• TABLE OF CONTENTS
HIDE
 Title Page
 Acknowledgement
 Table of Contents
 List of Tables
 List of Figures
 Introduction
 Literature review
 Experiment I
 Experiment II
 General discussion and conclus...
 Summary
 Literature Cited
 Appendix
 Biographical sketch
 Copyright














Title: Effect of level of nitrogen on growth and reproductive physiology of young bulls and rams
CITATION PDF VIEWER THUMBNAILS PAGE IMAGE ZOOMABLE
Full Citation
STANDARD VIEW MARC VIEW
Permanent Link: http://ufdc.ufl.edu/UF00091586/00001
 Material Information
Title: Effect of level of nitrogen on growth and reproductive physiology of young bulls and rams
Series Title: Effect of level of nitrogen on growth and reproductive physiology of young bulls and rams
Physical Description: Book
Creator: Meacham, Thomas Neil,
 Record Information
Bibliographic ID: UF00091586
Volume ID: VID00001
Source Institution: University of Florida
Holding Location: University of Florida
Rights Management: All rights reserved by the source institution and holding location.
Resource Identifier: alephbibnum - 000402342
oclc - 24680194

Downloads

This item has the following downloads:

Binder1 ( PDF )


Table of Contents
    Title Page
        Page i
    Acknowledgement
        Page ii
    Table of Contents
        Page iii
    List of Tables
        Page iv
        Page v
        Page vi
    List of Figures
        Page vii
        Page viii
    Introduction
        Page 1
        Page 2
    Literature review
        Page 3
        Page 4
        Page 5
        Page 6
        Page 7
        Page 8
        Page 9
        Page 10
        Page 11
        Page 12
        Page 13
        Page 14
    Experiment I
        Page 15
        Page 16
        Page 17
        Page 18
        Page 19
        Page 20
        Page 21
        Page 22
        Page 23
        Page 24
        Page 25
        Page 26
        Page 27
        Page 28
        Page 29
        Page 30
        Page 31
        Page 32
        Page 33
        Page 34
        Page 35
        Page 36
        Page 37
        Page 38
        Page 39
        Page 40
        Page 41
        Page 42
        Page 43
        Page 44
        Page 45
        Page 46
        Page 47
        Page 48
        Page 49
        Page 50
        Page 51
        Page 52
        Page 53
        Page 54
    Experiment II
        Page 55
        Page 56
        Page 57
        Page 58
        Page 59
        Page 60
        Page 61
        Page 62
        Page 63
        Page 64
        Page 65
        Page 66
        Page 67
        Page 68
        Page 69
        Page 70
        Page 71
        Page 72
        Page 73
        Page 74
        Page 75
        Page 76
        Page 77
        Page 78
        Page 79
        Page 80
        Page 81
        Page 82
        Page 83
        Page 84
    General discussion and conclusions
        Page 85
        Page 86
        Page 87
        Page 88
    Summary
        Page 89
        Page 90
        Page 91
    Literature Cited
        Page 92
        Page 93
        Page 94
        Page 95
        Page 96
        Page 97
    Appendix
        Page 98
        Page 99
        Page 100
        Page 101
        Page 102
        Page 103
        Page 104
        Page 105
        Page 106
        Page 107
        Page 108
        Page 109
        Page 110
        Page 111
        Page 112
        Page 113
        Page 114
    Biographical sketch
        Page 115
        Page 116
    Copyright
        Copyright
Full Text










EFFECT OF LEVEL OF NITROGEN ON

GROWTH AND REPRODUCTIVE

PHYSIOLOGY OF YOUNG

BULLS AND RAMS









By
THOMAS N. MEACHAM


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









UNIVERSITY OF FLORIDA


June, 1962














ACKNOWLEDGMENTS


The author wishes to express his sincere appreciation to

Dr. T. J. Cunha, chairman of his supervisory committee, and

Drs. A. C. Warnick and J. F. Hentges, Jr., for the many hours spent

in advising and assisting in the conduction of these experiments and

the preparation of this dissertation. He is most grateful to Dr.

R. L. Shirley and Dr. T. W. Stearns for their help and advice with his

graduate work.

To his fellow graduate students, D. D. Hargrove, D. L. Huffman,

R. E. Deese and J. H. Meade, Jr., go the author's appreciation for

their many hours of help in the collection of these data.

The patience and understanding of his wife, Carol, are sincerely

appreciated.






















ii
LY~fBC;















TABLE OF CONTENTS

Page

ACKNOWLEDGMENTS ... . . . . . . . . . . ii

LIST OF TABLES . . . . . . . . . . . . iv

LIST OF FIGURES ......................... vii

INTRODUCTION . . . . . . . . . . . . 1

LITERATURE REVIEW . . . . . . . . ...... 5

EXPERIMENT I . . . . . . . . . . . 15

Experimental Procedures . . . . . . . .. 15
Results and Discussion . . . . . . . . 25

EXPERIMENT II . . . . . .... . . . . . ... 55

Experimental Procedures . . . . . . . . 55
Results and Discussion . . . . . . . . 61

GENERAL DISCUSSION AND CONCLUSIONS . . . . . . . 85

SUMMARY . . . . . . . . . . . . . . 89

LITERATURE CITED . . . . . . . 92

APPENDIX .............. ..... ...... 98


iii















LIST OF TABLES


Table Page

1 Composition of Experimental Rations--Experiment I . .. 18

2 Vitamins Added to Ration Per Ton of Feed . . . 19

5 Mineral Composition of the Trace Mineralized Salt Fed
During the Experiment . . . . . . . . 19

4 Mean Interval to First Mount, Interval to First Mate
and Libido Score by 28-Day Periods . . . ... 57

5 Summary of the Semen Exhaustion Study for Individual
Bulls and Treatment Groups by Days and Total
Production .. . . . . . . . . . . 46

6 Quantitative Evaluation of Testes and Epididymis from
the Two Groups of Bulls . . . . . . . . 48

7 Quantitative Evaluation of the Seminal Vesicles,
Prostate and Cowper's Glands from the Two Groups
of Bulls--Experiment I . . . . . . . .. 49

8 Summary of Mean Gland and Organ Weights Taken at
Time of Slaughter. .. . . . . . . . 50

9 Composition of the Purified Rations Fed the Rams in
Experiment II . . . . . . . . . . 58

10 Summary of Libido Data for the Three Treatment Groups
of Rams by 14-Day Periods .. . . . .. 75

11 Quantitative Evaluation of the Testes and Epididymis
from the Three Groups of Rams . . . . ... 80

12 Quantitative Evaluation of the Seminal Vesicles and
Cooper's Glands from the Three Groups of Rams . . 81

15 Summary of the Mean Gland and Organ Weights Taken at
Time of Slaughter from the Three Groups of Rams . . 85

14 Biological Assay of Adenophypophyses from the Three
Treatment Groups of Rams . .. . . . . 84


iv













LIST OF TABLES--Continued


Table Page

15 Summary of Total Feed, Crude Protein Per Cwt. and TDN
Per Cwt. Consumption Per Day for the Two Treatment
Groups by Periods--Bulls . . . . .... ... 99

16 Summary of the Initial and 28-Day Period Weights of
Individual Bulls and Treatment Groups (Pounds) . .. 100

17 Summary of Blood Hemoglobin, Hematocrit and Serum
Protein Values for Individual Bulls and Treatment
Groups by 28-Day Periods . . . . . . .. 101

18 Summary of the Semen Volume, Motility and the Total
Sperm Cell Production for Individual Bulls and
Treatment Groups by Periods . . . . . . .. 103

19 Summary of Individual Ram and Treatment Group Mean
Feed Consumption by 14-Day Periods . . . . .. 105

20 Summary of Individual Ram and Treatment Group Mean
Body Weights by 14-Day Periods . . . . .... 106

21 Summary of Individual Ram and Treatment Group Mean
Blood Hematocrit, Hemoglobin and Serum Protein
Values by 28-Day Periods . . . . . . .. 107

22 Summary of Individual Ram and Treatment Group Mean
Semen Volume, Motility and Total Sperm Production
Values for 14-Day Periods . . . . . .... 109

25 Analysis of Variance Mean Squares for Semen Volume,
Motility and Total Sperm Cell Production in
Experiment I, Phase I . . . . . . .... 11

24 Analysis of Variance Mean Squares for Blood Hemoglobin,
Hematocrit, Serum Protein, Libido Scores, Intervals
to First Mount and Interval to First Mate--
Experiment I, Phase I . . . . . . .... ll

25 Analysis of Variance Mean Squares for Semen Volume,
Motility and Total Sperm Cell Production--
Experiment I, Phase II . . . . . . .... 112

26 Analysis of Variance Mean Squares for Blood Components
and Libido Evaluations--Experiment I, Phase II . .. 112











LIST OF TABLES--Continued


Table Page

27 Analysis of Variance Mean Squares for Blood Hemoglobin,
Hematocrit and Serum Protein--Experiment II,
Phases I and II . . . . . . . .... ... 113

28 Analysis of Variance Mean Squares for Semen and
Libido Criteria--Experiment II, Phase I . . . .. 114

29 Analysis of Variance Mean Squares for Semen and
Libido Criteria--Experiment II, Phase II . . .. 114















LIST OF FIGURES


Figure Page

1 Effect of Protein Deficiency on Daily Total Feed
Intake of Bulls . . . . . . . ... .. 24

2 Effect of Protein Deficiency on Crude Protein and
TDN Intake Per Cwt. Daily of Bulls .......... 25

3 Cumulative Average Daily Gains of Bulls . . . ... 27

4 Effect of a Low Protein Ration on Bull Growth ..... 30

5 Effect of Protein Deficiency on Average Hemoglobin
Levels of Bulls .................... 32

6 Effect of Protein Deficiency on Average Hematocrits
of Bulls . . . .. . . . . . . 534

7 Effect of Protein Deficiency on Average Serum Protein
Level of Bulls . . . . . . . . ... 36

8 Effect of Low Protein Intake on Mean Semen Volume
of Bulls ... . . .. . .. .. .. . .. 39

9 Effect of Low Protein Intake on Mean Sperm Motility
of Bulls ...................... .. . 41

10 Effect of Low Protein Intake on Total Sperm Production
of Bulls . .. .. .. ... . .... .. . . .. 43

11 Effect of a Protein Deficiency on the Testes and
Seminal Vesicles of Bulls . . . . . . .. 52

12 Effect of a Protein Deficiency on the Cowper' s and
Prostate Glands of Bulls . . . . . . .. 54

13 Effect of Source and Level of Nitrogen on Feed Intake
of Rams . . . . . . .... . .. .. . 62

14 Effect of Source and Level of Nitrogen on Cumultaive
Average Daily Gain of Rams . . . . . .... .64

15 Effect of Source and Level of Nitrogen on Ram Growth . 67











LIST OF FIGURES--Continued


Figure

16 Effect of Source and Level of Nitrogen
Hemoglobin Values for Rams . .

17 Effect of Source and Level of Nitrogen
Hematocrit Levels of Rams .. ...

18 Effect of Source and Level of Nitrogen
Serum Protein Values of Rams . .

19 Effect of Level and Source of Nitrogen
Semen Volume of Rams . . . .

20 Effect of Level and Source of Nitrogen
Semen Motility of Rams . . .

21 Effect of Source and Level of Nitrogen
Cell Production of Rams . . .


Page


on Mean


on Mean


on Mean


on Mean


on Mean



on Total
. . .


viii


. .















INTRODUCTION


Maximum production and profit in any livestock breeding operation

depends to a large extent on reproductive efficiency. The role of the

female in achieving this efficiency has been the object of consider-

able study in the past. The part of the male, while of equal importance,

has received little or no attention in efforts to improve reproductive

efficiency in breeding programs.

Protein, one of the most essential nutrients for growth and

development, is frequently a limiting factor in the feeding value of

pasture and roughage. Since pasture and roughage usually make up a

major portion of the ration fed to both mature and growing bulls and

rams, these animals may be subjected to a marginal or deficient protein

intake. This situation is particularly acute during periods of re-

stricted plant growth when the protein content of forages may be reduced

markedly.

Several studies have been conducted recently to ascertain the

effect of reduced protein intake on the growth and reproductive per-

formance of heifers and cows. These studies indicate that when the

protein intake is reduced to 50 percent of the recommended level,

retarded growth and impairment of estrus and ovulation result. A recent

study by the author (Meacham, 1960) has revealed that the reproductive

performance of the bull, while more resistant than that of the heifer

or cow, is adversely affected by a low protein intake. With these







2



facts in mind, the experiments reported herein were conducted to

provide additional information concerning the effect of source and

level of nitrogen and protein on the growth, blood composition, and

reproductive performance of young bulls and rams.















LITERATURE REVIEW


The literature contains little information concerning the

effect of a protein deficiency on bulls or rams. In recent years,

several workers have studied the effect of reduced feed and energy

intake on the growth and reproduction of dairy bulls and boars, but

little work has been done on a protein deficiency in the male of any

of the farm animals.

In an extensive review of the relationship of nutrition to

fertility Reid (1949) reports that reduced protein intake results in

decreased appetite, inanition and loss of weight in laboratory animals

as well as large animals. The accompanying depressing effect on the

pituitary gland adversely affects the secretion of gonadotrophic

hormones which regulate the reproductive processes. Supplementation

with protein has improved the reproduction rate in beef cows (Guilbert

and Hart, 1951) and increased the testicular development and activity

in the rat (Horn, 1955). In view of these findings, a definite need

for protein for normal reproduction does exist.


Dietary Protein and Growth

A number of workers have investigated the problem of how much pro-

tein is required for normal growth of beef calves. Winchester et al. (1957)

stated that 0.59 pounds of digestible protein (IP) daily would maintain

calves for six months with little or no gain in weight. Blaxter and










Mitchell (1948) estimated that 1.1 pounds of IP were needed daily for

dairy calves for adequate growth. Armsby (1917), the National Research

Council (NRC) committee for beef cattle in 1958, Morrison (1956), and

Guilbert and Loosli (1951) reported EP values for young calves falling

in the range of 0.50 to 0.82 pounds of EP per day for normal growth.

Darlow et al. (1946), the NRC committee for beef cattle (1958),

Lofgreen et al. (1951) and Morrison (1956) gave DP requirements for

older calves of 0.8 to 0.9 pounds daily. Both the NRC (1958) and

Morrison (1956) recommended 1.2 pounds of DP per day for young bulls.

This increased protein requirement for bulls over that for steers and

heifers is in agreement with the work of Speer et al. (1957) who

indicated that young boars may have a higher protein requirement for

optimum growth than do barrows or gilts.

The protein requirements for lambs are quite varied. The NRC

committee for sheep (1957) recommends 0.17 to 0.18 pounds IP for grow-

ing rams and 0.17 to 0.20 pounds per day for fattening lambs. Morrison

(1956) gives a range of 0.19 to 0.26 pounds of DP per day for growing

rams and a similar range for fattening lambs. Gallup et al. (1952),

Ross et al. (1954) and Bush et al. (1955) reported the optimum EP

level to be 0.20 to 0.22 pounds per day for fattening lambs. Very

few research data are available related to the protein requirements

of young rams.

Oltjen et al. (1959, 1960 and 1961), Meacham et al. (1961),

Repp et al. (1955) and Gallup et al. (1952) have all reported on the

use of urea in replacing a part or all of the protein in rations for











growing lambs and rams. Slen and Whiting (1955) studied the effect of

supplemental urea in rations for gestating and lactating ewes. In all

cases, urea was found to be a satisfactory source of nitrogen although

it was usually not as effective as the conventional protein sources.

As the percentage of nitrogen supplied by urea increased, the perform-

ance of the animals tended to decrease. Repp et al. (1955) stated

that when 15 to 30 percent of ration nitrogen was supplied as urea

lamb growth was essentially the same as that when all protein-nitrogen

was used. When urea was used as the sole source of nitrogen, Oltjen

et al. (1959) and Meacham et al. (1961) found that weight gains were

lower than those of lambs and rams fed protein sources of nitrogen.


Dietary Protein and Blood Composition

Hemoglobin. The normal hemoglobin values for beef cattle

reported by various workers are quite varied. Long et al. (1952) gave

a range of 8.0 to 15.0 grams (gm.) of hemoglobin per 100 milliliters

(ml.) of blood. Dukes (1955), Byers et al. (1952), Blucker (1952),

Flipse (1957) and Davis et al. (1958) reported normal hemoglobin values

for cattle that fell in the range given by Long et al. (1952). Kehar

and Murty (1945) reported a marked decrease in hemoglobin levels in

Zebu cattle fed protein deficient rations for short periods of time.

The protein deficiency reduced the hemoglobin levels from 11.0 to 8.9

gm. per 100 ml. of blood. A low protein intake lowered the hemoglobin

levels in young bulls from 11.9 to 8.01 gm. per 100 ml. of blood

(Meacham, 1960). Bedrak (1958) and Cruz et al. (1961) have reported











similar decreases in hemoglobin levels in beef heifers and cows fed

protein deficient rations.

Dukes (1955) gives the normal hemoglobin level of sheep as

12.4 gm. per 100 ml. of blood. Weir et al. (1948) reported an average

hemoglobin level of 10.16 gm. per 100 ml. of blood for young lambs.

Klosterman et al. (1950) stated that mature ewes had normal hemoglobin

of 11 gn. per 100 ml. of blood. Klosterman et al. (1948), Weir et al.

(1948) and Klosterman et al. (1950) all reported that low protein

intake in both ewes and lambs will result in a lowered hemoglobin level.

Hematocrit. A fairly wide range of values for normal hemato-

crits is found in the literature for cattle. Maynard.and Loosli (1956)

give a range of 30 to 45 percent of the blood as packed cells or

hematocrit. Blucker (1952) reported 31 to 38 percent as normal for

Hereford cattle. Dukes (1955) states that a normal hematocrit for

cattle as a whole is around 40 percent. Davis et al. (1958) reported

that Florida cattle have a hematocrit of 46 percent. Blincoe and

Brody (1951) give a range of 36 to 46 percent as normal for several

dairy breeds.

Kehar and Murty (1945) and Meacham (1960) found that low protein

rations reduced the hematocrit of beef cattle from about 45 percent to

40 and 27 percent for Zebu and Hereford and Angus cattle respectively.

Low protein rations similarly reduced the hematocrit of beef heifers

as reported by Bedrak (1958) and Cruz et al. (1961).

The average hematocrit for sheep is considerably lower than

that for cattle. Dukes (1955) gives a figure of 32 percent as normal

for sheep.










Serum Protein. The normal serum protein level in the blood

of cattle is 7.60 percent according to Dukes (1955). Gardner (1950)

gives a range of 8.0 to 11.0 percent serum protein in cattle. Crook-

shank and Sims (1955) give a lower range of 6.44 to 9.10 percent for

serum protein levels in cattle.

Kehar and Murty (1945) in their work with Zebu.cattle state

that adequately fed cattle have a serum protein level of 8.60 percent

compared to 7.32 percent in protein deficient animals. Beef bulls

fed low protein rations had serum protein levels of 6.07 percent as

compared to 7.63 percent serum protein in control bulls (Meacham,1960).

Other workers have reported similar decreases in serum protein per-

centages in beef heifers fed low protein rations (Bedrak 1958, and

Cruz et al.,1961).

Klosterman et al. (1950) reported a range of 6.9 to 7.3 per-

cent serum protein in the blood of adequately fed pregnant ewes. Dukes

(1955) gives a normal serum protein value of 5.58 percent for sheep.

Klosterman et al. (1948) report that low protein intake in ewes will

lower the serum protein in both the blood of ewes and the blood of

their lambs.


Dietary Protein and Reproduction

With the exception of a few amino acid and protein studies

with the rat and the human, the literature is practically devoid of

information concerning the effect of a protein deficiency on reproduc-

tion in the male. A preliminary study with bulls by the author (Meacham,

1960) provides the only information available concerning a protein











deficiency and reproduction, in males, with large animals. This work

indicated that such a deficiency had adverse effects on growth and

reproduction as shown by semen quality and libido.

Keller et al. (1946) reported that young male and female rats

fed a tryptophan deficient diet for as little as three days manifested

sterility in both sexes. Later work by Berg and Rohse (1947) however,

failed to confirm the work of Keller et al. The former workers were

unable to produce sterility in the young rats on a similar deficient

diet. Shettles (1960) in a recent report indicated that tryptophan,

as well as lysine and arginine, was essential for spermatogenesis

in man. The argininedeficient diet resulted in decreased sperm con-

centration and lowered motility in human semen. Leathem (1956)

reported that mature rats fed a protein free diet exhibited normal

spermatogenesis for 30 days. After 90 days, testes weights had

decreased 50 percent and complete atrophy had occurred in some cases.

Spermatogenesis was sharply reduced or stopped entirely. Working with

immature male rats, Horn (1955) found that a protein free diet in-

hibited testes maturation and spermatogenesis. Replacing the protein

in the diet brought about increased size and activity of the testes

with a corresponding increase in sperm production.

An early indication of a protein deficiency is the loss of

appetite and subsequent decrease in feed intake. With this in mind,

the work done on restricted energy and feed intake might well be

pertinent to the problem of protein deficiencies and reproduction.

Numerous workers have studied the effect of restricted feed

intake or energy intake on the reproductive performance of laboratory










and large animals. Friedman and Turner (1959) stated that an inhibi-

tion of spermatogenesis occurred when male rats were restricted to

feed intakes 25 to 50 percent below that of self-fed controls. When

feed intake was reduced to the point where body growth was halted,

testes atrophied and spermatogenesis was ultimately halted. Studies

with the male reviewed by Reid (1949) revealed a similar deficiency

situation when the energy intake was reduced 15 to 30 percent of the

self-fed controls. Talbert and Hamilton (1955), however, reported

that initiation of sperm production was not influenced by a reduction

of growth rate to one-third that of normal young male rats. In con-

trast to these data, Teitelbaum and Gantt (1956) reported an increase

in sperm production in dogs subjected to short periods of starvation.

When the dogs were returned to a normal ration, the level of sperm

output dropped back to normal, pre-starvation levels. Chang and

Sheaffer (1956) took exception to these data and reported that the

sperm they were measuring had been produced before the starvation

period was started and thus had not been affected by the treatment.

It appears from these studies that the initiation and maintenance of

spermatogenesis is impaired by a reduction in either feed or energy

intake severe enough to stop body growth. Apparently, as long as the

animal is able to grow, attainment of puberty and spermatogenesis are

not materially affected.

Studies with large animals have indicated fairly similar

effects from reduced energy and total feed intake on reproduction.

Jones et al. (1942, 1945) report that young dairy bulls restricted











to a roughage ration after the first seven months of age reached

puberty later and grew at a slower rate than bull calves supplemented

with concentrates and vitamins. There was no difference in the fer-

tility of the two groups of bulls at the normal breeding age of 24

to 40 months. The roughage-fed bulls did, however, reach puberty at

a later date than the supplemented bulls. These workers concluded

that any ration which permitted normal growth to 24 months of age

would be adequate for normal reproductive development. This is in

agreement with statements made by Walton (1949) and Rice and Andrews

(1951) that rations sufficient for normal body growth were also ade-

quate for reproduction. Reid (1949) reported essentially the same

results in a study where young dairy bulls were grown out on a reduced

energy ration.

Baker et al. (1953, 1955a, 1955b) and Flipse (1957) report

that the first ejaculate with viable sperm was obtained at 9 to 10

months of age with dairy bulls. Flipse et al. (1955) stated that young

dairy bulls restricted to 70 percent of the energy recommended by the

(NRC) committee for dairy cattle were delayed 12 to 14 weeks in reach-

ing puberty. They found that feeding 130 percent of the energy

recommended did not hasten the onset of puberty materially. Semen

concentration and motility were also adversely affected by the restric-

tion of the energy intake to 70 percent.

[Bratton (1953, 1957) and Bratton et al. (1959) have published

an extensive study conducted at Cornell University in which three

levels of total digestible nutrients(TDN) were fed to young dairy bulls.










The bulls were started at one week of age and the study was continued

over an 80-week period. These workers report that feeding 60 percent

of the recommended TDN allowance for dairy heifers delayed the onset

of spermatogenesis eight weeks beyond that of the 100 percent TDN

group. Feeding 160 percent of the allowance hastened the onset by

six weeks. It was found in the study that essentially the same total

amount of TDN was required to raise a bull to sperm-producing age

regardless of the feeding level. Those bulls on the high level merely

reached puberty at an earlier age. The non-return rates of females

bred by artificial insemination from the three groups of bulls were

identical, all 74 percent. This would indicate that the fertility of

the bulls was not influenced by the level of TDN feeding. Histo-

logical investigations were made of the tissue from the testes of

these bulls slaughtered at various intervals during the study. This

work indicated that the delay in sperm production in the low TDN level

bulls was due to a retarded differentiation of the seminiferous

tubules and interstitial tissue in the testes.

Davies et al. (1957) conducted a total feed intake deficiency

experiment with identical twin beef calves. These workers studied

the androgenic activity and the appearance of spermatozoa in the semen

of deficient and control calves. One twin from each pair was assigned

to the deficient group and the other to a normally-fed control group.

The appearance of fructose and citric acid in the semen was considered

an indication of the onset of testosterone production. The normally-

fed bulls began secretion of fructose and citric acid at five months











of age while the deficient bulls were delayed an average of four months

beyond this time with testosterone production not starting until the

bulls were nine months old. The low feed intake delayed the start of

spermatogenesis, but to a lesser extent. Sperm were observed in the

ejaculate obtained by electrical stimulation at 9.5 and 10.5 months

of age for the control and the deficient bulls respectively. Histo-

logical studies indicated that the differentiation of the seminiferous

tubules and interstitial tissue was delayed in the testes of the

deficient bulls. This accounted for both the retarded hormone secre-

tion and the delayedspermatogenesis. Shirley et al. (1960) reported

a marked decrease in fructose and citric acid concentrations in semen

obtained from protein deficient bulls. There was a corresponding

decline in spermaogenesis and libido in these same bulls (Meacham,

1960).

Mann and Walton (1953) reduced the feed intake of one mature

Dexter bull to 50 percent of the required amount and later to a near

starvation level for 25 weeks. This drastic reduction in feed intake

was without effect on sperm concentration, motility or sperm morphology.

The volume of semen produced was, however, reduced somewhat.

Hulet et al. (1956) studied the effect of roughage versus

roughage and concentrates feeding on the fertility of rams. They

found no significant differences in the fertilizing capacity of the

semen from the two groups of rams. Matousek et al. (1959) investigated

the effect of high and low energy rations on various semen character-

istics in mature rams. Rams fed a bulky, low energy ration produced










semen equal to that of rams fed concentrates with respect to volume,

motility, sperm concentration, fructose content and pH.

Theorizing from the data available in the literature, it seems

that once sexual maturity is attained, spermatogenesis and fertility

are not easily affected by restricting the energy or total feed intake

of large animals. Work by Dutt and Barnhart (1959) with young boars

indicates that perhaps a different situation exists with the boar.

These workers were unable to affect the concentration, motility or

morphology of the semen from young boars fed as little as 50 percent

of the NRC requirements for TDN. The volume of semen produced by the

boars on the 50 percent level was significantly smaller than those

on the 70 to 100 percent levels. Puberty, libido and fertilizing

capacity of the semen were not influenced by the low levels of TDN

intake. Steverman et al. (1961) studied the effect of reduced feed

intake in mature boars. Three levels of feed intake were used;

a level supplying 120 to 150 percent of the NRC requirements, a level

just meeting these requirements and a level providing only 50 to 75

percent of the required intake. The results of this study were quite

similar to those reported above. The volume of semen produced by the

boars on the low level was reduced somewhat, but there was no differ-

ence in total sperm production, motility or fertility between any of

the three groups. Some decline in sperm concentration was noted after

the boars on the low intake level had reached a point of lethargy.

Conflicting reports appear in the literature concerning the

role played by quality of protein in reproduction. Reid (1949)











reviewed work by Russian workers who reported beneficial responses in

semen quality from dairy bulls fed animal protein sources. These

workers fed bulls, in an Artificial Insemination Center, blood meal

and fish meal and found an increase in sperm concentration and motility.

Branton et al. (1948) and Flipse et al. (1956) attempted to duplicate

this favorable response from animal protein sources. Flipse and co-

workers fed both liquid and dried skim milk and soybean and linseed

meal to young dairy bulls. They reported that the source of protein

had no effect on growth, age at puberty, motility or morphology of

the sperm cells. There was an increase in sperm concentration with

the liquid skim milk, but the milk reduced the total volume of semen

produced. Branton and his co-workers fed corn gluten meal, dried

skim milk, and soybean meal to young dairy bulls. They were unable

to show any advantage attributable to the animal protein sources in

growth rate, age at puberty or semen quality. Apparently, it is

questionable whether or not animal protein feeds offer an advantage

in well-balanced rations for young breeding bulls. Flipse and

Almquist (1954) investigated the effect which dehydrated young grass

had on the reproductive efficiency of dairy bulls. They reported

no differences in the non-return rates of females for the treated and

control bulls. Hultnas (1959), in his work with dairy bulls in

Sweden, stated that part of the variation in libido, total sperm per

ejaculate and percent abnormals could be due to changes in forage

quality. He associated the decline in reproductive performance with

seasons of the year when the protein content of the forage was low.















EXPERIMENT I


Experimental Procedures


Animals

In this experiment 12 weaned Angus bull calves were used.

The average weight and age at the start of the experiment was 590

pounds and approximately 315 days of age. The bulls were purchased

from the M&M ranch at Loxahatchee, Florida. The bulls were brought

to the Purebred Beef Unit shortly after weaning and fed on pasture for

approximately two and a half months before the experiment was initiated.

Assignment to the treatment groups was accomplished in the

following manner. The bulls were exposed to an "estrogenized" heifer

of suitable size. As the bulls attained puberty, indicated by suc-

cessfully mating the heifer, they were assigned to a control and a

protein deficient group. Every third bull to mate the heifer was

placed in the control group and the other two bulls were placed in

the low protein group. This procedure was followed in an effort to

obtain bulls of the same physiological age in both treatment groups.

The average weights of the four control bulls and the eight low

protein bulls were 420 and 575 pounds, respectively, when the exper-

iment was started on October 8, 1960.

Facilities

The experiment was conducted at the South Experimental Barn











at the Purebred Beef Unit. The barn contained individual feeding pens

equipped with feed boxes and waterers. The bulls had access to two

small lots on each side of the barn at night. Water was available

to the animals in these lots.

Scales, working pens and chutes were available at the main

barn a short distance away. A stanchion was constructed in one of

the small lots by the barn to secure a heifer for libido checking.


Feedstuffs and Rations

The composition of the experimental rations, their crude pro-

tein analysis and estimated total digestible nutrients (TDN) content

are given in table 1. Ground corn cobs and shuck meal was used to

supply roughage in the rations. Molasses was included in the rations

to improve their palatability and insure a uniform blending of the

fine textured ingredients with the coarser materials in the rations.

The concentrate portion of the rations was mixed and bagged

at the beginning of the trial for the entire experimental period.

This was done to obtain rations as uniform as possible throughout the

study. The molasses and corn cob and shuck meal were mixed with the

concentrates periodically as needed during the trial. Vitamins A, D

and E were mixed in the concentrate portion along with the trace

mineralized salt and bone meal. The vitamin and mineral mixtures are

given in tables 2 and 5, respectively.

The average crude protein content of the control and low pro-

tein rations were 13.75 and 1.61 percent, respectively. Protein











analyses were made on each lot of feed mixed during the experiment.

Estimated TDN content was 67.41 and 73.45 percent for the control and

low protein rations, respectively. A higher energy level in the low

protein ration was used to counteract in part the reduction in total

feed intake which occurs on a low protein ration.


Procedures

Feeding and Management. The bulls were fed once daily in the

morning in individual feeding stalls. The bulls received 2 percent

of their body weight of the mixed ration daily throughout the exper-

iment. The animals remained in the stalls until late afternoon. At

this time they were turned out into the small lots adjacent to the barn

for the night.

Feed refused was weighed back daily and the net feed intake

recorded.- Feed consumption was summarized for each bull weekly and

for the treatment groups every 28 days.

Termination of the experiment was to occur when four of the

eight protein deficient bulls had died or been sacrificed.

Body Weights. The bulls were weighed every 14 days in the

morning before feeding. The feed intake for the following two-week

period was based on these weights. Average daily gains (ADG) per

weigh period and cumulative daily gains were summarized for each

14-day period and for each 28-day period.

Blood Evaluation. Blood samples were taken initially and at

intervals of 28 days for the first two months. After this, bleedings

were done every 14 days for the remainder of the experiment.













TABLE 1

COMPOSITION OF EXPERIMENTAL RATIONS--EXPERIMENT I


Control Ration Low Protein Ration
Ingredient C in Ration S in Ration

Granulated Corn Starch 16.00 42.00

Granulated Corn Sugar 13.00 27.00

Standard Cane Molasses 3.00 3.00

Corn Cob and Shuck Meal 25.00 25.00

Yellow Corn Meal 16.00

Cottonseed Meal, 41% 25.00

Trace Mineralized Salt 1.00 1.00

Steamed Bone Meal 1.00 2.00

Vitamin Supplementa
100.00 100.00

Crude Proteinb 13.75 1.61

Estimated TDNc 67.41 73.45

aVitamins were added in corn starch carrier to each ration
at the time the concentrate portion was mixed.
bActual crude protein analysis of the rations at the Nutrition
Laboratory.
CTDN estimated from Morrison's Feeds and Feeding ingredient
tables.











TABLE 2

VITAMINS ADDED TO RATION PER TON OF FEED


Vitamin International Units

Vitamin Aa 1,800,000

Vitamin D2b 250,000

Vitamin Ec 3,468


aVitamin A used was the Dawe's Laboratories "Fixtay";
10,000 IU of vitamin A per gram.
bVitamin D2 was supplied by Dawe's Laboratories Dry
D2; 4 million USP units of vitamin D2 per pound.
cVitamin E was obtained from the American Cyanamid
Co.; 20,000 IU of vitamin E per pound.




TABLE 5

MINERAL COMPOSITION OF THE TRACE MINERALIZED SALT
FED DURING THE EXPERIMENT


Mineral in Mixture

Salt (NaC1) 98.800

Manganese 0.250

Iron 0.270

Copper 0.033

Cobalt 0.010

Iodine 0.007

Zinc 0.005

Inert Material 0.625
100.000










Blood was drawn from the jugular vein and analyzed for the

following components: blood hemoglobin, hematocrit and serum pro-

tein. Hemoglobin values were determined by the method of Cohen

and Smith (1919). Hematocrit values were obtained by the Wintrobe

method using the Wintrobe tubes and centrifuging the blood for 50

minutes at 3,000 rpm. Serum protein concentration was determined

by the Kjeldahl method as described by the Association of Official

Agricultural Chemists (1950).

The blood data were summarized for each 14-day period and by

treatment groups every 28 days.

Libido Evaluation. The bulls were checked for libido by

exposing each bull individually to an "estrogenized" heifer secured

in a stanchion. Each bull was allowed a maximum of ten minutes with

the heifer. Records were kept of the amount of interest displayed

by the bull, the number of mounts made and the number of matings

completed. The following system of scoring the interest displayed

was used: 1, no interest; 2, interest but did not mount; 3, mounted

without an erection; 4, mounted with an erection; and 5, mated the

heifer.

Libido was checked in the described manner every 14 days.

The data were summarized for each 14-day and 28-day period by indi-

Vidual and treatment group.

Semen Evaluation. Semen was collected routinely throughout

the study to detect and evaluate changes in the various semen char-

acteristics. Semen was collected by means of an electro-ejaculator.











The criteria studied were semen volume, motility, sperm

concentration and total sperm production. Volume of semen was

determined directly by collecting the semen in calibrated tubes.

Motility was evaluated to the nearest 5 percent by microscopic

examination of the cells with progressive movement. A blood hema-

cytometer was used to determine the number of sperm per cubic

millimeter.

Single samples of semen were collected from each bull for

the first four months and duplicate samples collected for the final

two months of the experiment.

Midway through the experiment on January 5, 1961, semen was

collected and frozen for use in a fertility study. Ampules of semen

containing equivalent numbers of motile sperm were prepared from each

of 10 bulls (four control and six low protein animals). Two of the

low protein bulls were not included due to the poor quality of the

semen collected from them. An average of 24 ampules from each con-

trol bull and 21 from each low protein bull were used to inseminate

210 two-year-old heifers at the Deseret Farms of Florida, Melbourne,

Florida. The heifers were palpated to determine the number pregnant

on July 20, 1961.

On January 26 and 27, 1961, a semen exhaustion study was

conducted. Four ejaculates from each bull on each of the two days

were collected. Two collections were made in the morning and two

in the afternoon on each day. Evaluation of the semen characteristics

was made for each of the collections.










Histological Investigation. Tissue samples from the testes,

epididymis, prostate, Cowper's Gland and the seminal vesicles were

taken and fixed in formalin solution for histological studies.

Samples were taken from the four low protein bulls that were sacrificed

prior to the termination of the experiment.

The eight bulls, four control and four low protein, remain-

ing at the end of the experiment were slaughtered at the Experiment

Station Meats Laboratory. Tissue samples were obtained from these

bulls at this time.

The procedures of Gurr (1956) were followed for fixing,

imbedding, slicing and staining with eosin-hematoxylin stains.

Microscopic evaluations were made (with a calibrated eye piece) of

the size and development of the secretary tissues in the secondary

sex glands. The ducts of the epididymis were studied for differences

in size and sperm content. The testes were examined for the relative

size and development of the seminiferous tubules, germinal epithelium

and secretary tissue.

Additional slaughter data collected were: size and weight

of the testes, weights of the adeno hypophysis, the secondary sex

glands, the thyroid and the adrenal glands.

Statistical Analysis. The blood, libido, and semen data were

summarized for 28-day periods to facilitate statistical analysis.

The data were then divided into two phases; phase I which included

all 28-day periods in which all eight of the low protein bulls were

represented and phase II where the number of protein deficient bulls










varied due to death losses. There were five periods in phase I for

the semen data and four periods for the libido and blood data.

Phase II contained the final two periods for the three criteria

analyzed. The two phases were analyzed separately.

The Analysis of Variance technique for repeated-measurement

experiments as described by Danford et al. (1960) was used.


Results and Discussion


Feed Intake

The low crude protein content of the deficient ration had

an immediate effect on the appetite of the low protein bulls. There

was a marked difference in consumption between the two treatment

groups at the end of the first 28 days. At this time, the control

and low protein bulls were consuming 7.54 and 4.17 pounds of feed

per head per day, respectively. The same pattern continued to hold

true throughout the experiment. Figure 1 illustrates this effect

of a protein deficiency on feed intake. The average feed intake for

the control and the low protein bulls was 9.78 and 3.28 pounds

daily, respectively, for the experimental period.

Considering the feed intake on a body weight basis, the

differences are not as great. The average total feed intake per

100 pounds (cwt.) of body weight (table 15, in Appendix) was 1.86

and 1.13 pounds daily for the control and the low protein bulls,

respectively. In figure 2 the changes in crude protein and TDN

intake per cwt. of body weight daily are shown. The control bulls





















CONTROL
S ----- LOW PROTEIN


- - - -


1 2 3 4 5 6 7
28-DAY PERIODS

Figure 1. Effect of protein deficiency on daily total feed intake of bulls.




















, --
-C


1.3


1.1


0.9


0.7


. CONTROL
--- LOW PROTEIN
I I I I I I p
1 2 3 4 5 6 7
28-DAY PERIODS


Figure 2.


Effect of protein deficiency on crude protein and TDN intake per cwt.
daily of bulls.


s^



RPLI


- CONTROL
S-----LOW PROTEIN


I' j


.25


.20


.020


.010


0


a
H3(
gd


at











consumed about 1.25 pounds of TDN per cwt. of body weight fairly

consistently during the trial. This was not unexpected since the

controls were fed according to body weight. The TDN intake by the

low protein bulls, on the other hand, declined steadily even on a

body weight basis. The low protein bulls were losing appetite faster

than body weight.

The intake of protein, however, was fairly stable throughout

for both groups. Again, this would be expected for the control

bulls due to the method of feeding. The lack of any appreciable

change in protein intake, in spite of the marked change in total

feed intake, was due to the very low protein level in the deficient

ration.

These data indicate rather conclusively the definite depress-

ing effect of low protein levels on feed intake. This response is

in good agreement with the work of Guilbert and Hart (1951) and

Reid (1949) which demonstrated a similar effect with low protein

rations in both large and small animals. The results compare closely

with, and hence tend to confirm, earlier work by the author (Meacham,

1960) with low protein rations for beef bulls.


Body Weight Changes

The immediate reduction in feed intake by the low protein

bulls was reflected in a corresponding loss of body weight. The

pattern of weight changes is shown in figure 5. The control bulls

gained rather slowly the first 28-day period and then maintained
































CONTROL

LOW PROTEIN


- - -


- -------- --


1 2 3 4 5 6 7
28-DAY PERIODS


Figure 3. Cumulative average daily gains of bulls.


-6


-.8


-1.0


-1.1


1.1


1.0










an ADG of about a pound a day for the remaining periods. The ADG for

the 182 days of the experiment for the control bulls was 1.1 pounds.

The low protein bulls lost weight continuously throughout the exper-

imental period with the greatest losses occurring in the early part

of the study. The weight losses were less severe during the final

periods, due possibly to a lack of any additional tissue that could

readily be mobilized. The low protein bulls had a negative daily

gain of 0.71 pounds for the 182 days. The individual bull weights

for the seven periods are summarized in table 16 in the Appendix.

The control bulls gained an average of 200 pounds and the low

protein bulls lost an average of 150 pounds during the trial. The

ranges in body weight changes were losses of 115 to 145 pounds for

the low protein bulls and gains of 190 to 210 pounds for the control

bulls. As indicated by these relatively close ranges, the bulls dis-

played a very uniform response to the experimental rations.

Figure 4 illustrates the effect of the low protein ration on

the growth, weight and general appearance of the bulls.

Four of the low protein bulls died or were sacrificed shortly

before dying. The death of the fourth bull terminated the experiment

in accordance with the experimental plan. The four bulls survived

120, 132, 169 and 182 days respectively. The cause of death was

primarily due to starvation.





















Figure 4. Effect of a low protein ration on bull growth.


A low protein bull at the end of the experiment.













































-4-
-i -

hi

pl
b'4








L_











Blood Composition

Hemoglobin. The effect of the low protein ration and reduced

feed intake on the hemoglobin levels of the bulls is shown in figure 5.

There was no significant difference between the two groups for the

first four 28-day periods. As indicated in figure 5, a definite

spread occurred in the mean hemoglobin values during the final two

periods. Statistical analysis of the data indicated a significant

(P < .01) difference due to treatment in these final periods. The

significant interaction (P < .05) between treatments and periods sig-

nified that the increasing difference between the means was due to

an increase for the controls and a decrease in hemoglobin values for

the low protein bulls.

The final hemoglobin values 13.00 and 7.91 grams of hemoglobin

per 100 ml. of blood for the control and low protein bulls, respec-

tively, fit the normal range for hemoglobin reported by Long et al.

(1952) of 8.0 to 13.0 g. per 100 ml. of blood. In view of this, it

can not be concluded that the protein deficiency produced a partic-

ularly severe anemic condition in these low protein bulls. The mean

hemoglobin value for the low protein bulls in this experiment compares

quite closely with that reported by the author (Meacham, 1960) for

similar protein deficient bulls.

Individual and group hemoglobin values are summarized for

each 28-day period in table 17 in the Appendix. The Analysis of

Variance mean squares for the blood components are given in tables

24 and 26 in the Appendix.















13.0
8
S12.0


a 11.0


S10.0 -


9.0 1
a N

P 8.0 .
o
CONTROL
7.0 ---LOW PROTEIN


o I I I I I I
1 2 3 4 5 6
28-DAY PERIODS

Figure 5. Effect of protein deficiency on average hemoglobin levels of bulls.











Hematocrits. The pattern of changes in the mean hematocrit

values for the two groups of bulls followed that of the hemoglobin

values as might be expected. Figure 6 illustrates these changes

graphically. Again no significant differences due to treatment

occurred during the first four 28-day periods. There was, however,

a significant (P < .01) period x treatment interaction. The values

for the low protein bulls tended to decrease and those for the control

bulls to increase during this phase.

In periods five and six the spread between the two groups

widened markedly. The mean hematocrit values for the control and low

protein groups in the final period were 54.54 and 33.17 percent,

respectively. This difference was significant (P < .01). Signif-

icant (P < .05) effects due to period and period x treatment inter-

action were found during this final phase also. The increasing and

decreasing trends for the control and low protein groups, respectively,

continued to manifest themselves.

As was found with the hemoglobin analysis, the mean hematocrit

level for the low protein group was still within the normal range

reported in the literature. In spite of the significant difference

(P < .01) between treatments, the packed cell volumes for the surviving

bulls on the low protein ration had not reached a critical level.

Hematocrit levels were higher for both the control and the low

protein groups compared to those reported earlier (Meacham, 1960) for

bulls under similar nutritional conditions. Both the control and the

low protein group means were approximately 20 percent higher in the

























-L -


CONTROL

LOW PROTEIN


28-DAY PERIODS


Figure 6. Effect of protein deficiency on average hematocrits of bulls.


54L


48 L


50L


~^











present study. The magnitude of the difference between the treatment

groups, however, was similar. In both experiments, the low protein

groups had mean hematocrit levels 40 percent lower than the controls.

Table 17 in the Appendix summarizes the individual and mean

hematocrit values for each 28-day period.

Serum Protein. As shown in figure 7, the percent protein in

the serum appeared to be more sensitive to the low protein intake by

the bulls than the two other blood components. A significant difference

(P < .01) occurred between the two ration groups during the first four

periods. The significant period effect (P < .01) and a lack of a sig-

nificant period x treatment interaction indicated that both groups

declined in percent protein in the serum. Final mean values for the

control and low protein bulls were 6.76 and 5.34 percent, respectively.

The control value is in the lower end of the normal range reported by

Crookshank and Sims (1955) and is lower than the normal value of 6.70

percent given by Dukes (1955). The control and low protein group means

are both lower than corresponding serum protein values obtained by the

author (Meacham, 1960) in an earlier study similar to the present one.

The reduced protein level in the blood of the low protein bulls in this

study would be expected considering the severely restricted dietary

protein intake and diminished tissue protein supplies, but the com-

paratively low serum protein levels for the control group cannot readily

be explained. It is possible that the lower feed intake by the control

bulls in the present experiment may have influenced this somewhat.

The individual and mean serum protein values are summarized in

table 17 in the Appendix.















8.0


7.0





S6.0



CONTROL
5.0 LOW PROTEIN

i 1 ,I I
1 2 5 4 5 6
28-DAY PERIODS

Figure 7. Effect of protein deficiency on average serum protein
level of bulls.












Libido Evaluation

Statistical analysis of the libido data revealed a significant

difference due to treatment only in the interval to first mating

(P < .05) during phase I. The interval to first mount and libido score

were not affected sufficiently to overcome the wide variation within

treatment groups. These values are summarized by treatment and

period in table 4.



TABLE 4

MEAN INTERVAL TO FIRST MOUNT, INTERVAL TO FIRST MATE
AND LIBIDO SCORE BY 28-DAY PERIODS


Interval to Interval to Libido
Period First Mount First Mate Score
(min.) (min.) (1-5)
Low Low Low
Control Protein Control Protein Control Protein

1 2.0 0.7 5.1 4.4 4.4 4.7
2 0.4 0.6 5.2 7.1 4.6 4.4
5 0.5 3.8 2.2 9.3 4.9 3.2
4 0.3 3.4 3.8 9.3 4.8 3.5
5a 0.7 2.8 3.8 10.0 4.9 5.2


aThree of the low protein bulls died during the fifth period.



In the final period, both the libido scores and the interval

to first mate for the low protein bulls were significantly (P < .01)

lower than the controls. Only two of the low protein bulls mated the

heifer during the final period. Mean libido score and interval to

first mate (minutes) for the control and low protein bulls were 4.9

and 3.8 and 3.2 and 10, respectively, during the second phase.











The increasing physical weakness of the low protein bulls

probably influenced the increase in time required to mount and mate.

The significant (P < .01) period effect and treatment x period inter-

action in the first phase indicated that the control bulls tended to

increase in libido while the low protein bulls decreased rapidly.

There was considerable variation in libido among the bulls in

both treatment groups. One control bull mated the heifer only three

times in the entire experiment. One of the low protein bulls mated

the heifer in the preliminary period, was assigned to a treatment

group, and then failed to mate again.

In general, the lack of libido seemed to be closely associated

with the physical condition of the bulls. As the bulls lost weight

and became weak, libido decreased and eventually reached the point

where they displayed little or no interest in the heifer. It would

be difficult, therefore, to separate a possible endocrine disturbance

from a simple state of physical weakness.

Analysis of Variance mean squares for the various libido

criteria are given in tables 24 and 26 in the Appendix.


Semen Characteristics

Volume. The volume of semen produced by the two groups of

bulls was quite varied as shown in figure 8. This variation was

noted both between bulls within groups and within bulls between

collections. There were, however, significant differences in volume

between the two groups in both the first (P < .05) and the second

phase (P < .01). The within group variation was not as great in the


















8.0


i 7.0


6.0


5.0- \ />


4.0 CONTROL N "
LOW PROTEIN N ".
3.0



1 2 5 4 5 6 7
28-DAY PERIODS
Figure 8. Effect of low protein intake on mean semen volume of bulls.











second phase as in the first. The significant period effect (P < .01)

and interaction between period and treatment (P < .05) during the five

periods indicated that while the semen production for both groups tended

to decline, the low protein bulls were more severely affected.

In phase II, the significant (P < .01) treatment effect and

lack of a period or period x treatment interaction of significance

was indicative of an uncomplicated treatment effect. The low protein

bulls produced a smaller volume of semen in both periods.

The semen volume, motility and total sperm cell production

figures for individual bulls and treatment groups are given by

periods in table 18 in the Appendix. Analysis of Variance mean

squares for the various semen characteristics are presented in tables

25 and 25 in the Appendix.

Motility. As indicated in figure 9, motility values were

extremely variable. The variation within treatment groups precluded

assigning any statistical significance to the differences between

means. There was, however, a definite trend, particularly during the

latter part of the experiment, for the motility values of the control

bulls to be higher than those of the low protein bulls. Mean per-

centages for the control and low protein bulls were 70 and 24,

respectively, during the final period of the study.

The sharp decline in motility in the first part of the exper-

iment was due in part to low environmental temperatures. The semen

was collected in an exposed area where it was subjected to appreciable

cold shock. Changes in the collection technique whereby the semen















80


70 CONTROL
--0 ---- LOW PROTEIN
60


50 O


4O

30


20
SI
0 - -

1 2 3 4 5 6 7
28-DAY PERIODS


Figure 9.- Effect of low protein intake on mean sperm motility of bulls.










was protected from the cold weather improved the motility ratings

of both treatment groups. This drop and subsequent rise in percent

motility is indicated in figure 9 between periods two and five. The

significant period effect (P < .01) during phase I substantiates this

cold weather effect.

A significant treatment x period interaction (P < .01)

indicates the motility of the low protein bulls declined more rapidly

and did not recover as fast or as completely as the motility of the

control bulls. It would appear from this that the treatment was

influencing sperm motility even though there was no significant

treatment effect.

Total Sperm Production. During the first two 28-day periods,

both treatment groups had similar increases in total sperm production

as shown in figure 10. After the second period the low protein bulls

produced progressively fewer sperm per ejaculate. Their production

dropped from 1,877 x 106 in the second period to 337 x 106 sperm per

ejaculate in the final period. The control bulls had a total sperm

count average of 1,079 x 106 at the second period and a mean count

of 1,767 x 106 at the final period. Due to the variation and fluc-

tuation between bulls and between periods, the difference between

treatment means was not statistically significant until the final

two periods. At this time, the control bulls produced significantly

(P < .05) more sperm per ejaculate than did the low protein bulls.

There were no significant period or interaction effects in either

part of the experiment.









2100.


1900


o 1700- \/


1500-


1500 /
I/ \

S1100


90 /
o /
70C -


50C
CONTROL
300 LOW PROTEIN


1 2 3 4 5 6
28-DAY PERIODS


Effect of low protein intake on total sperm production of bulls.


Figure 10.











The marked difference in total cell production during the

final periods was not unexpected in light of the significant drop

in semen volume that occurred during these same periods.

It is noteworthy that even though the low protein bulls had

become so emaciated and weak that four of them had been sacrificed

by the end of the experiment, only one of the bulls had stopped

producing semen completely. The one low protein bull failed to pro-

duce semen for two consecutive collections 'just before death. As

indicated in figure 10, there was a marked reduction in sperm produc-

tion, but spermatogenesis was not completely halted.

Fertility Study. The 210 heifers used in the study were

inseminated with an ampule of semen from an experimental bull at

their first detected estrus during the breeding season. Heifers that

returned in heat were rebred with semen from a commercial insemination

source.

The results of the insemination, as determined by rectal

palpation approximately 60 days after the insemination period, were

rather disappointing. Only 68 of the 210 heifers were pregnant at

this time, Of these 68 pregnancies, 29 of them were attributed to

insemination with semen from the experimental animals. The remaining

39 pregnancies were considered to have resulted from the reinsemina-

tions with the commercial semen. This distinction was made on the

basis of fetal. size at the time of palpation.

Many of the heifers did not become pregnant regardless of the

source of semen or number of inseminations and others were inseminated











three or four times before conceiving. These difficulties, plus the

fact that bulls were turned in with the breeding herd on the week-ends

during the breeding season, precluded an accurate evaluation of the

fertility data obtained. With the beforementioned problems in mind,

an approximate fertility percentage was calculated. This percentage

was based on the number of heifers considered pregnant with semen

from each group of experimental bulls divided by the total number of

heifers inseminated with semen from the respective treatment groups

that were pregnant at the time of palpation. These percentages were

49 and 34 for the control and low protein bulls, respectively. These

data are subject to question and the confounding factors involved

need to be kept in mind when considering the apparent differences

between the two groups.

Semen Exhaustion Study. The exhaustion study was conducted

for two days during the last part of phase I. The data from this

study are summarized in table 5.

The total volume of semen and the total number of sperm cells

produced on the first day, averaged by groups, were 24.5 ml. and

4555 x 106 cells and 17.2 ml. and 1190 x 106 cells for the control

and low protein bulls, respectively. On the second day, correspond-

ing values were 15.0 ml. and 1564 x 106 cells and 11.0 ml. and 568

x 106 cells for the control and low protein bulls, respectively.

The control bulls produced a greater amount of semen contain-

ing more sperm cells than did the low protein bulls on both days. Both

the control and the low protein bulls had a marked reduction on the














TABLE 5

SUMMARY OF THE SEMEN EXHAUSTION STUDY FOR INDIVIDUAL BULLS AND
TREATMENT GROUPS BY DAYS AND TOTAL PRODUCTION


Treatment Groups First Daya Second Daya Total
and Animal No. Vol.b TCPc Vol.b TCPc Vol.b TCP

Control
6 18.4 2,784 12.3 1,898 30.7 4,682
9 43.9 11,824 20.3 1,787 64.2 13,611
11 19.6 1,344 15.6 122 35.2 1,466
14 16.0 2,260 12.0 2,450 28.0 4,710

Av. 24.5 4,553 15.0 1,564 39.5 6,117

Low Protein
4 22.9 2,136 26.4 1,472 49.2 3,608
8 23.2 88 6.4 0 29.6 88
10 29.2 2,872 12.0 960 41.2 3,832
12 18.7 132 4.0 0 22.7 132
13 14.6 568 8.0 144 22.6 712
15 9.6 0 12.0 0 21.6 0
16 11.2 2,324 10.8 683 22.0 3,00
17 8.4 1,404 8.4 1,289 16.8 2,695

Av. 17.2 1,190 11.0 568 28.2 1,759


aEach observation is the sum of four determinations.
bVolume of semen in milliliters.
cTotal cells production per ejaculate times 106.











second day of the study. One of the low protein bulls did not pro-

duce sperm cells on the first day and three of the bulls in the low

protein group failed to produce sperm on the second day.

Examination of the semen for abnormal sperm cells did not

indicate any treatment effect. The percentages of abnormal sperm

in the semen of the two treatment groups were quite similar and fairly

normal for bulls of this age.

It is evident from these data that there was a definite

difference in spermatogenic capacity between the two treatment

groups. It was possible to collect considerably more total sperm

from the control bulls as compared to the low protein bulls. This

is in agreement with earlier work by the author with bulls under

similar experimental conditions (Meacham, 1960).


Histological Investigation

The results of the histological investigation are given in

tables and 7. The objective measurement of the thickness of the

epithelial tissue of the organs and glands revealed a marked and

consistent reduction in the tissues from the low protein bulls.

The Cowper's Gland presented the greatest difference and the spermatic

ducts of the epididymis the smallest. The diameter of the semini-

ferous tubules of the testes and the ducts in the epididymis were

approximately 30 percent larger in the control bulls than in the

low protein bulls. The amount of interstitial cells in the control

bull testes was greater than that in the low protein bulls.










TABLE 6


QUANTITATIVE EVALUATION OF TESTES AND EPIDIDMIS FROM THE TWO GROUPS OF BULLS


Testes Epididymis
Treatment Epithelium Tubule Inter- Epithelium Duct Sperm
Group and Thickness Diameter Sperma stitialb Thickness Diameter Content
Animal No. (mm x 10-2) (mm) Content Tissue (mm x 10-2) (mm)
Control
6 5.4 .21 2 2 6.6 .55 2
9 4.8 .20 2 5 5.4 .51 5
11 3.0 .24 1 3 6.6 .28 2
14 4.8 .19 1 5 6.0 .30 5

Av. 5.0 .20 1.5. 3.7 6.1 .51 2.5

Low Protein
4 2.4 .15 1 2 4.8 .22 1
8 1.2 .15 1 1 3.6 .17 1
10 -_c 4.8 .50 2
12 1.8 .14 1 1 4.2 .22 2
15 5.6 .15 1 2 5 6 .14 5
15 2.4 .15 1 1 2.4 .21 1
16 4.8 .15 1 2 6.0 .23 1
17 1.8 .12 1 2 4.2 .20 1

Av. 2.6 .155 1 1.6 4.2 .21 1.5


aSperm content scored from 1 to 5 with
bInterstitial tissues scored from 1 to
cTestis sample from this bull was lost


1, no sperm, and 5, abundant.
5, with 1, sparse, and 5, abundant.
in processing.









TABLE 7


QUANTITATIVE EVALUATION OF THE SEMINAL VESICLES, PROSTATE AND COOPER'S
GLANDS FROM THE TWO GROUPS OF BULLS--EXPERIMENT I


Seminal Vesicles Prostate Cowpers
Treatment Epithelium Ratio of Epithelium Epithelium
Group and Thickness Secretory Tissuea Thickness Thickness
Animal No. (mm x 10-2)c to Total Area (mm x 10-2) (mm x 10-2)
Control
6 3.6 2 1.2 1.8
9 3.0 2 1.2 2.4
11 3.0 2 1.2 --b
14 3.0 2 __-b 1.8

Av. 3.1 2 1.2 2.0

Low Protein
4 1.8 2 .6 1.2
8 1.2 3 --_b .6
10 1.2 1 --b 1.2
12 2.4 3 .6 .6
13 1.8 3 .6 .6
15 1.8 3 -__b .6
16 1.8 5 .6 .6
17 1.8 3 --.b .8

Av. 1.7 2.6 .6 .78

aScale is based on values from 1 to 3, 1 being a narrow ratio of
secretary tissue to total area and 3 being a wide ratio.
bSamples lost in processing.
cMeasurements were made with a calibrated eyepiece and converted
to millimeters.











Perhaps the most notable difference was in the germinal

epithelium of the seminiferous tubules. The numbersof layers of germ

cells in the low protein bulls were greatly reduced and the existing

layers were characterized by a general disorganization. The germinal

epithelium of the tubules of the control bulls on the other hand,

was well developed and had numerous and orderly layers of developing

sperm cells. / These findings are in very good agreement with the

sperm production records of the two groups of bulls as noted by the

routine semen analysis. The reduction in size and secretary capacity

of the secondary sex glands in the low protein bulls was also con-

sistent with the reduced volume of semen produced by these bulls.

These differences are illustrated in figures 11 and 12.

The gross weights of these various organs and glands were

obtained at the time of slaughter and are summarized in table 8.


TABLE 8

SUMMARY OF MEAN GLAND AND ORGAN WEIGHTS
TAKEN AT TIME OF SLAUGHTER

Weight (grams)
Control Low Protein
Gland or Organa Per Cwt. Per Cwt.
Mean Body Weight Mean Body Weight

Testes (4)b 197.2 31.81 (8) 67.0 27.35
Epididymis (4) 15.6 2.52 (8) 6.4 2.62
Seminal Vesicle (4) 26.0 4.19 (8) 8.3 5.38
Cowper's Gland (3) 6.3 1.03 (7) 1.4 .57
Adrenal Gland (4) 6.8 1.10 (6) 4.0 1.63
Thyroid Gland (2) 22.8 3.52 (6) 11.9 4.86


aProstate gland tissue was taken, but the direction was
not quantitative.
bNumber in parenthesis is the number of samples represented
in the mean.





























Figure 11. Effect of a protein deficiency on the testes and
seminal vesicles of bulls.

A and B: Sections of the testis from a control
and a low protein bull, respectively

C and D: Sections of the seminal vesicle from a
control and a low protein bull, respectively.

( x 645 )



















II- - *


** *




"3U


;* .
*- *. *




* P lt .
~ -a -





f ",, r *.: ".

p 5. ... .I
*rj~ atW 55/


~c2"~IILat


St i
,I



S .'




S
'S.'..

K






























Figure 12. Effect of a protein deficiency on the Cowper' s and
prostate glands of bulls.

A and B: Sections of the Cowper's gland from a control
and a low protein bull, respectively.

C and D: Sections of the prostate gland from a control
and a low protein bull, respectively.

( x 645 )











'a-


* .. ".S-, r,

a' i V -. -" ". a ' . "
--
- C.. A ,



' . .I,





.. b->'
Ir IL
^ .--a a *e;*C! 10W-' n^ rp










S.*m *-, -
-, b kt. . r "v'-" "
*: 7 -r :w... *4 .*.






. .. . .- ;
..** ." 1 i

I.:, at r1


*r1 t. i


, j -P : ": t \, *" "',















*i * ff' -
- i
s'- -"r :"Y 5 ..* ,-.
; - .. ".




*. " -




6 61
+,+' , ) -A

"* * .,















EXPERIMENT II


Experimental Procedures


Animals

Eighteen Dorset or Hampshire x Rambouilet crossbred rams

approximately 11 months old and weighing an average of 99 pounds were

used in this experiment. The rams were purchased from Auburn Univer-

sity in Alabama, and brought to the Experiment Station as three-month-

old weaned lambs.

The rams were used in preliminary studies with purified rations

prior to the start of the present experiment. The rams were all placed

on a basal purified ration several weeks before the experiment was

initiated to minimize any effect of previous treatment.

During the preliminary feeding period, three semen collections

were made. The average total sperm production value for each ram

from these three collections was used as the basis for assignment to

treatment groups. The 18 rams were ranked according to total sperm

production and the top, middle and low ramspulled out. The remaining

15 rams were then stratified into three uniform groups of five rams

each. A protein-nitrogen ration, a urea-nitrogen ration and a nitrogen-

free ration were assigned at random to the three groups of rams. The

three rams pulled out during the ranking were then added to the group

receiving the nitrogen-free ration. In this manner two groups of five










rams and a group of eight rams were obtained that were relatively

uniform in total sperm production.

At the start of the experiment on September 1, 1960, the

average initial weights of the three groups were 97.6, 92.5 and 100.2

pounds for the urea-nitrogen, protein-nitrogen and nitrogen-free rams,

respectively.


Facilities

The rams were maintained in individual stalls in metal metab-

olism racks under a shed roof at the Experiment Station Sheep Unit.

Each stall was equipped with a feed and a water container. Platform

scales were available at the unit. Pens and chutes for working the

rams were adjacent to the shed area. A small, portable stanchion was

constructed to secure an "estrogenized" ewe for libido checking.


Feedstuffs and Rations

Purified ingredients were used in the experimental rations.

Drackett C-1 assay protein, a purified soybean protein, was used as

the protein source and urea as the non-protein source of nitrogen.

The composition of the two control rations and the nitrogen-free

ration is given in table 9. The levels of Drackett protein and urea

were calculated to supply approximately 11 percent crude protein in

each control ration. The methionine analogue was added to the control

rations as a source of sulfur containing amino acid. The total pro-

tein content was 11.54 and 11.05 for the Drackett and urea rations,

respectively. The corn sugar and corn starch levels were increased











in the nitrogen-free ration to replace the nitrogen source. Chemical

analysis indicated essentially no nitrogen in the nitrogen-free ration.

Four percent corn oil was added to all three rations to reduce the

dustiness and improve the texture and palatability.

The mineral mixture used in the rations is shown in table 9.

The mixture containing the 13 essential mineral elements was essentially

that reported by Oltjen et al. (1959).

Vitamins A, D, E and choline in a corn sugar carrier were

added to each ration. Table 9 gives the vitamins and the levels

at which they were fed.

The rations were mixed in 100-pound lots in a stainless steel

mixer. The mixed rations were stored under refrigeration at approx-

imately 400 F.


Procedures

Management and Feeding. The rams were fed twice daily all

the feed they would clean up by the next feeding. Feeding was done

at 7:00 a.m. and 5:00 p.m. Feed refused was weighed back daily at

the evening feeding. Fresh water was available at all times. The

rams were maintained in the individual stalls at all times except

when removed to collect experimental data.

Feed consumption data were summarized individually every seven

days and by treatment groups for 28-day periods.

Weighing. The rams were weighed every seven days on platform

scales. Individual and group average daily gains were summarized for














COMPOSITION OF THE
IN


TABLE 9

PURIFIED RATIONS
EXPERIMENT II


FED THE RAMS


Ingredients


Cellulosea

Corn Starch, granulated

Corn Sugar

Corn Oil

Drackett Protein

Urea (262 Protein)

Methionine (Hydroxy analogue)

Mineral Mixtureb

Vitamin Mixturec


Drackett
Control
(lbs.)

50.00

23.00

15.45

4.00

15.00



0.55

10.00

4.00
100.00


Rations
Urea
Control
(lbs.)

50.00

28.00

19.45

4.00



4.00

0.55

10.00

4.00
100.00


aHydrolyzed wood pulp, Solka Floc, was used as a source of
cellulose.
bThe mineral mixture supplied the following amounts of the
13 essential minerals per 100 pounds of feed: CaHP04*2H20, 1444.2 gm.;
1H
K2C03'1~H20, 950.7 gm.; MgSO43H20, 316.9 gm.; NaC1, 227 gm.; FeSO4
2H20, 59.8 gm.; Na2B407lOH20, 5.6 gm.; MnSO4*H20, 2.9 gm.; ZnS04-7H20,
2.0 gm.; KI, 0.88 gi.; CuSO4, 1.18 gm.; 2CoC03*5Co(0H)2, 0.008 gm.;
MoOg, 0.025 gm.; Corn sugar carrier, 1568.90 gm.
CThe vitamin mixture supplied 100,000 IU of vitamin A palmitate,
15,000 ICU of vitamin Dg, 200,000 units of vitamin E, 200 gm. of choline
chloride (254 in carrier) and corn sugar carrier to make four pounds per
100 pounds of feed. The vitamins were furnished by Chas. Pfizer and Co.,
Terre Haute, Indiana.


Nitrogen
Free
(lbs.)

50.00

30.00

22.00

4.00







10.00

4.00
100.00











each seven and 28-day period. The weighing was done in the morning

before the rams were fed.

Blood Evaluation. Procedures followed in the blood evaluation

were similar to those used in Experiment I. Blood samples were obtained

at 28-day intervals in Experiment II.

Libido Studies. Libido evaluations were conducted in the same

manner as in Experiment I.

Semen Evaluation. The procedures followed in the routine

semen evaluation were basically the same as those of Experiment I.

Fertility and semen exhaustion studies were not conducted in Exper-

iment II. Single ejaculates per ram were collected in the second

experiment.

SHistological Investigations. The same procedures used in

Experiment I were employed in Experiment II. The prostate gland,

however, was not used in these investigations in Experiment II.

Adenohypophysis Assay. The gonadotrophic activity of the

adenohypophyses from the three groups of rams was determined by the

chick testes weight technique. The gonadotrophic assay was performed

using day-old White Leghorn chicks injected with a total of 10 mg.

of dried adenohypophysis powder in a saline solution.

Due to the small number of chicks that could be used on

individual glands, the powder from rams within ration groups was

pooled. Twelve, 14 and 15 chicks were used for the urea, Drackett

and nitrogen-free groups, respectively. The 10 mg. of dried powder

in 0.75 ml. of 0.9 percent saline solution was injected subcutaneously











in three injections 12 hours apart. Four groups of 10 chicks each

received 250, 500, 750 and 1000 IU of A standard gonadotrophin'.

An additional group of 10 chicks received saline injections only and

a control group of 10 chicks received no injections.

Sixteen hours after the last injections, the chicks were

weighed and sacrificed. The testes were removed and wet weights of

both testes were taken. The degree of stimulation from the adenohypo-

physis powder was determined by comparing the testes weights of the

test chicks with those of the chicks injected with saline only. An

approximation of the potency of the adenohypophysis powder from the

treatment groups was estimated from the curve established with the

testes weights of the chicks injected with the graded doses of the

standard gonadotrophin.

Statistical Analysis. The blood, semen, and libido data were

subjected to the same statistical analysis used in Experiment I. In

this experiment, however, there were only four periods in phase I and

a single period in phase II. The analysis used for phase II involved

comparisons of values for the various criteria for the four surviving

rams in the nitrogen-free group.











Results and Discussion


Feed Intake

The purified rations fed in the second experiment were not

consumed readily by any of the treatment groups. One ram in the

Drackett control group and one in the urea control group would not

eat the ration and had to be removed from the experiment. The

Drackett control, urea control and the nitrogen-free groups consumed

an average of only 1.60, 1.46 and 0.61 pounds of feed per day,

respectively, during the experiment. There was a great deal of var-

iation in feed intake between rams in the control groups as indicated

in table 19 in the Appendix. Feed intake in the nitrogen-free group

was more uniform, possibly due to the rather severe restriction

brought about by the lack of nitrogen. The reason for the limited

feed intake of the two control rations is not immediately apparent.

It may have been related to the bulky nature of the ration or to some

environmental factor associated with the experiment.

While the two control groups consumed fairly similar amounts

of feed the lack of nitrogen had a marked and immediate effect on the

appetite of the rams fed the nitrogen-free ration. Intake dropped off

the first week of feeding, and continued to decline throughout the

study. The feed intake pattern for the three treatment groups is

given in figure 15. The effect of a complete absence of nitrogen

was even more severe than that found with the low protein ration in

Experiment I and earlier work reported by the author (Meacham, 1960).

Four of the eight rams on the nitrogen-free ration died or were






















N
N
N


N


N - - -
.. -


DRACKETT CONTROL
UREA CONTROL
NITROGEN-FREE


2.00



1.75


1.50


1.25


1.00



.75


.50



.25


5%.


I -I I__J I -II
1 2 3 4 5 6 7
14-DAY PERIODS

Figure 13. Effect of source and level of nitrogen on feed intake of rams.


--- -'
,,,.,










sacrificed an average of 78 days after the start of the study. In

Experiment I, the low protein bulls survived 120 days on the low

protein ration before the first bull died. The nitrogen-free rams

consumed an average of 40 percent of the amount consumed by the con-

trol rams whereas the low protein bulls in Experiment I ate only 33

percent as much as the control bulls.

There was little difference in feed intake between the two

sources of nitrogen, Drackett protein and urea. The average intake

of the urea rams tended to be lower during the first part of the study.

This was due in part to the performance of one ram in this group that,

after starting on the ration, became very erratic in feed consumption

and finally went completely off-feed. This ram, no. 19, was subse-

quently sacrificed after 84 days on the experiment.

This feed consumption response is somewhat at variance with

that observed in earlier work with these rations by the author

(Meacham et al., 1961) and work by others (Oltjen et al., 1959) with

these same sources of nitrogen. The performance of the protein-

nitrogen was usually much better than that of the non-protein-nitrogen.


Body Weight Changes

The average daily gains of the treatment groups reflected

very closely their respective feed intakes. Figure 14 shows the

cumulative ADG for the three groups. The Drackett control group had

an average gain of 0.09 pounds daily for the experimental period.

The urea control group lost considerable weight during the second

period and then gradually recovered, finishing the experiment with a






















/


DRACKETT CONTROL

UREA CONTROL

-.. NITROGEN-FREE


. .-


Figure 14.


14-DAY PERIODS

Effect of source and level of nitrogen on cumulative average daily gain
of rams.


.10 L


.10


.20


.30


-.40


0


_ .- -











cumulative ADG of 0.06 pounds. As was the case with feed intake for

the urea control group, ram no. 19 tended to hold the group average

down. This ram lost 42 pounds in the 84 days he was on the experiment.

The other rams in the group held their initial weight or gained

slightly. These individual weight changes are summarized by treat-

ment group and period in table 20 in the Appendix.

The nitrogen-free group lost weight consistently throughout

the study. This was not unexpected in view of their reduced feed

intake. The four rams surviving the 98-day experimental period lost

an average of 37 pounds or over one third of their initial weight.

The weight losses for all eight rams in this group tended to be

fairly uniform. The cumulative ADG for this group was -.58 pounds.

Figure 15 illustrates the appearance of these rams near the

end of the experiment.


Blood Composition

Hemoglobin. The effect of the three experimental rations on

blood hemoglobin levels in the rams is shown in figure 16. The mean

hemoglobin value for the nitrogen-free rams dropped rapidly during

the second and third periods and then tended to level off. The two

control groups, however, increased slightly during the second period

and then dropped during the third period with a similar leveling off

for the remainder of the study.

While the hemoglobin of the two control groups tended to be

higher than that of the nitrogen-free group during the experiment,























Figure 15. Effect of source and level of nitrogen on ram growth.

A, B and C: A representative Drackett control, urea
control and nitrogen-free ram, respectively, at the
end of the experiment.






























































ar)
















-
N---


DRACKETT CONTROL
_- UREA CONTROL
S .- NITROGEN-FREE


Initial


1 2
28-DAY PERIODS


Figure 16. Effect of source and level of nitrogen on mean
hemoglobin values for rams.


aPeriod 4 only seven days.


15.0 L


12.0 L


11. L


10.0 1


- -


9.01


8.0L


I -


c. --
\










the differences were not statistically significant. The lack of

statistical significance was due primarily to the wide variation in

hemoglobin levels within treatment groups.

Initial and final mean hemoglobin values for the Drackett

control, urea control and nitrogen-free groups were 11.06 and 11.09,

12.02 and 10.05 and 12.74 and 8.74 grams of hemoglobin per 100 ml. of

blood, respectively. It would appear that had the nitrogen-free rams

survived longer, real differences might have developed.

Individual and treatment group blood values for the five

periods are summarized in table 21 in the Appendix.

Hematocrit. As indicated in figure 17, hematocrit values for

the three treatment groups fluctuated markedly. Statistical analysis

of the data revealed no significant differences due to treatment or

period. The urea-control and the nitrogen-free groups tended to fluc-

tuate together and both were somewhat lower than the Drackett controls

during the last part of the study.

Mean hematocrit values for the initial and final periods were

44.06 and 42.87, 44.22 and 37.80 and 48.34 and 55.88 percent, respec-

tively, for the Drackett control, urea control and the nitrogen-free

groups. All groups, however, were well above the normal hematocrit

figure of 32 percent reported by Dukes (1955).


Serum Protein. Serum protein values varied somewhat between

periods during the experiment, but practically no difference was ob-

served between treatment groups. This situation is shown graphically

in figure 18. Statistical'analysis of the serum protein data failed













/ \


/




/
/


/ \

/ /\ \\
,/ / \ \
\* \\
/


DEACKETT CONTROL
UREA CONTROL
NITROGEN-FREE \/


Initial


22
28-DAY PERIODS


Figure 17. Effect of source and level of nitrogen on mean
hematocrit levels of rams.
Period 4 only seven days.


55


51 1


47L


391.


35 L


r


4


I I_ r ---


--



i















9.0 .





S8.0 /





7.0



DRACKETT CONTROL
6.0 UREA CONTROL
-.-.-. NITROGEN-FREE

01 1 a I I.
Initial I 2 3 4a
28-DAY PERIODS

Figure 18. Effect of source and level of nitrogen on mean
serum protein values of rams.

aPeriod.4 is a seven-day period.











to indicate any treatment effect also. Apparently, the nitrogen-free

rams were maintaining blood protein levels through tissue breakdown.

The initial and final serum protein values for the three groups were

7.77 and 7.36, 7.81 and 7.44,and 7.78 and 6.77 percent for the Drackett

control, the urea control and the nitrogen-free groups, respectively.

Analysis of Variance mean squares for the various blood com-

ponents are given in table 27 in the Appendix.


Libido Evaluation

The three dietary treatments had very little effect on the

libido of the rams. No significant differences were noted for the

interval to first mount, interval to first mating or libido score.

As indicated in table 10, the Drackett control and the nitrogen-free

rams tended to have shorter mean intervals to first mount and mating

and higher libido scores than the urea control rams. Two of the four

rams in the urea group, nos. 19 and 14, displayed little or no interest

in the estrogenized ewe throughout the trial. No. 14 did mate the ewe

once near the end of the study. These two rams were undoubtedly

responsible for most of the difference between the libido performance

of the urea control group and the other two groups. One of the

nitrogen-free rams also showed no interest in the ewe during the ex-

periment.

Comparing the effect of low protein rations on the libido of

bulls with the effect of nitrogen-free rations on rams, reveals a pos-

sible species difference. In Experiment I and earlier work by the

author (Meacham, 1960) libido was adversely affected in the protein










TABLE 10


SUMMARY OF LIBIDO DATA FOR THE THREE TREATMENT GROUPS OF RAMS
BY 14-DAY PERIODS


Periods
Criteria and Periods
Treatment Groups-- 1 2 3 4 5 6
Interval to First Mount, min.a
Drackett Control 1.0 .5 .8 .5 .5 .5
Urea Control 5.9 5.2 5.2 5.6 5.0 .8b
Nitrogen-free 1.8 1.6 1.7 1.6 1.6 2.9c

Interval to First Mating, min.a
Drackett Control 1.4 1.1 1.5 .9 2.8 .9
Urea Control 5.3 6.1 8.0 8.0 3.8 4.6b
Nitrogen-free 2.2 1.9 .9 7.2 5.5 5.9c

Libido Scored
Drackett Control 5.0 5.0 5.0 5.0 4.8 5.0
Urea Control 3.5 5.0 2.8 3.0 4.2 4.7b
Nitrogen-free 4.5 4.6 4.5 4.4 4.6 4.2c


aRams allowed a total of 10 minutes with the ewe; rams
mating in 10 minutes received a value of 10.
bMean of three animals only in period 6.
cMean of four animals only in period 6.
dLibido score based on a 1 to 5 scale; 1, no interest;
as described on page 20.


not mounting or



5, mated ewe,











depleted bulls, while a complete lack of nitrogen had little or no

effect on the libido of these rams. Even the four rams that did not

finish the experiment mated the ewe at the last libido check before

they died or were sacrificed.

The obvious physical weakness of the nitrogen-free rams did not

have the same deterring effect on the libido that was observed in the

low protein bulls in Experiment I.

Analysis of Variance mean squares for the various libido criteria

are given in table 28 in the Appendix.


Semen Characteristics

Volume. The mean volume of semen produced by the urea control and

nitrogen-free groups was quite similar until the latter part of the study

when the nitrogen-free group produced slightly smaller amounts. There

was an appreciable drop in volume during period 2, followed by a more uni-

form production for both groups. These changes are shown in figure 19.

The mean for the Drackett control group, on the other hand, fluctuated

widely, being both above and below the other two groups.

Statistical analysis of these data indicated no significant

difference between treatment groups. There was a significantly

greater volume (P < .01) produced in period 1 as compared to the

other periods. Lack of a significant interaction indicated that the

higher volume in period 1 was consistent for all groups. Initial and

final semen volume figures for the three groups were 1.28 and 1.38,

1.85 and 1.12, and 1.45 and 1.12 ml. per ejaculate for the Drackett

control, urea control and nitrogen-free groups, respectively.














DRACKETT CONTROL
- - UREA CONTROL
- -.- NITROGEN-FREE


1.0 .


.8 1


1 2 5 4 5 6
14-DAY PERIODS


Figure 19.


Effect of level and source of nitrogen on mean semen volume
of rams.


2.0


1.8


1.6


1.4


1.2











Semen volume, motility and total cell production figures for

individual rams and treatment groups are summarized in table 22 in

the Appendix.

Motility. Mean estimates of the percent motile sperm for the

three groups of rams are shown in figure 20. There was a character-

istic decline for all groups as the experiment progressed. This

decline was due primarily to the progressively colder environmental

temperatures as the experiment went into the winter months. The semen

collections were made in an exposed area and the cold shock had a

marked influence on the motility scores.

In view of this environmental effect, the motility values are

not too meaningful except from a relative standpoint. The average

motility for the nitrogen-free group tended to drop more than that

of the control group, but the differences were not statistically

significant. There was a significant period effect as would be ex-

pected. The motility of all groups was lower during the latter part

of the experiment. A great deal of variation was observed within

rams between collections and within treatment groups for any one

collection. This would tend to preclude assigning any statistical

significance to the apparent differences also.

Total Sperm Cell Production. Subjecting the total sperm cell

production data to a statistical analysis revealed no significant

differences due to treatment or period. Again, the small number of

animals involved and the considerable variation within treatment

groups tend to minimize the treatment effects.















80 L


o %


I'
/
/


.0 N


* -,


/
\~ /


DRACKETT CONTROL
UREA CONTROL
NITROGEN-FREE


1 2 3 4 5 6
14-DAY PERIODS


Figure 20.


Effect of level and source of nitrogen on mean semen motility
of rams.


\ \


. -.---











The pattern of total sperm cell production for the groups is

presented in figure 21. While there was considerable fluctuation in

the two control groups, the nitrogen-free rams had a fairly uniform

production. The larger number of animals in this group could explain

part of this uniformity. The total cell production of the urea control

group appeared to be consistently higher than the others and the nitro-

gen-free group generally lower than the two control groups. This was

particularly true during the last part of the study.

The average total cells per ejaculate for the experimental

period for the three treatment groups was 2609, 4660, and 1665 times

106 sperm for the Drackett control, the urea control and the nitrogen-

free rams, respectively. In spite of the lack of statistical evidence,

there appears to be some treatment effect on the total cell production,

particularly when the urea control and nitrogen-free groups are

compared.

Analysis of Variance mean squares for the semen characteristics

are given in table 28 in the Appendix.


Histological Investigations.

Microscopic examination of the reproductive organs and tissues

did not reveal any marked differences due to treatments. Measurements

of the epithelial tissue, tubule diameters and secretary areas of the

tissues studied are given in tables 11 and 12. There was some reduc-

tion in the number of layers of germ cells in the seminiferous tubules

of the testes from the nitrogen-free rams. Cellular disorganization

was observed in the seminal vesicles of several of these nitrogen-free

rams.












DRACKETT CONTROL
UREA CONTROL
SNITROGEN-FREE


/


N
N
N


N
'N
N


I I
1 2


3 4
14-DAY PERIODS


Figure 21.


Effect of
of rams.


source and level of nitrogen on total cell production


6000 L


5000 L


4000 1


3000 L


2000 .


1000 L


[] I, I


!







TABLE 11


QUANTITATIVE EVALUATION OF THE TESTES AND EPIDIDYMIS FROM THE THREE GROUPS OF RAMS
Testes Epididymis
Treatment Epithelium Tubule Inter- Epithelium Duct Sperm
Group and Thickness Diameter Sperma stitialb Thickness Diameter Content
Animal No. (rmm x 10-2) (rmm) Content Tissue (mm x 10-2) (mm)
Drackett Control
2 3.0 .10 1 3 6.0 .20 1
5 4.8 .17 4 2 6.0 .28 5
9 6.0 .17 3 1 7.2 .35 5
17 4.2 .13 1 1 6.0 .22 5
Av. 4.5 .14 2.2 1.7 6.5 .26 3.5
Urea Control
4 3.6 .17 4 2 7.2 .28 2
13 3.6 .14 1 2 6.5 .27 4
14 4.8 .13 1 2 7.8 .32 5
19 3.6 .11 1 4 4.8 .28 1
Av. 3.9 .14 1.8 2.2 6.6 .29 3.0
Nitrogen-free
1 4.8 .11 1 3 4.8 .22 1
6 3.6 .10 1 5 4.8 .20 1
7 3.6 .10 1 4 7.2 .28 2
8 4.8 .11 1 5 6.0 .22 1
15 4.8 .12 1 3 6.6 .25 1
16 4.8 .12 1 2 4.8 .24 1
18 4.2 .10 1 4 4.2 .24 1
20 6.0 .12 1 2 6.6 .28 2
Av. 4.6 .11 1 3.2 5.6 .24 1.2


aSperm content scored from 1 to 5: 1, no sperm, and 5, abundant.
bInterstitial tissue scored from 1 to 5; 1, sparse, and 5, abundant.







TABLE 12


QUANTITATIVE EVALUATION OF THE SEMINAL VESICLES AND
FROM THE THREE GROUPS OF RAMS


COWPEIS GLANDS


Seminal Vesicles Cowpers Glands
Treatment Epithelial Rate of Epithelial
Group and Thickness Secretory Tissuea Thickness
Animal No. (mm x 10-2) to Total Area (mm x 10-2)
Drackett Control
2 1.8 1 1.2
5 1.2 5 1.5
9 5.0 5 1.2
17 5.0 2 1.2
Av. 2.2 2.2 1.3
Urea Control
4 2.1 .2 1.8
15 1.8 1 1.5
14 1.8 2 1.8
19 1.8 2 0.9
Av. 1.9 1.7 1.5
Nitrogen-free
1 1.2 2 .9
6 1.2 1 .6
7 1.8 3 --b
8 1.8 1 1.5
15 2.1 1 --b
16 1.5 2 .6
18 1.8 1 .9
20 1.8 1 .9
Av. 1.6 1.6 .9

aScale is based on values from 1 to 5, 1 being a wide ratio of
secretory tissue to total area and 5 being a narrow ratio.
"Samples lost in processing.











Absence of any major differences in these testes and secondary

glands is not completely without explanation. In fact, it would be

difficult to explain differences had they occurred. Both libido and

secondary sex gland development and activity are controlled by testos-

terone secretion from the testes primarily. Since the nitrogen-free

ration had little or no effect on libido, differences of any magnitude

would not be expected in these glands.

The results here are somewhat different from those obtained in

Experiment I. With the low protein bulls, both libido and histological

differences were observed. Again, it is possible that a species dif-

ference may exist.

The gross weights of these glands and organs were obtained at

time of slaughter. The gross weights, along with the weights per cwt.

of body weight, are summarized by treatment groups in table 13. Very

little difference is noted between treatment groups when the weights

per cwt. of body weight are considered.


Adenohypophysis Assay

The results of the adenohypophysis assay are given in table 14.

The Drackett control and the nitrogen-free groupswere identical in

gonadotrophic activity according to the assay. The urea control group

was slightly lower than the other groups. When the extremely wide

ranges in chick testesweights for any one group are considered, it

must be concluded that there was no real difference between the three

treatment groups in gonadotrophic activity.










TABLE 13


SUMMARY OF THE MEAN GLAND AND ORGAN WEIGHTS TAKEN AT TIME OF SLAUGHTER
FROM THE THREE GROUPS OF RAMS


Weights (grams)
Drackett Control Urea Control Nitrogen-free
Gland or Organ Per Cwt. Per Cwt, Per Cwt.
Mean Body Weight Mean Body Weight Mean Body Weight

Testes 221.97 220.0 168.12 163.00 102.47 163.00
Seminal Vesicle 8.10 8.0 7.70 7.50 4.77 7.22
Cowper's Gland 2.70 2.6 3.22 3.10 1.85 2.80
Pituitary Gland .76 .75 .65 .65 .51 .81
Thyroid Gland 3.62b 5.60 4.07c 4.00 3.77 6.00


aMean of four samples only.
bMean of three samples only.
CMean of two samples only.











This lack of treatment effect is quite comparable with the

lack of treatment effect on libido, reproductive tissue histology and

semen production. The gonadotrophic hormones, Follicle Stimulating

Hormone (FSH) and Leutinizing Hormone (LH), from the adenohypophysis

regulate semen production and testosterone production which in turn

regulates libido and activity of the secondary sex glands.



TABLE 14

BIOLOGICAL ASSAY OF ADENOHYPOPHYSES FROM
THE THREE TREATMENT GROUPS OF RAMS


Groups


No. of
Chicks


Non-injected Chicks
Saline-injected Chicks

Standard
250 IUa
500 IU
750 IU
1000 IU

Ram Treatment Groups
Drackett Control
Urea Control
Nitrogen-free


Mean Chick
Testes Weight
(mg.)
5.8
5.0


9.9
12.5
11.6
16.2


S12.0
9.9
12.0


Range in
Testes Weight
(mg.)
5.6 9.0
1.6 7.2


5.6 15.7
6.2 10.4
6.4 21.1
12.2 21.1


4.0 19.8
6.9 19.0
5.9 19.1


aStandard used was unfractionated sheep pituitary powder
obtained from Abbott Laboratory.














GENERAL DISCUSSION AND CONCLUSIONS


Before discussing the effects of the experimental treatments

on the growth and reproductive performance of the bulls and rams, it

is necessary to consider the fact that a confounding multiple defi-

ciency condition existed in both experiments. The depressing effect

of low protein and nitrogen-free rations on feed intake created defi-

ciencies of energy, vitamins, minerals and other dietary factors, as

well as a nitrogen or protein deficiency. It should be stated, how-

ever, that these deficient rations would have supplied the required

amounts of these other nutrients had they been consumed at the same

level as the control rations. The basic cause of this multiple defi-

ciency is therefore, a protein or nitrogen deficiency.

Comparing the effects of the low protein ration on the per-

formance of the bulls in Experiment I with those of the nitrogen-free

ration in Experiment II on ram performance reveals several notable

similarities and several striking differences.

The immediate response in both experiments to a low level or

absence of nitrogen was a pronounced drop in feed intake. As a result

of this reduction in feed intake, losses in body weight rapidly

developed. This response is in good agreement with earlier work by

the author (Meacham, 1960) and that of Guilbert and Hart (1951).

Under the conditions of these experiments, it may be concluded that











low levels or complete absence of nitrogen will result in marked

reduction in feed intake and losses in body weight.

In Experiment II, it appeared the source of nitrogen, whether

it was of a protein or non-protein source, did not have an appreciable

effect on feed intake or weight gains. These findings are not in com-

plete agreement with the work of other investigators (Meacham et al.,

1961, and Oltjen et al., 1959) who reported that a protein source of

nitrogen was more effective than a non-protein source. It is possible,

however, that the conditions of the experiment did not permit full

manifestation of the potential differences.

The experimental rations affected the blood composition of

the bulls and rams to differing extents. Bulls receiving the low

protein ration in Experiment I had significantly lower hemoglobin,

hematocrit and serum protein levels by the end of the experiment than

the control animals. The rams receiving the nitrogen-free ration in

Experiment II did not, on the other hand, differ significantly from

the control rams in these blood components. The nitrogen-free rams

did tend to be lower in all three components by the end of the trial.

The differences in length of experimental periods, 98 days in Exper-

iment II and 182 days in Experiment I, undoubtedly played a part in

this differential response. It should be pointed out, however, that

in both experiments hemoglobin and hematocrit levels for all treatment

groups were near the normal values reported in the literature. The

differences observed in Experiment I were usually due to increases in

the control levels rather than marked decreases in the low protein











group. The serum protein levels at the end of the first experiment

for the low protein bulls were slightly below the normal levels

reported in the literature.

The two sources of nitrogen used in Experiment II were fairly

comparable in maintaining normal blood composition. The urea tended

to be slightly less effective in this respect than the protein source,

but the differences were not statistically significant.

The effect of nitrogen deprivation on libido presented one

of the more striking differences between the two experiments. The low

protein ration in Experiment I brought about a significant decline in

libido whereas the nitrogen-free ration had very little effect on the

libido of the rams in Experiment II. Keeping in mind the small num-

bers of animals involved, there may be a species difference in the

effect of a nutritional stress on the libido of rams and bulls.

This lack of effect on the libido of the rams was confirmed

by a similar lack of differences in the histology of the primary and

secondary sex glands and in the gonadotrophic activity of the adenohy-

pophysis of the three groups of rams. The histological changes ob-

served in the reproductive tissues of the low protein bulls in turn

confirmed the differences observed in the libido of these bulls.

The lack of protein had rather distinct effects on these tissues.

No real difference was obtained between the two sources of

nitrogen in the histology study with the rams. On the basis of these

limited data, it appears that non-protein nitrogen can be effectively

utilized for these reproductive tissues.










A comparison of the semen production in the two experiments

revealed a rather similar treatment response. In both cases, the

deprived group tended to be lower in volume, motility and total sperm

cell production. The differences in volume and total cell production

were significant for the bulls in Experiment I, but not for the rams

in Experiment II. The nitrogen-free rams were sufficiently low by

the end of the study to indicate a possible treatment effect had the

numbers in each group been larger.

The results of these experiments are basically in agreement

with the limited data available in the literature. Mann and Walton

(1953) stated that severe starvation resulted in only reduced volume

of semen per ejaculate in a mature bull. Dutt and Barnhart (1959) and

Steverman et al. (1961) working with boars and Baker et al. (1953) and

Flipse (1957) working with dairy bulls, all reported little effect on

semen characteristics other than semen volume in animals restricted

in feed intake.

In Experiment II, the non-protein-nitrogen had no adverse

effect on any semen characteristic. Moreover, the total cell pro-

duction for this group was generally higher than that of the other

two groups. On the basis of these data, it could be concluded that

the ram is able to utilize non-protein nitrogen adequately for sperm-

atogenesis and semen production.















SUMMARY


Two experiments were conducted to study the effect of source

and level of nitrogen on the growth and reproductive performance of

young bulls and rams.

In Experiment I, 12 bulls averaging 390 pounds were allowed

to two groups of four and eight bulls each as they reached puberty.

The four bulls received a ration adequate in all nutrients. The

eight bulls were fed a ration containing 1.61percent CP, but other-

wise adequate. Feed intake, weight changes, blood composition,

semen characteristics, libido and reproductive tissue histology were

studied.

The low protein ration depressed appetite severely. Average

daily feed consumption was 9.78 and 3.28 pounds for the control and

low protein bulls, respectively.

At the end of the 182-day experiment, the control bulls had

gained 200 pounds and the low protein bulls had lost an average of

130 pounds. Four of the low protein bulls died or were sacrificed

during the experiment. Average survival time was 151 days.

The low protein bulls had final blood hemoglobin, hematocrit

and serum protein levels significantly (P < .01) lower than the

controls.

Libido was adversely affected by the low protein ration. The

low protein bulls had significantly longer intervals to first mating

and lower libido scores (P < .01).











Semen volume (P < .01) and total sperm cells per ejaculate

(P < .05) were significantly lower for the low protein bulls. Sperm

motility was quite varied and not significantly affected by treatments.

Artificial insemination of heifers revealed no treatment effect

on semen fertility.

The semen exhaustion study indicated reduced spermatogenic

capacity in the low protein bulls. The controls produced greater

volumes of semen containing greater numbers of sperm.

Histological investigation of the reproductive organs and

glands revealed marked differences in size and development. The thick-

ness of the epithelium in the glands and organs of the low protein

bulls was greatly reduced. Tubule diameters in the testes and epididy-

mis were similarly reduced. The control bulls tended to have greater

amounts of interstitial tissue.

In Experiment II, 16 crossbred rams weighing 99 pounds were

assigned to three purified rations. Two groups of four rams each

received control rations, one containing protein-nitrogen and the other

non-protein-nitrogen. Eight rams received a nitrogen-free ration.

The experimental period was 98 days. The same criteria studied in

Experiment I were studied in Experiment II.

The nitrogen-free ration brought about marked decreases in

feed intake. Average intakes were 1.60, 1.46 and 0.61 pounds daily

for the Drackett control, urea control and nitrogen-free rams,

respectively.











ADG were 0.09, 0.06 and -.58 pounds for the Drackett control,

urea control and nitrogen-free groups, respectively. Four of the

nitrogen-free rams died or were sacrificed during the experiment.

Mean survival time was 78 days. One urea control ram died after

84 days.

Blood components were not significantly affected by the treat-

ments. All groups were within the normal ranges.

The libido criteria were not significantly different for any

of the treatment groups.

Semen volume, motility and total sperm production were not

affected significantly by the treatments. The urea control group

tended to have the higher total sperm production average and the

nitrogen-free group the lower average.

Histological examination of the reproductive tissues revealed

very little treatment effect. There was some reduction of epithelium

in the seminiferous tubules of the nitrogen-free ram testes.

The gonadotrophic activity of the adenohypophyses of the rams

was not affected by the treatments.

The differential response to the deficient rations of the bulls

and rams was due to the shorter experimental period for the rams, 98

versus 182 days. The depleted rams would die before libido, blood

components and tissue histology were affected.















LITERATURE CITED


Armsby, H. P. 1917. The Nutrition of Farm Animals. Macmillan Book
Company, New York, N.Y.

Association of Official Agricultural Chemists. 1950. Official Methods
of Analysis. Eighth Edition.

Baker, F. N., N. L. Van Demark and G. W. Salisbury. 1955. A year's
study of the semen characteristics and libido of young bulls
subjected to various frequenciesof ejaculation. Journal of
Animal Science, 12:942.

1955a. Growth of Holstein bulls and its relation to sperm
production. Journal of Animal Science. 14:5 75.

1955b. The effect of frequency of ejaculation on sperm
production. Journal of Dairy Science. 38:1000.

Bedrak, E. 1958. Effect of protein intake on weight changes, blood
constituents and reproduction in beef heifers. Ph.D. disser-
tation. Department of Animal Husbandry, University of Florida.

Berg, C. P. and W. G. Rohse. 1947. Is sterility induced in growing
rats on a tryptophan deficient diet? Science. 105:96.

Blaxter, K. L. and H. H. Mitchell. 1948. Protein requirements of
ruminants. Journal of Animal Science. 7:25.

Blincoe, C. and S. Brody. 1951. Environmental physiology with special
reference to domestic animals. XVII. The influence of tem-
perature on blood composition of cattle. University of Missouri
Research Bulletin No. 488.

Blucker, J. D. 1952. Mineral and management studied with range beef
cattle. Master of Science Thesis, Oklahoma Agricultural and
Mechanical College, Stillwater, Oklahoma.

Branton, C., R. W. Bratton and G. W. Salisbury. 1948. Semen produc-
tion and fertility of dairy bulls fed rations containing
proteins of plant and animal sources. Journal of Dairy
Science. 34:292.




University of Florida Home Page
© 2004 - 2010 University of Florida George A. Smathers Libraries.
All rights reserved.

Acceptable Use, Copyright, and Disclaimer Statement
Last updated October 10, 2010 - - mvs