938; Woyke 1969)]. However, despite opportunity for
hybridization, Africanized honey bees have maintained behavioral,
chemical and morphological characteristics similar to their African
parental population and distinguishable from European honey bees:
colony defense behavior (=stinging behavior) (Collins, Rinderer, Harbo
and Bolten 1982; Stort 1974, 1975a, 1975b, 1975c, 1976); reproductive
rates (Fletcher 1978; Fletcher and Tribe 1977a; Otis 1980, 1982a);
absconding behavior (reviewed by Fletcher 1978; Winston, Otis and Taylor
1979; Winston, Taylor,and Otis 1983); foraging and hoarding behavior
(Nunez 1973, 1979a, 1982; Rinderer, Bolten, Collins and Harbo 1984;
Rinderer, Bolten, Harbo and Collins 1982; Rinderer, Collins and Tucker
1985; Winston and Katz 1982); worker bee longevity (Winston and Katz
1981); development times (Chapters II and IV; Harbo, Bolten, Rinderer
and Collins 1981); selection preferences for nest cavity sizes


121
these bees evolved. Two lines of evidence support this selectionist
argument. First, the foraging behavior of the Africanized honey bees,
characterized by solitary foraging and less colony recruitment, would be
more adaptive in tropical areas with rich, but dispersed, resources
(Rinderer, Bolten, Collins and Harbo 1984; Rinderer, Collins and Tucker
1985). Second, there is much historic evidence that European honey bees
have not been successfully maintained (probably due to starvation) in
many areas of tropical South America that are now densely populated with
Africanized honey bees (Bolten, personal observation; Michener 1972;
Winston, Taylor and Otis 1983).
The selectionist argument allows for hybridization between
Africanized and European populations, with the African genotype being
selected for under the physical and biotic parameters characteristic of
tropical areas. Selection for the African genotype would then account
for the present population in South and Central America being
behaviorally, chemically and morphologically similar to the African
parental type.
These two alternative hypotheses for the maintenance of the African
parental characteristicsreproductive isolation between the Africanized
and European honey bee populations versus selection for the African
genotype in tropical regionssuggest different scenarios for the
potential impact of Africanized honey bees in North America. One
scenario resulting from reproductive isolation would limit Africanized
honey bees in their northern movement because of their inability to
overwinter (Taylor 1985; Taylor and Spivak 1984). This is based on the
observation that the parental African population as well as Africanized
bees do not have the thermoregulatory capabilities to survive the cold


TABLE 4-6. Total development time (in days from oviposition
to adult emergence) of Africanized queens in
Africanized and European cell-producing colonies
median, (sample size).
NURSE BEE GENOTYPE3 AFRICANIZED EGG GENOTYPE (A26)
AFRICANIZED5
A43
14.4
(10)
A
A37
14.6
(6)
B
A43 & A37
14.4
(16)
C
EUROPEAN
HI
14.4
(16)
D
IBR
14.2
(14)
E
HI & IBR
14.4
(30)
F
ANALYSES0
A x D NS
A x E NS
B x D NS
B x E NS
C x F NS
3Queen-cell-producing colonies.
^Africanized comb cell size.
Kolmogorov-Smirnov one-tailed test, chi-square
distribution, df = 2, alpha = 0.05.


148
Venezuela also reduce the size of their nest entrances to a greater
extent than European honey bees, which helps to protect against
invertebrate predators, particularly ants (Bolten, personal
observation).
Besides colony defense, another response to predation is to abscond
(relocate). Because predation by both vertebrates and invertebrates and
infestation by wax moths (Galleria me!lone!1 a and Achroia arise!la) on
honey bee colonies is extensive in tropical regions, disturbance-induced
absconding would be an advantageous behavior and is frequently observed
in tropical honey bees (Fletcher 1976; Seeley 1983; Seeley, Seeley and
Akratanakul 1982; Winston, Taylor and Otis 1983). Disturbance-induced
absconding was more frequently observed in Africanized honey bees than
in European honey bees under similar conditions in Venezuela-,
particularly with respect to attacks by ants (Bolten, personal
observation).
In addition to predation on the colony-level, Africanized worker
bees may have behaviors that are better adapted to avoiding predators
and parasites while foraging. The rapid, zig-zag flight of worker bees
in the African parental population may be more advantageous in avoiding
predators (invertebrate as well as vertebrate) compared with the slower,
less erratic flight of European honey bees (Fletcher 1977b). Also,
queen honey bees on mating flights are susceptible to predators.
Fletcher (1977b) suggests that queens from the African parental
population have shorter mating flights than European queens which may
reduce predation. The differences in flight patterns and behaviors of
Africanized and European honey bees need to be investigated under
similar conditions.


123
taken into areas with only European honey bees so the reciprocal cross
was not evaluated. However, evaluating the success of the European
queen x Africanized drone cross is important because it represents the
most probable initial hybridization that will occur when Africanized
honey bees invade North America (Mexico and U.S.A.).
Methods
The Africanized queen mother (A26) was removed from a feral colony
in eastern Venezuela where there were no known European honey bees. The
colony was identified as Africanized by its behavior and small comb cell
size characteristic of the Africanized population (4.5-5.0 mm between
opposite sides of the hexagon, Chapter III). The European queen mother
(L13) was produced by a commercial queen producer in the southeastern
U.S.A. and shipped to Venezuela.
Experimental queens were produced from each queen mother by the
standard queen rearing technique of transferring (grafting) 12 to 18-
hour-old larvae into artificial queen cells that were then introduced
into queen-cell producing colonies (Laidlaw 1979). After the queen
cells were sealed, each cell was protected by a wire mesh cylinder (mesh
size = 3.0 mm). Virgin queens were allowed to emerge in the cell-
producing colonies. Newly emerged virgins were marked for
identification and then stored in a strong, queenless colony. The
following day they were introduced into individual, five-frame colonies
with Africanized worker bees from which natural matings could occur.
Queens were released into these mating colonies using standard three-
hole mailing cages (Laidlaw 1979). Based on earlier calculations,
natural release from the mailing cages was estimated to take 2.5-3.0
days. Marked, virgin queens were introduced into mating colonies rather


TABLE 4-1. Experimental matrix for the comparison of total
development times (oviposition to adult emergence)
for both Africanized and European honey bee queens.
EGG GENOTYPES
NURSE BEE GENOTYPE3 AFRICANIZED (A26) EUROPEAN (Y5)
AFRICANIZED
A43 A
A37 C
COMBINED E
EUROPEAN
19 G
27 I
28 K
F M
H 0
IBR Q
COMBINED S
B
D
F
H
J
L
N
P
R
T
COMBINED AFRICANIZED
AND EUROPEAN U V
aQueen-cell-producing colony.


APPENDIX C
CHANGES IN QUEEN PUPAL WEIGHT WITH AGE


132
TABLE 7-1. Mating success of Africanized and European honey bee queens.
TIME TO
NO. SPERM
OVIPOSITION3
(x 106)b
CORRELATIONS'
AFRICANIZED
11
4.09 + 0.50
NS
GENOTYPE (A26)
(8)
(8)
EUROPEAN
10
4.12 + 0.58
NS
GENOTYPE (L13)
(11)
(11)
ANALYSES
P<0.001d
NSe
Median days post-emergence to initiation of oviposition (sample size).
One-day-old virgins were introduced into mating colonies.
Mean + SD (sample size) of spermathecal spermatozoa number.
^Spearmans rank correlation coefficient, two-tailed, alpha = 0.05.
Kolmogorov-Smirnov two-tailed test, chi-square distribution, df = 2.
et-test, two-tailed, alpha = 0.05.


TABLE 4-2. Total development times (in days from
oviposit ion to adult emergence) for
Africanized and European honey bee queens:
median (sample size).
EGG GENOTYPES
NURSE BEE GENOTYPE3 AFRICANIZED (A26) EUROPEAN (Y5)
AFRICANIZED
A43
14.0
(1)
15.0
(4)
A37
14.5
(7)
15.0
(4)
COMBINED
14.5
(8)
15.0
(8)
EUROPEAN
19
14.5
(15)
15.0
(19)
27
14.5
(17)
15.0
(13)
28
14.0
(13)
14.5
(12)
F
14.0
(10)
14.5
(9)
H
14.0
(7)
14.5
(8)
IBR
14.0
(8)
14.8
(8)
COMBINED
14.2
(70)
15.0
(69)
COMBINED AFRICANIZED
AND EUROPEAN
14.5
(78)
15.0
(77)
aQueen-cell-producing colony.


ACKNOWLEDGEMENTS
I would like to thank the members of my committee: Dr. Thomas C.
Emmel, my chairman, for his continued support, guidance and
encouragement throughout my graduate education; Dr. Malcolm T. Sanford
for stimulating discussions and his thorough editing; and Dr. Jonathan
Reiskind for his enthusiasm and helpful suggestions. I appreciate the
comments made by Drs. James Nation and Frank Nordlie on the
dissertation. I am also grateful to Professor Frank Robinson for
introducing me to the excitement and challenges of honey bee research
and management. Drs. John Harbo, Anita Collins and Tom Rinderer were
excellent field companions, sharing their knowledge of honey bee
research techniques, and creating a stimulating research environment,
both in Venezuela and during my work in Baton Rouge. I particularly
want to thank Dr. John Harbo for the instrumental inseminations and
acknowledge his collaboration on both the bee size and egg laying rate
experiments. I would also like to thank Dr. Orley Taylor for giving me
the opportunity to study Africanized honey bees.
This research was supported by the U.S. Department of Agriculture
Cooperative Agreement No. 58-7B30-8-7 with the University of Kansas (0.
R. Taylor, principal investigator). The Ministerio de Agricultura y
Cria de Venezuela provided research facilities near Maturin. I would
like to thank Med. Vet. Ricardo Gomez Rodriguez for his hospitality and


177
Roubik, D.W. 1982. Ecological Impact of Africanized honeybees on
native neotropical pollinators. Pages 233-247 in P. Jaisson (ed.),
Social insects in the tropics. Universite Paris-Nord, Paris.
Roubik D.W. 1983. Experimental community studies: Time-series tests
of competition between African and neotropical bees. Ecology
64:971-978.
Roubik, D.W., and S.L. Buchmann. 1984. Nectar selection by Mel 1ipona
and Apis me!1ifera (Hymenoptera: Apidae) and the ecology of nectar
intake by bee colonies in a tropical forest. Oecologia 61:1-10.
Ruttner, F. 1968. Les races dabeilles. Pages 27-44 in R. Chauvin
(ed.) Traite de biologie de labeilles, 1. Masson et Cie Paris.
Ruttner, F. 1975. Races of bees. Pages 19-38 in Dadant and Sons
(eds.) The hive and the honey bee. Dadant and Sons, Inc.
Hamilton, Illinois.
Ruttner, F. 1976a. The races of bees of Africa. Proc. 25th Int.
Beekeap. Congr. 1975, Grenoble. Apimondia, Bucharest, Rumania.
Ruttner, F. 1976b. Honeybees of the tropics: Their variety and
characteristics of importance for apiculture. Pages 41-46 in E.
Crane (ed.), Apiculture in tropical climates. International Bee
Research Assoc. London.
Ruttner, F. 1981. On the taxonomy of honey bees of tropical Africa.
Pages 278-287 in Proc. 28th Int. Congr. Apimondia, Acapulco,
Mexico.
Seeley, T.D. 1977. Measurement of nest cavity volume by the honey bee
(Apis mellifera). Behav. Ecol. Sociobiol. 2:201-227.
Seeley, T.D. 1978. Life history strategy of the honey bee, Apis
mellifera. Oecologia 32:109-118.
Seeley, T.D. 1979. Queen substance dispersal by messenger workers in
honeybee colonies. Behav. Ecol. Sociobiol. 5:391-415.
Seeley, T.D. 1982. Adaptive significance of the age polyethism
schedule in honeybee colonies. Behav. Ecol. Sociobiol. 11:287-293.
Seeley, T.D. 1983. The ecology of temperate and tropical honeybee
societies. Am. Sci. 71:264-272.
Seeley, T.D., and R.D. Fell. 1981. Queen substance production in honey
bee (Apis me!1ifera) colonies preparing to swarm (Hymenoptera:
Apidae). J. Kans. Ent. Soc. 54:192-196.
Seeley, T.D. and R.A. Morse. 1976. The nest of the honey bee (Apis
mellifera L.). Insectes Sociaux 23:495-512.


2
1 iaustica is native to the Italian peninsula (Ruttner 1975). Because
these European honey bee populations were not very successful in
tropical and subtropical habitats of Brazil (Michener 1972) researchers
believed that they could improve Brazil's honey production by breeding a
honey bee better adapted to local conditions (Woyke 1969). With this
intention, honey bee queens from South Africa (A. m. scute!lata,
formerly classified as adansonii, see Ruttner 1976a, 1976b, 1981) were
imported into southeastern Brazil in 1956 (Kerr 1967). The following
year, swarms escaped and hybridized with established European honey
bees. The descendents from this hybridization are known as Africanized
honey bees (Goncalves 1982). Details of the introduction and subsequent
spread throughout South America have been extensively reviewed
(Goncalves 1974, 1975, 1982; Kerr 1967; Michener 1972, 1975; Taylor
1977, 1985; Taylor and Williamson 1975; Woyke 1969).
In the 30 years since African honey bees were imported into
southeastern Brazil, their hybridized offspring have rapidly dispersed
throughout tropical South and Central America and are now as far north
as Honduras and El Salvador (Rinderer 1986). The dispersion from their
original importation site into new areas has been rapid200-500 km per
year (Taylor 1977, 1985; Winston 1979a). As Africanized honey bees have
colonized new areas, they have achieved dramatic population densities
(Michener 1975). There are now probably more than ten million feral
colonies in South and Central America (Winston, Taylor and Otis 1983).
Their success in these new habitats, compared with the lack of success
of European honey bee populations, may be attributed to their foraging
behavior which is more suited to the resource patterns of the tropics
(Nunez 1973, 1979a, 1982; Rinderer, Bolten, Collins and Harbo 1984;


137
regions? Finally, were the relatively high reproductive rates observed
for Africanized honey bees in these studies (Otis 1980, 1982a) simply an
artifact of experimental conditions, particularly with respect to brood-
nest crowding?
Brood-nest crowding is a primary stimulus for reproductive swarming
in honey bees (Baird and Seeley 1983; Simpson 1966, 1973; Simpson and
Riedel 1963). However, the experimental conditions affecting brood-nest
crowding for Africanized colonies in South America were significantly
different from the experimental conditions for European colonies in
North America: the nest cavity volume for Africanized colonies was 22
liters (Otis 1980; Winston 1979b) compared with 42 liters for European
colonies in North America (Winston 1980a). Despite these problems with
respect to making valid comparisons, the earlier studies (particularly
Winston 1979b) leave one with the impression that the apparent
differences in reproductive rates between the two populations were
primarily due to differences in demographic parameters and not to
differences in environmental and experimental conditions, resource
utilization, or some combination of factors.
What is the consequence of comparing reproductive rates of
Africanized honey bees in South America with those of European honey
bees in North America without considering differences in environmental
conditions? Certainly, environmental conditions in temperate regions
impose strict limits on the length of the reproductive (= swarming)
season for honey bees because of a reduced growing season when floral
resources (nectar and pollen) are available. The honey bee reproductive
season is significantly shorter than the growing season, because
colonies first have to grow to reproductive size before swarming can


TABLE B-2
160
TABLE B-2.
Africanized
mean + SD,
and European adult honey bee weights (mg):
coefficient of variation (CV), (sample size).
GENOTYPE
COMB CELL
TYPE3
FRESHLY
EMERGED
DRIEDb
CORRELATIONS'
AFRICANIZED
A26
EUR
94.9 + 5.1
CV = 5.4
(28)
13.5 + 0.6
CV = 4.4
(28)
#**
A57
EUR
88.6 + 6.2
CV = 7.0
(30)
11.9 + 0.8
CV = 6.7
(30)
***
B39
EUR
87.4 + 4.7
CV = 5.4
(16)
12.3 + 0.5
CV = 4.1
(16)
NS
A60
AFR
95.2 +5.1
CV = 5.4
(30)
13.2 + 0.6
CV = 4.5
(30)
***
EUROPEAN
WE2
EUR
107.1 + 4.8
CV = 4.5
(29)
14.6 + 0.6
CV = 4.1
(29)
**
Y (K)
EUR
116.1 + 4.7
CV = 4.0
(27)
15.5 + 0.7
CV = 4.5
(27)
***
aEUR = European comb cell diameter = 5.4 mm.
AFR = Africanized comb cell diameter = 4.8 mm.
Dried at 50C for 48 hrs.
Pearson's correlation coefficient alpha = 0.05;
** = PcO.Ol; *** = P<0.001.


134
The selection advantage for Africanized bees may be a result of
behavioral and/or physiological characteristics that may include
differences in resource utilization and/or colony demography. It is not
surprising that Africanized honey bees are better adapted to tropical
conditions than are European honey bees, considering the former are
derived from imported African bees that evolved under similar tropical
and sub-tropical conditions in Africa. Fletcher (1978) has reviewed the
biological characteristics of the parental population of African honey
bees in Africa.
The spread and impact of Africanized honey bees in South America
must, however, be kept in perspective. European honey bees introduced
into North America early in the 17th century (Pellett 1938) dispersed
throughout North America, also achieving high population densities. In
general, honey bees are very successful not only in their native
habitats but in almost every region where they have been introduced.
Their success is based on a highly developed social system that allows
honey bees to: 1) develop large, perennial colonies that are able to
buffer climatic changes; 2) efficiently utilize resources because of
advanced communication and recruitment systems; and 3) defend against
both vertebrate and invertebrate predators because of their very
effective colony defense behavior.
The question with which we are concerned in these studies is not
what makes A. me!1 ifera more successful than other species nor what
impact introduced honey bees have on native pollinator communities (see
Roubik 1978, 1979, 1980, 1982, 1983; Roubik and Buchmann 1984). Nor is
it a question of comparing Africanized honey bees in tropical regions
with European honey bees in temperate regions (see Winston, Dropkin and


36
of age-related tasks. However, the sequence and duration of the
different stages are flexible and depend on the needs of the colony.
An advantage of worker size variation within bumble bee colonies
may be efficient utilization of diverse nectar and pollen resources that
may be size dependent. Different sized workers within a colony
specialize on those resources that they can most efficiently exploit
(Heinrich 1979a). However, highly eusocial bees are not at a
disadvantage with respect to resource utilization because they have
evolved complex communication systems that allow foragers to monitor
changing nectar conditions and to recruit workers from the colony to a
particular resource. Therefore, both the species characterized by
workers of highly variable sizes and those species characterized by
uniformly-sized workers have evolved behaviors that enhance the
efficiency of nectar and pollen exploitation.
The difference in intra-colony worker size variation between
primitively eusocial and highly eusocial species of bees is so
significant that Kerr and Hebling postulated that "some controlling
mechanism leads to reduced variances among mature workers [Meliponinae
and Apis], which are therefore of relatively uniform size" (1964, p.
267). Waddington (1981) hypothesized that the evolution and maintenance
of the complex communication systems in Apis, Triaona and Mel ipona
depend upon uniformity of worker bee size within a colony. Differences
in bee size may result in miscommunication because resource
"profitability" may be size dependent. For example, a high quality
resource for a small bee may not be a high quality resource for a larger
bee. However, a regulatory mechanism for reduced size variation has not
been identified.


157
TABLE A-2. Mortality during different developmental stages. Mortality
was measured in European comb cell size with European nurse
bees.
El3 E2b L1C L2d Sb8 Nf
AFRICANIZED EGG GENOTYPES
30
28
29
30
30
30
fMortality during first 24 hours in test colony (acceptance).
^Mortality between 24-72 hours (before hatching).
'-Mortality between 72-96 hours (at time of hatching).
Mortality during older larval stages, before sealing.
^Mortality during the pupal stage.
fN = total eggs monitored.
9Not distinguished between and L2.
A53
A26
A25
EUROPEAN EGG GENOTYPES
W18
HI
Y(A5)
0
0
1
0
0
0
0 5
0
0 9
0 0
0 3
0 4
0 0
199 o
0 0
2 0
8 0
0 0


29
TABLE 2-4. Unsealed brood development times. Hypotheses were tested
using Kolmogorov-Smirnov one-tailed test chi-square
distribution, df = 2, alpha = 0.05 (Siegel 1956).
HI: Worker bee development is faster for Africanized genotypes than for
European genotypes.
A X B ***a
C X D **
E X F ***
G X H NS
A X H NS
AC X BD ***
EG X FH ***
AE X BF ***
CG X DH ***
ACEG X BDFH ***
H2: Worker bee development is more rapid in Africanized comb cells than
in European comb cells.
AXE NS
C X G NS
B X F NS
D X H NS
AC X EG NS
BD X FH NS
H3: Worker bee development is more rapid with Africanized nurse bees
than with European nurse bees.
A X C NS
E X G NS
B X D NS
F X H NS
AE X CG NS
BF X DH NS
H4: Worker bee development is more rapid with Africanized comb cells
and Africanized nurse bees than with European comb cells and
European nurse bees.
A X G NS
B X H NS
a ** = P<0.01
*** = Pco.001.
bAnalysis may be NS because test used is conservative for small sample
sizes using chi-square distribution.


127
as well as Africanized bees may not have the thermoregulatory
capabilities to survive the cold temperatures of temperate winters
(Nunez 1979b; Woyke 1973). Based on the temperature limits of the
parental population, Taylor (1985) and Taylor and Spivak (1984)
predicted the northern limits of Africanized honey bees in North
America. However, Africanized honey bees may acquire, through
hybridization with European honey bees in Mexico and southern U.S.A.,
the ability to overwinter farther north than is presently expected.
That is, the overwintering genome of the European honey bees may become
introgressed into the Africanized genome. Or, the corollary, the
stinging behavior characteristic of the Africanized honey bees may
become introgressed into the overwintering European population.
Successful genetic introgression of these traits may not be a rapid
process because these traits are polygenic and/or may involve coadapted
genomes. However, because hybridization occurs, the potential for
successful genetic introgression exists and must be considered.
Hybridization may, therefore, result in the stinging behavior of the
Africanized honey bees becoming a potential public health hazard
throughout North America, not just in the warmer southern regions. That
this may be the unfortunate outcome of hybridization is supported by
recent investigations in Argentina, which have demonstrated that
Africanized honey bees are distributed farther south than predicted
based on temperature limits of the parental population (Dietz, Krell and
Eischen 1985; Krell, Dietz and Eischen 1985).
The U.S. Department of Agriculture Economic Research Service has
recently evaluated the potential impact of Africanized honey bees in the
U.S.A. (McDowell 1984). Unfortunately this report does not consider the


74
TABLE 4-5. Analyses of queen development times for the
European egg genotype in the different cell-
producing colonies. Letters represent different
cell-producing colonies; see Table 4-1 for
explanation. Kolmogorov-Smirnov two-tailed test
chi-square distribution df = 2 alpha = 0.05
(Siegel 1956).
B x D
NS
B x H
NS
B x J
NS
B x L
NS
B x N
NS
B x P
NS
B x R
NS
D x H
NS
D x J
NS
D x L
NS
D x N
NS
D x P
NS
D x R
NS
H x J
NS
H x L
NS
H x N
NS
HxP
NS
H x R
NS
J x L
NS
J x N
NS
J x P
NS
J x R
NS
L x N
NS
L x P
NS
L x R
NS
N x P
NS
N x R
NS
P x R
NS


95
A third variable, comb cell size, had to be controlled. If bees
were allowed to build their own comb, comb built by Africanized workers
would be smaller than comb built by European bees (Chapter III). The
larger, European comb was selected for these experiments for two
reasons: 1) European queens do not lay eggs in a uniform pattern in
Africanized comb; and 2) there is a higher brood mortality in colonies
with European nurse bees on Africanized combpossibly because the
larger European nurse bees have difficulty feeding the developing larvae
in the smaller, Africanized cells (Chapters II and III). On the other
hand, Africanized queens and worker bees appear to behave normally when
managed on European comb. Only Africanized bees that had been reared in
managed colonies with European combs were used in the experimental
colonies to avoid any delay in adjusting to larger comb cell size.
Finally, the fourth variable controlled was resource availability.
Africanized and European queens were compared simultaneously so that
floral resource conditions were identical. Surplus honey was also
provided for each experimental colony to reduce the effects of
differential foraging success between Africanized and European honey
bees in tropical resource conditions (Rinderer, Bolten, Collins and
Harbo 1984; Rinderer, Collins and Tucker 1985).
In addition to colony-level variables, conditions under which
experimental queens are produced may affect queen fecundity, e.g., age
of larvae used to produce queens, quantity and quality of food fed to
developing queen larvae, and temperature during development (Beetsma
1979; Laidlaw 1979; Weiss 1974; Woyke 1971). The age of a queen may
also affect her fecundity (Ribbands 1953). In this study, all


LITERATURE CITED
Abdellatif, M.A. 1965. Comb cell size and Its effect on the body
weight of the worker bee Apis mel11fera L. Am. Bee J. 105:86-87.
Adams, J. E.D. Rothman, W.E. Kerr and Z.L. Paulino. 1977. Estimation
of the number of sex alleles and queen matings from diploid male
frequencies in a population of Apis mel1ifera. Genetics 86:
583-596.
Alies, P. 1961. Role of queen and worker bees 1n the hereditary
transmission of certain characteristics. Apic. Absts. 12:167.
Alpatov, W.W. 1929. Biometrical studies on variation and races of the
honey bee. Quart. Rev. Biol. 4:1-58.
Alpatov, W.W. 1933. South African bees biometrically investigated.
Bee World 14:62-64.
Anderson, R.H., B. Buys and M.F. Johannsmeier. No date. Beekeeping in
South Africa. Technical Services Bulletin 394. South African
Dept, of Agriculture, Pretoria.
Baird, D.H., and T.D. Seeley. 1983. An equilibrium theory of queen
production in honeybee colonies preparing to swarm. Behav. Ecol.
Sociobiol. 13:221-228.
Baudoux, U. 1933. The influence of cell size. Bee World 14:37-41.
Beetsma, J. 1979. The process of queen-worker differentiation in the
honeybee. Bee World 60:24-39.
Boch, R. 1957. Rassenmabige Unterschiede bei den Tanzen der
Honigbiene. Z. Vergl. Physiol. 40:289-320.
Boch, R., and C.A. Jamieson. 1960. Relation of body weight to
fecundity in queen honeybees. Can. Ent. 92:700-701.
Boch R. D.A. Shearer and J.C. Young. 1975. Honeybee pheromones:
Field tests of natural and artificial queen substance. J. Chem.
Ecol. 1:133-148.
Bodenheimer, F.S. 1937. Studies in animal populations, II. Seasonal
population trends of the honey-bee. Quart. Rev. Biol. 12:406-425.
168


BIOGRAPHICAL SKETCH
Alan Bolten was born on May 27, 1945 In Newark, New Jersey. In
1959, he was graduated from Maple Avenue Grammar School. Four years
later, he completed h1s secondary education at Weequahic High School in
Newark. He received his undergraduate education from Union College in
Schenectady, New York, where he was graduated with honors in biology in
1967. Alan began graduate studies in the Department of Zoology at the
University of Florida in 1977. He is married to Karen Bjorndal.
182


80
ovarioles, more rapid egg maturation, and reduction in egg size (Hagan
1954 and Iwata and Sakagami 1966 cited in Wilson 1971; Wilson 1971).
Honey bee queens have a very large number of ovarioles (>300) and, for
European queens, the number of ovarioles has been shown to be an
inherited character (Eckert 1934) which is positively correlated with
queen pupal weight (Hoopingarner and Farrar 1959). Queen weight was
also found to be correlated with brood production (Boch and Jamieson
1960). If it is assumed that both Africanized and European honey bees
have the same relationship between queen weight and brood production,
then weights of queens from the two populations can be compared to
determine potential differences in fecundity.
In honey bees, differentiation between worker and queen castes is
not genetically determined, but rather is regulated by the quantity and
quality of food fed to developing larvae during the first 3 days
(Beetsma 1979). Therefore, a number of factors other than genotype
affect queen size, e.g., age of larvae used to produce queens,
population of the cell-producing colony, quantity and quality of food
fed to developing larvae, and temperature (Beetsma 1979; Johansson and
Johansson 1973; Laidlaw 1979; Weiss 1974; Woyke 1971). Because of
differences in queen rearing methods and experimental conditions,
previous comparisons of size between Africanized and European queens may
be inappropriate. This study was undertaken to compare queen pupal
weights for Africanized and European honey bees under identical
experimental conditions in Venezuela.
Methods
Four Africanized honey bee lines (A26, A57, A61 and A62) were
established from queens removed from feral colonies in an area in


41
and Willis 1939). This was confirmed for fresh pupal weights by
weighing a sample of pupae every 24 hours from day 11.5 post oviposition
to 17.5 days post oviposition (Table B-5). Although Africanized bees
develop one day faster than European bees (Chapter II), pupal weights
can be compared because there is no significant difference in weights
between adjacent days during this period of pupal development (Table B-
5).
Pupal weights were used instead of adult weights in order to reduce
variation resulting from differences in food engorgement and/or feces
accumulation. Pupae were carefully removed from their comb cells by
first removing the cappings and then spreading the cell walls with a
forceps in order that the pupae could easily be removed without
rupturing. Weights (to 1.0 mg) were recorded using Mettler Type H4 and
H6 balances. Comb cell diameters were determined by measuring ten
adjacent cells; three sets of measurements were made from each comb.
Results
Table 3-1 presents the experimental design matrix. The interaction
of egg genotype and comb cell size on worker bee pupal weights for each
of the nine genotypes is summarized in Table 3-2. Table 3-3 presents
the results of the statistical analyses. When Africanized and European
genotypes are reared simultaneously in the same colony (same comb cell
size, nurse bee genotype, temperature and humidity, and colony size),
the weights of the worker bees produced are different. Africanized bee
pupae (111.1 +7.6 mg) that developed in Africanized comb cells were
smaller than European bee pupae (123.3 +6.3 mg) that also developed in
Africanized comb cells (ACE x MOO, P<0.001). When worker bees of
European genotypes are reared in Africanized comb cells, the cells are


128
possibility that further hybridization between Africanized and European
honey bees might result in the stinging behavior of Africanized honey
bees becoming established throughout the northern regions of the U.S.A.
The economic consequences as well as public health hazards may be more
widespread throughout North America than previously thought. For any
solutions to the Africanized honey bee problem to be successful a
realistic assessment of the potential problem is necessary.
However, hybridization may also have a positive impact. Coupled
with selection favoring both the foraging and thermoregulatory behavior
of European honey bees in temperate regions, hybridization between
Africanized and European bees may have the positive effect of increasing
the rate at which African genes become rare in the population. There is
a large population of European honey bees, both managed and feral, in
North America (perhaps greater than 15 million colonies in the U.S.A.
alone), which is particularly dense in the south where the Africanized
bees will first enter the U.S.A. The invading Africanized population
would be quite small relative to the existing European population,
increasing the frequency of hybridization and resultant "swamping" (or
diluting) of African genes.
The problem of Africanized honey bees may be reduced prior to their
entry into the U.S.A. because of both the potential for hybridization
and competition for available floral resources with European honey bees
in Mexico. Mexico has a larger population of European honey bees than
any other country in Latin America. When Africanized honey bees enter
Mexico, they will be entering a region that already has an extensive,
established population of European honey bees, both managed and feral
[2.6 million managed colonies alone (Zozaya cited in Taylor 1985)].


21
large as expected from previous reports, which underscores the
importance of making comparisons under identical conditions.
The proportional difference (5.7%) in egg development times between
Africanized (A26) and European (Y5) honey bees reported by Harbo,
Bolten, Rinderer and Collins (1981) is identical to the proportional
difference in total development time reported in the present study
(5.7%, see Table 2-8). Egg development time is a function of the
inherent rate characteristic of the particular genotype because colony-
level parameters (e.g., feeding) are not involved (Harbo, Bolten,
Rinderer and Collins 1981). Using egg development to evaluate
differences in total development between genotypes (or populations) is
advantageous because egg development times are easier to evaluate, take
less time, have fewer variables to control (temperature and humidity
only), and can be evaluated in an incubator rather than in a colony,
avoiding problems associated with disturbing the colony during
observations. It must be noted, however, that by using egg development
times one can only extrapolate proportional differences between
genotypes for total development time but cannot extrapolate the absolute
total development time.
A prerequisite for high reproductive rates would be a rapid colony
growth rate. However, the importance of worker development time to the
rate of colony growth (increase in numbers of bees in a colony) has
apparently been misunderstood, e.g., see Fletcher (1977a, 1978),
Fletcher and Tribe (1977a), Tribe and Fletcher (1977), Winston (1979b),
Winston, Dropkin and Taylor (1981), Winston and Katz (1982), Winston,
Taylor and Otis (1983). The difference in worker development times
observed for Africanized and European honey bees is not a factor


Abstract of Dissertation Presented to the Graduate School
of the University of Florida in Partial Fulfillment of the
Requirements for the Degree of Doctor of Philosophy
BIOLOGY OF AFRICANIZED AND EUROPEAN HONEY BEES,
Apis me!1 ifera, IN VENEZUELA
By
Alan B. Bolten
August 1986
Chairmanj Thomas C. Emmel
Major Department: Zoology
To determine factors responsible for the greater success of
Africanized honey bees, Apis mel1 ifera, in tropical regions of South
America, demographic parameters affecting colony reproductive rates were
evaluated for Africanized and European honey bees under identical
conditions in Venezuela. Worker bee development time was evaluated as
an interaction between egg genotype, comb cell size and nurse bee
genotype. Africanized worker bees developed faster than European bees:
18.9 and 19.8 days, respectively. There was no significant effect of
comb cell size or nurse bee genotype. Mortality for different
developmental stages was recorded. The relationship of worker bee
development time to colony growth rate is discussed.
Africanized queens develop in 14.5 days post-oviposit ion compared
with 15.0 days for European queens. Queen pupal weights were not
significantly different. Post-emergence maturation rates for
Africanized and European queens were similiar as determined by both the


CHAPTER II
WORKER BEE DEVELOPMENT TIMES
Introduction
The presence of both Africanized and European honey bees, Apis
mel1 ifera, in South America allows for comparisons to be made under
identical conditions between a population that has evolved in the
tropics and one that has evolved in temperate regions. Africanized
honey bee populations in South America are reported to have a high
colony reproductive (swarming) rate compared with European honey bee
populations in North America (Otis 1980, 1982a; Seeley 1978; Winston
1979b, 1980a; Winston, Dropkin and Taylor 1981; Winston, Taylor and Otis
1983). However, these investigations have not been conducted under
similar environmental or experimental conditions. Therefore,
comparisons of reproductive rates between Africanized and European honey
bees using these data are inappropriate either for identifying
differences in reproductive rates between tropical and temperate honey
bee populations or for identifying factors responsible for the success
of Africanized bees in tropical regions.
Reproductive rates in honey bees are a function of colony growth
rates which are a result of an interaction of at least three factors:
resource availability, resource utilization efficiency (foraging
success, brood production efficiency, and bee size), and colony
demographic parameters. Worker bee longevity is the only demographic
9


3
Rinderer, Collins and Tucker 1985; Winston and Katz 1982). As a result
of both foraging success and the length of time throughout the year that
resources are available in the tropics, Africanized honey bees have a
high annual reproductive rate, which is responsible for both their rate
of dispersal into new areas and their high colony densities. Net
reproductive rates for Africanized bees are estimated to be 16 colonies
per colony per year based on demographic data collected in French Guiana
(Otis 1980, 1982a), compared with 0.92-0.96 (Seeley 1978) or, when
afterswarms are considered, 3-3.6 (Winston 1980a; Winston, Taylor and
Otis 1983) colonies per colony per year for European honey bees in North
America.
Characteristics of Africanized Honey Bees
The most well known characteristic that differentiates Africanized
honey bees from European honey bees is their stinging behavior (Collins,
Rinderer, Harbo and Bolten 1982; Stort 1974, 1975a, 1975b, 1975c, 1976).
Because of their stinging behavior, Africanized bees are a health hazard
for both humans and domestic animals (Taylor 1986). Collins, Rinderer,
Harbo and Bolten (1982) compared the colony defense behavior of
Africanized honey bees in Venezuela with European bees under identical
conditions in Venezuela and with a population of European bees in
Louisiana, U.S.A. Africanized honey bees responded more rapidly and in
much greater numbers, resulting in 5.9 times as many stings in a target
compared with European honey bees in Venezuela and 8.2 times as many
stings compared with European bees in Louisiana. Two additional
components of Africanized honey bee defense behavior increase their
potential as a health hazard. Compared to European bees, Africanized
bees pursue a source of disturbance for a greater distance (160 versus


58
Eggs of known ages were collected from the Africanized (A26) and
European (Y5) egg sources by the standard commercial queen-producing
technique of caging the queen on an empty comb within a colony (Harp
1973; Laidlaw 1979; see also Chapter II). After 6 hours, the combs with
the egg samples were moved to strong incubator-colonies for the eggs to
develop and larvae to hatch and be fed. Very young larvae, 12-18 hours
old, were transferred (grafted) into beeswax queen-cell cups primed with
royal jelly and then introduced into queen-cell-producing colonies
(=nurse bee colonies) (Laidlaw 1979). All cell-producing colonies had
large worker bee populations and were intentionally crowded into two
standard Langstroth hive bodies. Queens in the cell-producing colonies
were removed 48 hours before introducing the grafted cells. All young,
unsealed brood was also removed 2-4 hours before introducing the grafted
cells. Twenty grafted Africanized and twenty grafted European cells
were introduced into each cell-producing colony. There were twenty cell
cups to a frame, ten on the top bar and ten on the middle bar. Both
Africanized and European larvae were grafted into the same frame, five
each on the top bar and five each on the middle bar. All cell cups were
equally spaced about 8 mm apart, centered on the bars.
In one experimental trial, the effect of nurse bee genotypes on
queen development was evaluated by comparing queen development times for
both Africanized and European egg genotypes in both Africanized and
European cell-producing colonies. In another trial, development times
for Africanized queens in Africanized and European cell-producing
colonies were compared. In both trials, the Africanized cell-producing
colonies had comb cell sizes characteristic of Africanized honey bees
(4.8 mm wide; Chapter III).


43
of genetically similar, reciprocal hybrids are not the same. Because
reciprocal hybrids are phenotypically different from each other, but
phenotypically similar to their maternal lines, maternal genotype must
interact with cell size and egg genotype to determine pupal weight.
This is the first character in honey bees that has been shown to be
influenced by maternal inheritance. Other genetic mechanisms cannot
explain these results. The mechanism for maternal inheritance in worker
size may be through egg size, which has been shown to be inherited
(Roberts and Taber 1965; Taber and Roberts 1963).
Alies (1961) and Mel'nichenko (1962) suggested that differences
between nurse bee genotypes might affect size of developing larvae.
However, McGregor (1938) found that bee size was not affected by nurse
bee genotype. In the experiments presented in this chapter, pupae from
each of the genotypes were reared simultaneously in the same colony for
each comb size treatment. Therefore, differences between the pupal
weights of Africanized and European genotypes cannot be attributed to
either nurse bee differences, cell size, or temperature but must be a
result of both egg and maternal genotype differences.
The importance of maternal inheritance on bee size is that it
reduces worker bee size variation within a colony. If maternal
inheritance were not operating, worker bees of different sizes would be
produced within a colony because of cell size differences and genotype
differences. The effectiveness of maternal inheritance for reducing bee
size variation can be demonstrated by comparing the degree of variation
for the two parameters of bee size (comb cell volume and genotype) with
the degree of worker bee size variation. Abdel 1 at if (1965) showed that


112
TABLE 6-7. Comparison of daily egg laying rates for sister
Africanized and European nurse bees. Mean + SD
size).
queens with
(n = sample
AFRICANIZED
NURSE BEES
EUROPEAN
NURSE BEES
ANALYSES3
AFRICANIZED QUEENS
A57 (W41)
A57 (W42)
694.0 +39.1 (4)
699.8 + 39.5 (4)
NS
A62 (W85)
A62 (W81)
798.7 + 92.3 (3)
915.8 +30.7 (4)
NS
EUROPEAN QUEENS
GK30 (Y61)
GK30 (Y63)
823.5 +37.0 (4)
736.8 + 32.2 (4)
NS
^lann-Whitney U test, two-tailed, alpha = 0.05.


110
TABLE 6-5. Analyses of the effect of queen genotypes and nurse bee
genotypes on the daily egg laying rates of Africanized and
European honey bees. Letters represent the different
treatments presented in Table 6-4. Mann-Whitney U test,
two-tailed, alpha = 0.05 (Siegel 1956).
HI: There is no difference in egg laying rates between Africanized
and European queens.
i.Africanized nurse bees
A x B NS
ii.European nurse bees
D x E NS
iii.Africanized and European
nurse bees combined
(A + D) x (B + E) NS
H2: There is no differential effect of Africanized and European
worker bees on egg laying rates.
i.Africanized queens
A x D NS
ii.European queens
B x E NS
iii.Africanized and European
queens combined
C x F
NS


63
Res u1ts
Queen Development Times
Table 4-1 presents the experimental matrix for evaluating the
interaction of egg genotype and nurse bee genotype on queen development
times for Africanized and European queens. Table 4-2 presents the total
development times from oviposition to adult emergence for Africanized
queens and European queens in Africanized and European cell-producing
colonies. Table 4-3 presents the analyses for the paired comparisons in
each cell-producing colony. These paired comparisons avoid any
differences between cell-producing colonies because colony size (nurse
bee population), brood area temperature, and quantity and quality of
larval food are factors that affect queen development (Beetsma 1979;
Laidlaw 1979; Johansson and Johansson 1973). Africanized queens develop
in 14.5 days post-oviposition compared with 15.0 days for European
queens (P<0.001, Kolmogorov-Smirnov one-tailed test, chi-square
distribution, df = 2; Siegel 1956). There was no significant effect of
the cell-producing colony on queen development times (Kolmogorov-Smirnov
two-tailed test, chi-square distribution, df = 2, alpha = 0.05) (Tables
4-4 and 4-5).
Table 4-6 presents the development times for the Africanized queens
in Africanized and European cell-producing colonies. There was no
difference in Africanized queen development time between Africanized and
European cell-producing colonies (Kolmogorov-Smirnov one-tailed test,
chi-square distribution, df = 2, alpha = 0.05).
The median unsealed development times from oviposition to sealing
for both the Africanized and European queens was 7.5 days (Table 4-7).
However, the Africanized and European genotypes were significantly


147
mutually exclusive (Winston, Otis and Taylor 1979). Fletcher suggests
that resource-induced absconding may not always be appropriate,
considering
the distance that bees can fly in relation to the general
distribution of their food plants. The maximum flight range
is unlikely to exceed about 16 kilometres...and yet huge areas
of Africa inhabited by honey-bees consist of more or less
uniform grasslands and savannah. With certain exceptions,
therefore, such as movements up and down mountain slopes and
in and out of river valleys, there would appear to be little
advantage in absconding in such areas, for within their flight
range the bees would very often find only more of the same
type of country they had left. (Fletcher 1975, p. 13).
However, based on measurements of engorgement and estimates of metabolic
rates, the maximum flight range of absconding colonies of honey bees has
been calculated to be as great as 131 km (Otis, Winston and Taylor
1981). In addition, periodic foraging while in-transit could extend the
potential distance even further. Until comparative studies demonstrate
the advantages of either resource-induced absconding or hoarding,
resource-induced absconding behavior, which is common in tropical honey
bee populations, is concluded to be an advantageous behavior under some
tropical conditions.
Predation and Colony Defense
Ability to defend the nest from predators affects colony survival
and therefore reproduction. The colony defense behavior characteristic
of Africanized honey bees (Collins, Rinderer, Harbo and Bolten 1982;
Stort 1974, 1975a, 1975b, 1975c, 1976) may be more effective than that
of European honey bees. One aspect of colony defense behavior of
Africanized honey bees, their stinging behavior, is so extreme that
Africanized bees are a public health hazard for both humans and domestic
animals (Taylor 1986). This colony defense behavior is particularly
effective against vertebrate predators. Africanized honey bees in


102
better with Africanized bees and one sister of Africanized queen pair
(A57) performed better with European bees.
Egg laying rates for queens evaluated in the daily egg laying
experiment were compared with estimated egg laying rates derived from
the brood production experiment (Table 6-12). There was no correlation
between daily egg laying rates with either the estimated daily egg
laying rates for the first 12 days or the estimated daily egg laying
rates for 17 days. Estimated daily egg laying rates at day 12 was
significantly correlated to the estimated overall egg laying rates at
day 17 (P<0.05).
Adequate pollen and nectar resources were available during the
experiment. Each colony stored pollen and had an overall weight gain.
There was no significant difference between colonies with Africanized or
European worker bees with respect to pollen stored or weight gained.
The amount of pollen stored by each colony was not significantly
correlated with colony weight gain, total brood produced, or mean
estimated daily mortality. Colony weight gain was positively correlated
(P<0.05) with total brood produced and negatively correlated (P<0.05)
with mean estimated daily mortality.
Discussion
The purpose of these experiments was to compare egg laying rates
between Africanized and European queen honey bees and to determine if
Africanized and European worker bees differentially affect brood
production and/or the queen's egg laying behavior. Results from daily
egg laying rates indicate that there was no significant difference
between Africanized and European queens during the initial colony growth
period under identical experimental conditions in Venezuela. There was


115
TABLE 6-10. Brood production during first brood cycle expressed as a
percent of adult population.
DAY 12
DAY 17
USB3 SBb TBC USB SB TB
AFRICANIZED NURSE BEES
AFRICANIZED QUEENS
A62 (W81) 92.3
A57 (W42) 40.1
EUROPEAN QUEEN
GK30 (Y63) 115.9
EUROPEAN NURSE BEES
AFRICANIZED QUEENS
A62 (W85) 71.6
A57 (W41) 103.2
EUROPEAN QUEEN
GK30 (Y61) 108.5
84.3
64.2
176.6
104.3
239.4
104.4
209.9
173.2
449.3
274.6
53.4
169.3
113.3
149.6
262.9
65.1
33.9
136.7
137.1
56.4
100.7
176.1
124.2
232.5
224.9
48.9
157.4
145.8
140.1
285.9
ANALYSES^
AFR WORKERS x EUR WORKERS NS NS NS NS NS NS
aUnsealed brood (eggs and larvae); colony adult population estimated
for Day 12 and Day 17 as described in methods.
Sealed brood (pre-pupae and pupae).
^Total brood.
Mann-Whitney U test, one-tailed, alpha = 0.05. AFR = Africanized;
EUR = European.


174
Merrill, J.H. 1924. Observations on brood-rearing. Am. Bee J. 64:
337-338.
Michener, C.D. 1964. Reproductive efficiency in relation to colony
size in hymenopterous societies. Insectes Sociaux 11:317-341.
Michener, C.D. 1972. Final report of the Committee on the African
Honey Bee. Nat. Res. Counc., Nat. Acad. Sci., Washington, D.C.
Michener, C.D. 1974. The social behavior of the bees. Belknap Press,
Cambridge, Mass.
Michener, C.D. 1975. The Brazilian bee problem. Ann. Rev. Ent.
20:399-416.
Michener, C.D. 1979. Biogeography of the bees. Ann. Missouri Bot.
Gard. 66:277-347.
Milum, V.G. 1930. Variations in time of development of the honeybee.
J. Econ. Ent. 23:441-447.
Moeller, F.E. 1958. Relation between egg-laying capacity of queen bee
and populations and honey production of their colonies. Am. Bee J.
98:401-402.
Moeller, F.E. 1961. The relationship between colony populations and
honey production as affected by honey bee stock lines. Production
Research Report 55. USDA, Washington, D.C.
Morse, R.A. 1984. The mating behavior of African queens. Glean. Bee
Cult. 112:125.
Morse, R.A., D.M. Burgett, J.T. Ambrose, W.E. Conner and R.D. Fell.
1973. Early introductions of African bees into Europe and the New
World. Bee World 54:57-60.
Nelson, J.A., and A.P. Sturtevant. 1924. The rate of growth of the
honeybee larvae. Department Bulletin 1222. USDA, Washington, D.C.
Nolan, W.J. 1925. The brood-rearing cycle of the honeybee. Department
Bulletin 1349. USDA, Washinton, D.C.
Nolan, W.J. 1928. Seasonal brood-rearing activity of the Cyprian
honeybee. J. Econ. Ent. 21:392-403.
Nunamaker, R.A., and W.T. Wilson. 1981. Comparison of MDH allozyme
patterns 1n the African honey bee (Apis mel1 ifera adanson i i L.) and
the Africanized populations of Brazil. J. Kans. Ent. Soc. 54:
704-710.
Nunez, J.A. 1973. Quantitative investigation of the behaviour of Apis
mellifera 1 iqustica Spinola and Apis mellifera adansonii Latreille:
Energy factors, forager recruitment and foraging activity. Apiacta
8:151-154.


CHAPTER I
INTRODUCTION
Evolutionary Origin and Distribution of Honey Bees
Honey bees of the genus Apis have their greatest diversity in Asia
(Michener 1979). Earliest fossil evidence for the genus is from
01igocene deposits in Europe (Zeuner and Manning 1976). The
evolutionary relationships of the four generally recognized species of
Apis are reviewed by Michener (1974). Three of the species (A. cerana.
A. dorsata and A. florea) are native only to Asia (Michener 1979;
Ruttner 1975). The western honey bee (A. me!1 ifera) is native to
Africa, western Asia and Europe and may have evolved in tropical or
subtropical Africa (Wilson 1971) or the Near East (Ruttner 1975). The
widely different climatic conditions and floral resources under which
populations of A. mel1ifera evolved have resulted in a number of
geographically recognizable subspecies (Alpatov 1929, 1933; Br. Adam
1966; Dupraw 1965; Ruttner 1968, 1975, 1976a, 1976b; Smith 1961; Wafa,
Rashad and Mazeed 1965).
Importation of African Honey Bees into Brazil and Their Dispe.rs.a_l
Throughout South and Central America
European honey bees (A. mel1ifera mel1ifera and A. m. 1igustica)
had been introduced into Brazil by 1845 (Gerstaker cited in Pellet 1938
Woyke 1969). A. m. mel1ifera is native to Europe in the regions west
and north of the Alps and extending east into Central Russia; A. m.
1


81
eastern Venezuela that had no known European honey bees. They were
identified as Africanized honey bees by their comb cell size, which was
significantly smaller than European comb cell size (Chapter III). Two
European lines (YK and WE) were established from queens shipped to
Venezuela by commericial queen producers in the southeastern U.S.A.
Three additional European lines (YD, N and GK) were established from
queens shipped to Venezuela from the U.S. Department of Agriculture Bee
Breeding and Stock Center Laboratory, Baton Rouge, Louisiana, U.S.A.
Queens were produced from these nine lines by standard queen
rearing methods (Laid!aw 1979). Egg samples from the nine queen mothers
were collected by confining the queens to an empty comb within their own
colonies using an 8 x 8 cm push-in cage made from 3 mm mesh hardware
cloth. -Queen excluder material was soldered to the tops of the push-in
cages, allowing worker bees to move in and out in order to feed and tend
the queen (Harbo, Bol ten, Rinderer and Collins 1981). Both Africanized
and European eggs were collected in European size comb. After
approximately 4-6 hours, the queens were released, and combs containing
the eggs were put into a strong colony in order for the eggs to be
incubated and for the larvae to be fed. Africanized and European eggs
were both put into the same incubator-colony in order to control for any
differences in early larval feeding and temperature.
Young larvae approximately 12-15 hours old were transferred
(grafted) into artificial, beeswax, queen-cell cups and then introduced
into the cell-producing colonies. Twenty larvae from one of the
Africanized lines and twenty larvae from one of the European lines were
grafted into each cell-producing colony. To control for extrinsic
factors affecting queen size, analyses of Africanized and European


124
than mature queen cells so that the identity of the experimental queens
would later be certain. The mating colonies were located in an area in
eastern Venezuela where there was no known European honey bees, but
which was densely populated with feral Africanized colonies.
Eighteen days after introduction into mating colonies, the queens
were collected, and the spermatazoa in their spermatheca were counted
using hemacytometers (Mackensen and Roberts 1948; Mackensen and Tucker
1970). The criterion for mating success was the number of spermatozoa
in the spermatheca. In addition, the age of the queen when oviposition
first started was calculated by determining the age of the oldest brood
in each colony.
Results
Numbers of spermatozoa counted in the spermatheca of Africanized
and European queens are summarized in Table 7-1. The mean number of
spermatozoa in Africanized queens (4.09 +0.50 million) was not
different from the mean number in European queens (4.12 +0.58 million;
t-test, two-tailed, alpha = 0.05). European queens began oviposition on
the 10th day post-emergence, one day sooner than the Africanized queens
(P<.001; Kolmogorov-Smirnov two-tailed test, chi-square distribution, df
= 2). The time from adult emergence to initiation of oviposition
reported here (Table 7-1) is longer than the maturation interval
reported in Chapter IV, which may be a result of having introduced
virgin queens into the mating colonies rather than mature queen cells.
There was no correlation between the time post-emergence to initiation
of oviposition and the number of spermatozoa in the spermatheca
(Spearman's rank correlation coefficient, two-tailed, alpha = 0.05).
The acceptance of the Africanized and the European virgin queens


CHAPTER IV
QUEEN DEVELOPMENT AND MATURATION
Introduction
African honey bees, Apis mel1ifera scutellata (formerly classified
as adansonii; Ruttner 1976a, 1976b, 1981), were introduced into
southeastern Brazil in 1956 (Kerr 1967; Michener 1975; Woyke 1969). The
following year, swarms escaped and hybridized with the established
European honey bees (primarily A. m. 1iaustica and mellifera) that had
been introduced by 1845 (Gerstaker cited in Pellet 1938; Woyke 1969).
The descendents from this hybridization are known as Africanized honey
bees (Goncalves 1982).
Africanized honey bees in South America have a very high annual
reproductive rate compared with European honey bees in temperate
regions. Based on demographic data collected in French Guiana, the net
reproductive rate for Africanized bees is estimated to be 16 colonies
per colony per year (Otis 1980, 1982a). In comparison, the annual rate
determined for European honey bees in North America was 0.92-0.96
(Seeley 1978) or 3-3.6 when afterswarms are considered (Winston 1980a;
Winston, Taylor and Otis 1983). This dramatic difference in
reproduction between these two honey bee populations may be a result of
length of time throughout the year that resources are available in the
tropics compared with temperate regions (see Chapter VIII) and/or
55


APPENDIX B
HONEY BEE SIZE, COMB CELL SIZE AND SIZE VARIATION


126
23 minutes separating the mean times of peak flight activity for each
population (Taylor Kingsolver and Otis in press). This difference does
not provide a satisfactory mechanism for reproductive isolation between
the Africanized and European honey bee populationsparticularly because
any unfavorable climatic conditions e.g. high winds cloudiness high
humidity, or rain (Gary 1975), cause mating flights of reproductives
from both populations to more completely converge to times of favorable
weather conditions. The data on reproductive success (determined by the
number of spermatozoa in the spermatheca) of European queens mating with
Africanized drones presented in this study demonstrate that any
differences in mean peak flight times did not effectively prevent
hybridization of European queens with Africanized drones.
Evidence that extensive hybridization has already occurred between
the introduced African honey bees and the previously established
European honey bees can be demonstrated by the increase in genetic
diversity in the Africanized population. For example, Adams, Rothman,
Kerr and Paulino (1977) concluded that the large increase in number of
sex alleles in the population of honey bees in southeastern Brazil is a
result of hybridization between African and European honey bees. Page
and Erickson (1985) also suggest that hybridization has occurred based
on the variation in behavior and appearance of Africanized bees in
Venezuela.
Impact of Hybridization
Demonstration that successful hybridization can and does occur has
important implications with respect to the potential impact the
Africanized honey bee will have in North America. First is the negative
impact that hybridization would have. The parental African population


154
TABLE 8-1. Factors affecting colony survival and reproductive success
for Africanized and European honey bees in Venezuela.
POPULATION
FACTOR WITH ADVANTAGE REFERENCES
COLONY DEMOGRAPHY
Growth Rate
Egg Laying Rate No Difference Chapter VI; Chapter V
Worker Longevity European Winston & Katz 1981
Swarm Age Structure No Data
Brood Mortality No Data
Brood Production No Data
during Resource Dearth
Interval3
Worker Development Time Africanized
Queen Maturation European
REPRODUCTIVE OUTPUT
Number of Afterswarms No Data
per Swarming Cycle
RESOURCE UTILIZATION
Foraging Behavior Africanized
Brood Production
Efficiency
Bee Size
Resource-Induced
Absconding
PREDATION
Colony Defense
No Data
Africanized
Africanized
Africanized
Disturbance-Induced Africanized
Absconding
Flight Behavior No Data
NEST CAVITY VOLUME No Data
DENSITY-DEPENDENT FACTORS No Data
Chapter II
Chapter IV
Rinderer, Bolten, Collins
& Harbo 1984; Rinderer,
Bolten, Harbo & Collins
1982; Rinderer, Collins &
Tucker 1985; Nunez 1979,
1982; Winston & Katz 1982
Chapter III
Winston, Otis & Taylor 1979
Winston, Taylor & Otis 1983
Collins, Rinderer, Harbo &
Bolten 1982
Bolten, pers. observation;
Winston, Taylor & Otis 1983
aInterval from swarming to beginning of population increase.


39
1982). The European genotypes (YD28 and WEI) were imported into
Venezuela from the U.S. Department of Agriculture Bee Breeding and Stock
Center Laboratory in Baton Rouge, Louisiana, U.S.A., and from a
commercial queen producer from southeastern U.S.A., respectively. Queen
YD28 was artificially inseminated with the spermatozoa from one drone;
queen WEI was naturally mated. The third European genotype (SDY1) was a
daughter from line YK produced by another commercial queen producer from
southeastern U.S.A. and artificially inseminated in Venezuela with a
single drone from the same commercial line.
Two reciprocal hybrid lines were established from artificially
reared queens (Laidlaw 1979) that were instrumentally inseminated with
spermatozoa from single drones: Africanized queen x European drone
(SDA12) and European queen x Africanized drone (SDY10 and SDY11). The
Africanized queen and drone source was A26. The European queen and
drone source was line YK. The hybrid lines were therefore genetically
similar, but were the reciprocal of each other with respect to their
queen and drone sources.
Queens were produced by the standard method of transferring young
larvae from the desired queen line into artificial queen cells which
were then introduced into cell-producing colonies (Laidlaw 1979).
Mature queen cells were put into an incubator (35 + 1C) 72 hours prior
to adult emergence. Newly emerged virgins were marked for individual
identification and then put into individual cages and maintained in a
strong, queenless colony for approximately one week until they were
artificially inseminated.
Drones for instrumental inseminations were produced by caging drone
comb containing sealed drone pupae from the desired drone source lines.


TABLE 6-11. Comparison of overall egg laying rates
between Africanized and European sister
queens with Africanized and European
nurse bees during the first brood cycle.
AFRICANIZED EUROPEAN
NURSE BEESa NURSE BEES
AFRICANIZED QUEENS
A62 (W81)
696

A62 (W85)

447
A57 (W42)
328
A57 (W41)

568
EUROPEAN QUEENS
GK30 (Y63)
772
__
GK30 (Y61)
775
aOverall egg laying rate = (TBjj/17)(4.25).