The relationship of carbohydrate reserves to the quality of bare-root Pinus elliottii var. elliottii (Engelm.) seedlings...

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Title:
The relationship of carbohydrate reserves to the quality of bare-root Pinus elliottii var. elliottii (Engelm.) seedlings produced in a northern Florida nursery
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x, 146 leaves : ill. ; 28 cm.
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McNabb, Kenneth Lee, 1949-
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Slash pine -- Seedlings -- Quality -- Florida   ( lcsh )
Carbohydrates   ( lcsh )
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non-fiction   ( marcgt )

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Thesis:
Thesis (Ph. D.)--University of Florida, 1985.
Bibliography:
Includes bibliographical references (leaves 137-145).
Statement of Responsibility:
by Kenneth Lee McNabb.
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Typescript.
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Vita.

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THE RELATIONSHIP OF CARBOHYDRATE RESERVES TO
THE QUALITY OF BARE-ROOT Pinus elliottii
var. elliottii (Engelm.) SEEDLINGS PRODUCED
IN A NORTHERN FLORIDA NURSERY






By

KENNETH LEE McNABB


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



UNIVERSITY OF FLORIDA


1985











ACKNOWLEDGEMENTS


I want to especially thank my major advisor, Dr. Earl Stone.

He impressed upon me the art of logical writing, an enthusiasm

for discovery, and to never forget the basics. Also, I wish to

thank the other members of my committee; Drs. Ed Barnard, Sue

Kossuth, and Sherlie West, for their encouragement and critical

evaluation of my work. The contributions of Drs. Jon Johnson and

Tom Humphries are also appreciated. And I am grateful for the

constant encouragement and financial support of Dr. Arnett Mace.



I am deeply endebted to Sarah Mesa and Joel Smith for

laboratory and field assistance over the past several years.

Their personal support was also invaluable.



I would like to thank Dale Rye of the Container Corporation

Nursery in Archer, Florida. His cooperation and attitude to

nursery research is exemplary.



Last but certainly not least, I wish to thank my family.

The support of my wife Miriam throughout the course of these

studies has been essential. Finally, I want to acknowledge my

children: they gave me the purpose for which it was done.












TABLE OF CONTENTS

ACKNOWLEDGEMENTS...................... .......... ....... ii

LIST OF TABLES. ...................................... ..... v

LIST OF FIGURES.............................................. vi

ABSTRACT..................................................... ix

GENERAL INTRODUCTION......................................... 1

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

Introduction.................... ................. .. 5
Lateral Root Formation............................... 6
Seasonality of Carbohydrate Physiology.................. 8
Stress Physiology................................. ..... 13
Root Regeneration in Southern Pines..................... 18

CHAPTER I. CHANGES IN MORPHOLOGY AND CARBOHYDRATE
RESERVES DURING THE NURSERY SEASON

Introduction........................... ......... ............ 20
Materials and Methods ................................. 21
Field Procedure. .......... .............. ........ 21
Laboratory Procedure............................ 22
Statistical Analysis............................. 24
Results....... ........................... ........... 25
Morphological.................. .................... 25
Carbohydrates ................................. 28
Proportionality of Dry Weight Increases............ 37
Discussion............................ ................ 39

CHAPTER II. CHANGES IN SEEDLING MORPHOLOGY AND
CARBOHYDRATE RESERVES AFTER OUTPLANTING

Introduction...................................... 44
Materials and Methods .. ........ ........................ 45
Nursery.................................................. 45
Field.............................................. 48
Laboratory....................... ...... ......... 49
Statistical Analysis............................... 49
Results................................................ 51
Morphological Characteristics at Lifting........... 51
Morphological Characteristics after Outplanting.... 53
Changes in Carbohydrate Reserves after
planting ........................................ 62
Survival and Height at 1 Year..................... 71

Discussion. ............................................ 78












CHAPTER III. THE EFFECT OF WATER STRESS ON SEEDLING
MORPHOLOGY AND CARBOHYDRATE PHYSIOLOGY

Introduction.................................... ...... 85
Materials and Methods ........................... ........ 86
Nursery .... ......... ................ .......... 86
Field............. ... ................... ..... 92
Laboratory............ ........................ 92
Statistical Analysis................. ........... 93
Results....... ........ ............................... 94
Soil Moisture.......... ...... ................... 94
Seedling Water Potentials ......................... 97
Seedling Morphological Characteristics............. 101
Carbohydrate Concentrations....................... 104
Amount of Carbohydrate per Seedling................ 112
Field Performance after Outplanting................ 114
Discussion... .... .................. ........... ....... 120
Seedling Growth................................... 120
Seedling Carbohydrate............................. 122
Outplanting ............................. ......... 126

SUMMARY AND IMPLICATIONS
Summary of Findings........................... .......... 130
Practical Implications.................................. 133

REFERENCES ............................ ..................... 137

BIOGRAPHICAL SKETCH ................. ................... ...... 146












LIST OF TABLES

1-1 Seasonal changes in dry weight and total non-structural
carbohydrate per seedling.............................. 38

2-1 Analysis of variance design............................ 50

2-2 Treatment means for morphological variables measured at the
time of lifting......................................... 52

2-3 Seedling morphological measurements after outplanting
averaged across treatments by sampling date............ 54

2-4 Treatment means for morphological variables averaged across
replications and sampling times 0 through 12 weeks..... 60

2-5 Carbohydrate values by treatment averaged across sampling
times after outplanting ............................... 63

2-6 Average values for carbohydrate variables for each sampling
date averaged across treatments........................ 64

2-7 Linear regression models for which the test B A 0 proved
significant at alpha = .05.... ........................ 75

3-1 Treatment averages for morphological variables measured at
the time of lifting................. ................... 102

3-2 Replication averages for morphological variables measured
at the time of lifting............................... 103

3-3 Average carbohydrate concentration at time of lifting.. 105

3-4 Best fitting models for carbohydrate concentrations at time
of lifting.. ........................................ 108

3-5 Average mass of sugar, starch, and total carbohydrate per
seedling at lifting.............................. ...... 113

3-6 Treatment averages for survival, height, and height
increment 1 year after planting........................ 115

3-7 Independent variables tested in linear regression against
the dependent variable of first year height increment.. 118











LIST OF FIGURES

1-1 Section 14 of the Archer nursery indicating experiment
locations.......... ....... ........... ..... ......... 23

1-2 Average shoot height and primary root depth............ 26

1-3 Average root collar diameter......................... .... 27

1-4 Average root, shoot, and total dry weight per
seedling................. ....... ... ....... ......... 29

1-5 Average root/shoot ratio per seedling (dry weight
basis) ............. ................... .............. 30

1-6 The relationship of log shoot dry weight to log root dry
weight for biweekly samples ........................... 31

1-7 Average seedling sugar concentrations................. 33

1-8 Average seedling starch concentrations................ 34

1-9 Average sugar and starch concentrations of entire
seedlings........ .............. ..................... 35

1-10 Average amount of carbohydrate per seedling............ 36

2-1 Archer nursery root pruning study indicating replication,
treatment, and sample plot location.................... 47

2-2 Average root, shoot, and total dry weight per seedling
after outplanting compared to seedlings remaining
in the nursery................ ........................ 57

2-3 Root and shoot moisture content of outplanted seedlings as
compared to those remaining in the nursery............. 59

2-4 Average dry weight of new roots per seedling after
outplanting............................................ 61

2-5 Average sugar and starch concentration per seedling after
outplanting, compared to those of seedlings remaining in
the nursery............................................ 67

2-6 Average root starch concentration after outplanting com-
pared to that of seedlings remaining in the
nursery................................................ 68











2-7 Average carbohydrate concentrations and absolute amounts
for whole seedlings after outplanting compared to those of
seedlings remaining in the nursery..................... 70

2-8 The regression of survival at 1 year over the number of
undercutting prior to outplanting .................... 72

2-9 The regression of first year height growth over the number
of undercutting prior to outplanting.................. 73

2-10 Relationship of survival at 1 year after outplanting to
average mass of new roots per seedling during the first 12
weeks after outplanting ............................ 76

2-11 Relationship of average new root mass per seedling during
the 2-12 week period after outplanting to total non-
structural carbohydrate concentration during the same
period................................................. 77

3-1 Structure covering the water stress experiment at Archer,
Florida, nursery....................................... 88

3-2 Layout of Archer water stress experiment.............. 89

3-3 Cross section of Bed 6, Section 14, Archer nursery showing
average seedling height and tap root depth relative to the
three tensiometer depths........................ ....... 91

3-4a Tensiometer readings at three depths for Treatment 1 on
successive dates during the study...................... 96

3-4b Tensiometer readings at three depths, January 31....... 96

3-5 The relationship of soil moisture content and soil tension
at three depths....................................... 98

3-6 Relationship of pre-dawn total stem water potential and the
number of days seedlings had not received water........ 99

3-7 The relationship of pre-dawn total stem waer potential and
soil moisture content at time of lifting............... 100

3-8 The regression of average shoot starch concentration at the
time of lifting and the total amount of water received
during the 8 week experiment.......................... 107











3-9 The regression of seedling root and sugar concentrations
and the number of days without receiving water prior to
sampling.................................. ............. 109

3-10 The regression of root and shoot starch concentrations and
the number of days without receiving water prior to
sampling............................................... 110

3-11 The regression of whole seedling sugar and starch
concentrations and the number of days without receiving
water before lifting...................... ............ 111

3-12 The regression of first year height increment and the
number of days seedlings had not received water prior to
lifting.. .......................................... 117

3-13 The regression of first year height increment and the
absolute amount of starch per seedling at the time of
lifting................................. ... ... .... 119


viii











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


THE RELATIONSHIP OF CARBOHYDRATE RESERVES TO
THE QUALITY OF BARE-ROOT Pinus elliottii
var. elliottii (Engelm.) SEEDLINGS PRODUCED
IN A NORTHERN FLORIDA NURSERY


By

Kenneth Lee McNabb

May 1985

Chairman: Earl L. Stone
Major Department: Forest Resources and Conservation



Three experiments explored relationships between carbohy-

drate reserves and quality of slash pine (Pinus elliottii var.

elliottii) seedlings in a northern Florida nursery.

In the first experiment, growth of roots and shoots varied

independently by season. Shoot dry weight increased most rapidly

in summer, root weight in winter. Total weight increased from

2.95 g on November 2 to 3.95 g on February 24. Carbohydrate con-

centration averaged 6% from May to October, then increased stead-

ily to 13.8% in late February. Root starch concentration

increased from 1.1% in October to 11.8% in February. Physiolog-

ical activity during the winter lifting season was therefore

substantial.











In a second experiment, seedlings were undercut at four fre-

quencies during November and December, then outplanted and per-

iodically sampled. Outplanted seedlings suffered no loss in dry

weight and reinitiated normal carbohydrate accumulation within 2

to 4 weeks. A temporary increase in sugar concentration occurred

simultaneously with decreased starch. Root growth after out-

planting was not dependent upon carbohydrate levels. Compared to

the controls, undercutting resulted in 115% more new root mass,

68% more root starch during the first 12 weeks after outplanting,

24% higher survival, and 6% more first year height increment.

Survival was linearly related to new root mass after outplanting

(R2 = 0.48).

In the third experiment, seedlings were subjected to five

levels of water stress during December and January and were then

outplanted. Pre-dawn xylem water potential of severely stressed

seedlings averaged -1.3 MPa at lifting. Seedling morphology was

not measurably affected. Sugar concentrations increased curvi-

linearly (R2 = 0.92) with stress intensity, whereas starch de-

creased linearly (R2 = 0.83) suggesting osmotic adjustment. The

starch-sugar conversion was rapidly reversed by watering.

Nursery water stressing did not affect field survival but

decreased first year height increment from 5.4 to 0.9 cm, proba-

bly related to impaired bud development. Height increment,
2
however, was also correlated with total seedling starch (R =

0.67).











GENERAL INTRODUCTION


Pine plantations provide an increasing portion of the supply

base for the numerous wood products industries in the southeast-

ern United States. Plantations are especially important to the

pulp and paper industry, which is committed to short-rotation

silviculture for fiber production. An essential part of the

plantation system is the regeneration of harvested stands, in-

cluding site preparation, planting, and cultural practices, such

as fertilization and weed control. The success of this expensive

enterprise depends upon the survival and early growth of planted

seedlings.

The vast majority of seedlings planted in the Southeast are

pines, which are usually produced in large-scale nursery opera-

tions. They are lifted bare-root and outplanted during the

months of November to March. Slash pine (Pinus elliottii

Engelm.) and loblolly pine (Pinus taeda L.) are by far the most

common species. In fact, the combined nursery production of

slash and loblolly pine exceeds the total production of all other

species in the entire U.S. (Abbott and Fitch 1977).

Slash and loblolly pines are native.to the Southeast and

have a long history of successful plantation establishment.











Recently, however, there has been a growing concern about sur-

vival after field planting. Weaver et al. (1980) reported that

overall average survival in pine plantations decreased from 82%

in the early 1960s to 73% in the 1970s. Even though some land-

owners have not experienced this drop, prudence dictates concern

for adequate survival.

Field survival depends upon many factors, notably weather,

site conditions, and handling. Another crucial factor is use of

a quality seedling, one which can cope with weather, site, and

handling conditions so that initial survival and growth are maxi-

mized. If a nurseryman is to produce a quality seedling, how-

ever, he must be able to define what "quality" is. There must be

specific seedling parameters which indicate whether a seedling is

optimally plantable.

The definition of a quality seedling has proven elusive.

For the past 50 years, nurserymen have used morphological char-

acteristics as a measure of seedling quality. Such character-

istics are height, root collar diameter, presence of secondary

needles, presence of a dormant (winter) bud, woodiness of the

stem, and the root/shoot ratio. Specific combinations of these

characteristics have been used to define morphological grades

(Wakeley 1954). Many studies indicate a strong relationship be-

tween morphological grades and seedling quality (Wakeley 1954,

Switzer and Nelson 1967, Burns and Brendemuehl 1971, Blair and

Cech 1974).











Yet, morphological grades are not always accurate indicators

of seedling performance. At an early date Wakeley (1949) noted

important exceptions to the morphological grade-quality relation-

ship. He proposed that there were "physiological grades" of

southern pine nursery stock which were the true determinants of

quality and that these might or might not coincide with morph-

ological grades. The inadequacy of morphological grades in de-

termining seedling quality has been further indicated by Stone

(1955), Shoulders (1960), Stone et al. (1962), Stone and Jenkin-

son (1971), Blair and Cech (1974), and Chavasse (1977). It is

now generally accepted that seedling quality is influenced by

physiological condition (Sutton 1979, Brissette et al. 1981).

Research on the numerous physiological factors that may be

related to seedling quality has concentrated on osmotic poten-

tial, nutritional balance, endogenous plant growth regulators,

and carbohydrate reserves. A hypothesized relationship between

carbohydrate reserves and slash pine seedling quality holds that

root regeneration potential of outplanted seedlings is dependent

upon carbohydrate reserves (Wakeley 1949). Recent research has

lent some support to this hypothesis (Barnard et al. 1981). The

lack of a strong photoperiod influence or chilling requirement

for the spring shoot flush of slash pine seedlings (Wakeley 1954,

Kaufmann 1977, Fisher 1981), as opposed to many other pine spec-

ies, strengthens this hypothesis inasmuch as it suggests reduced

hormonal involvement. Even so, there has been little work on the











contribution of carbohydrate reserves to seedling quality and

several basic issues have yet to be resolved.

The overall objective of the present study was to investi-

gate the relationship between carbohydrate reserves and seedling

quality for nursery-grown slash pine (Pinus elliottii var. el-

liottii Engelm.) in northern Florida. The study concentrated on

four issues.

A. To determine the "normal carbohydrate levels in nursery-

grown slash pine seedlings

B. To determine the influence of some cultural treatments on
"normal" carbohydrate levels

C. To determine what happens to these reserves after

outplanting

D. To relate carbohydrate reserves to survival in the field

During the course of this study, a freezer malfunction re-

sulted in the thawing of certain samples and their subsequent

loss for the purpose of carbohydrate analysis. While this re-

duced the total amount of information available, enough sound

data were produced to address the specified objectives.













LITERATURE REVIEW


Introduction



The ability of nursery-grown seedlings to survive outplant-

ing depends on the rate of new root production. As Switzer and

Nelson (1967, p.5) pointed out, "the typical planter reasons that

trees must survive to grow, while in reality they must grow to

survive."

Several studies relate seedling field performance to Root

Growth Capacity (RGC). Both survival and height increment of

boreal conifer seedlings have been related to RGC (Sutton 1980).

Burdett (1979) showed that in certain circumstances, survival of

lodgepole pine (Pinus contorta Dougl.) is largely dependent on

its RGC. Rapid root growth immediately after outplanting is also

critical to the survival of southern pines (Woods 1959, Switzer

and Nelson 1967, Kozlowski 1979). Wakeley (1954, p.123) states

that "high initial survival seems to depend, perhaps even more

than that of pines planted in other regions, upon formation of

considerable new root tissue after planting."











Lateral Root Formation



The physiology of lateral root formation is complex and in-

completely understood. Wightman and Thimann (1980) showed that

for Phaseolus the increase in lateral root growth after the root

tips are removed is a result of a specific promoter (an auxin),

which is produced in the leaves cotyledonss) and moves

acropetally to concentrate in the most apical segment of the de-

capitated roots. Growth of lateral root primordia is inhibited

by substances produced in the root tip. Removal of this inhib-

itory effect by root decapitation did not increase the number of

root primordia but allowed growth of existing primordia. Lateral

root extension, therefore, resulted from a synergistic effect of

a promoter produced in the cotyledons and an inhibitor produced

in the root tips.

The effect of IAA on lateral root emergence is directly con-

nected to carbohydrate physiology. Altman and Wareing (1975,

p.37), also working with Phaseolus, found that IAA affected car-

bohydrate transport directly, as well as inducing the formation

of a stronger "sink" in the root primordia. These two effects

were independent. One of the proposed effects of leaf-produced

IAA was a stimulation of the conversion of starch to sugar. The

authors stated that "where root initiation is easy, carbohydrates

transported from the leaves are the main limiting factor, rather

than unknown 'co-factors' of hormonal nature."











One of the possible mechanisms whereby carbohydrates

activate primordia is through an effect on the mitotic cycle.

With Vicia faba, Van't Hof et al. (1973) showed that carbohydrate

starvation caused cell arrest in the G1 and G2 phases of mitosis.

The longer the starvation, the more difficult to re-initiate mi-

tosis. Also, the first metabolic activity after sucrose feeding

of starved cells was protein synthesis.

One of the first events during root primordia development is

increased activity of enzymes directly involved in carbohydrate

physiology (Haissig 1982b). Application of napthaleneacetic acid

(NAA) to Pinus banksiana (Lamb) seedling cuttings caused an in-

crease in the activity of glyceraldehyde 3-phosphate dehydro-

genase (Haissig 1982a). This increase was temporally associated

with callus formation. Moreover, development of the callus pri-

mordium was positively correlated with total carbohydrate.

Sugars produced in the needles were translocated and concentrated

in the stem, with the exogenously applied auxin directing the

basipetal transport of these sugars.

Stoltz (1968) found that rooting ability of two difficult-

to-root cultivars of Chrysanthemum was correlated with total car-

bohydrate concentrations. The cultivar with the highest carbohy-

drate reserves produced more roots regardless of preparation

date.











Root cuttings of apple cultivars also showed maximum rooting

ability when carbohydrate contents were greatest (Robinson and

Schwabe 1977). Carbohydrate contents of apple tree roots vary

according to season. Levels of both carbohydrate and rooting

ability were highest in autumn and winter. During the growth of

these roots, polysaccharide reserve concentrations declined by 30

and 40%, presumably being used for root and shoot formation. The

authors held that a "minimum polysaccharide threshold level" of

approximately 30% of root dry weight was necessary for survival.

The same study found evidence of carbohydrate X IAA interactions,

however, and the authors felt that a suitable balance of each was

necessary for optimum rooting potential.

Similarly, a hormone/carbohydrate balance was also necessary

for root formation on etiolated stem segments of Populus (Nanda

et al. 1971, p.391). The effectiveness of an auxin in promoting

stem segment rooting depended upon the carbohydrate produced in

leaves. The authors concluded that the "rooting of 2.5 cm long

segments of Populus nigra is limited primarily by nutritional

factors."



Seasonality of Carbohydrate Physiology



Woody species, as a rule, have seasonal fluctuations in car-

bohydrate concentrations, especially starch. The amount of

photoassimilates going into storage increases in the fall











(Kruegar and Trappe 1967, Ursino et al. 1968, Schier 1970, Nelson

and Dickson 1981). Krueger and Trappe (1967) found that both

sugar and starch in Douglas-fir (Pseudotsuga menziesii Franco)

seedlings fluctuate considerably by season, with winter concen-

trations of sugars three times those in summer. The same was

true for starch except that concentrations peaked in early

spring.

Winter carbohydrate reserves may serve to fuel the flush of

growth in the spring, especially in deciduous species (Kramer and

Kozlowski 1979). Utilization of carbohydrate reserves for the

sprouting of aspen sucker shoots has been documented (Schier

1981, Fitzgerald and Hoddinott 1983). Decreases in total car-

bohydrate concentrations of seedling shoots during the spring

shoot flush have been demonstrated in Douglas-fir (Kruegar and

STrappe 1967). On the other hand, a study with 5 year old balsam

fir (Abies balsamea L.) seedlings transplanted into a nursery

soil found the spring flush independent of reserves (Little

1974). In this case, current photoassimilates supported the

spring top growth.

Apparently, environmental conditions influence the balance

between use of current photosynthate and reserves during the

spring flush. In the case of young red pines (Pinus resinosa

Ait.), unfavorable temperatures and moisture may delay the re-

establishment of the photosynthetic capacity of old needles and











thus increase the importance of reserves for the spring flush

(Gordon and Larson 1970).

Although Kruegar and Trappe (1967) found shoot reserves used

for new shoot expansion, there was no parallel decrease in root

starch concentrations during spring root growth. The authors

concluded that this indicated a "steady-state" situation, with

sugars from the top supporting root extension. Schier (1970),

however, showed that red pine root expansion from January to July

must have been at the expense of root reserves. Ursino et al.

(1968) also discovered little evidence for spring translocation

of photoassimilates to the roots in white pine (Pinus strobus

L.).

A number of studies have demonstrated seasonality of RGC in

forest nursery seedlings. Stone et al. (1962, p.296) found that

Douglas-fir nursery seedlings "displayed marked periodicity in

their root--regenerating potential," with winter-lifted seedlings

having much higher RGC than summer-lifted seedlings. Lavender

(1964) felt that early lifting decreased RGC due to disruption of

the physiological process of dormancy. Stone and Jenkinson

(1971) reported that the ability of ponderosa pine (Pinus ponder-

osa Laws) seedlings to regenerate roots is dependent upon the

number of hours that seedlings are exposed to low air tempera-

tures in the nursery. A minimum of 1500 hours below 10C is

needed to insure a high root growth capacity. Spring RGC for

northern hardwoods is also controlled by a chilling factor











(Farmer 1978). Exogenous applications of IAA or gibberellins did

not improve the RGC of either Douglas-fir (Lavender and Hermann

1970) or ponderosa pine (Zaerr 1967).

The relationship between dormancy controls, such as chil-

ling, and carbohydrate levels is quite complex. How they inter-

act to influence RGC of seedlings has not been completely

elucidated. Most of the studies on this topic have concentrated

on species with strong chilling requirements.

Ronco (1973, p.213), for example, stored 3-0 Engelmann

spruce (Picea engelmannii Parry) seedlings 4 months at 1-20C.

Total carbohydrates decreased from 265 to 120 mg/g dry weight,

and upon outplanting after storage survival was very poor. A

second group of seedlings that were lifted from holding beds and

planted immediately (no storage) had total carbohydrate reserves

of over 300 mg/g dry weight and survived better. When comparing

the performance of both groups, Ronco concluded that "survival

may be adversely affected by carbohydrate concentrations below

certain threshold levels when trees are planted."

McCracken (1979) stored Pinus mugo Turra and Pinus radiata

D. Don for up to 18 weeks at 10C. At 6, 12, and 18 weeks, seed-

lings were sampled for laboratory analysis as well as potted for

growth analysis. Total carbohydrates decreased "markedly" for

both species, especially Pinus radiata during cold storage. Fur-

thermore, in both species, decreases in carbohydrate levels were











associated with increases of new tissue after potting, indicating

that reserves were used during the growth process. This evi-

dence, along with a previous finding that cold storage reduces

photosynthetic efficiency (McCracken 1973), caused the author to

conclude that stored carbohydrates form a vital resource for

growth after cold storage and transplanting.

Ritchie (1982) obtained somewhat different results with

Douglas-fir. Seedlings were lifted periodically from November to

March, stored at 20C for different lengths of time, and their RGC

and carbohydrate content were determined. Total nonstructural

carbohydrate (TNC) decreased during storage, but RGC did not al-

ways do so. For example, with January-lifted seedlings, TNC de-

creased during the first 6 months of storage, whereas RGC (of

planted seedlings) actually increased. The author concluded that

changes in RGP are not driven by changes in carbohydrate

concentrations.

Van den Driessche (1979) presented similar results for red

pine and white spruce (Picea glauca Voss) seedlings. Root growth

capacity was low in fall and spring but high in midwinter. These

differences in RGC did not appear to be related to carbohydrate

reserves. The results also indicated that current photosynthate

was essential for new growth in Pinus resinosa.











Recent work with hardwoods has indicated a seasonal fluc-

tuation of starch reserves (Rose and McGregor 1982, Rietveld et

al. 1982). With four hardwood species in southern Illinois,

Rietveld et al. (1982) found a weak, but consistent, association

between later lifting dates, which had higher root starch con-

centrations and greater survival and/or growth in the field.

Unfortunately, TNC was confounded with lifting date, which did

not allow for the separation of their individual effects upon

field performance.



Stress Physiology



The process of lifting, storage, transport, and planting

imposes various types of stress on nursery-produced seedlings.

The chief stress factor is desiccation, which can occur at any

time before or soon after planting. It is evident that the

physiology of outplanting is to a great extent the physiology of

water-stressed seedlings.

Increasing levels of water stress usually result in decreas-

ing photosynthetic capacity. A water potential of -0.4 MPa has

been shown to reduce the photosynthetic capacity of loblolly

pine, with assimilation becoming negligible at -1.1MPa (Brix

1962). This effect was caused by closure of the stomates, which

reduces transpiration as well as photosynthesis. Upon rewater-

ing, tree seedlings re-initiate photosynthesis. The period of











time required by the plant to return to 100% efficiency, however,

depends upon the length and intensity of the previous water def-

icit, species, and humidity (Brix 1962, Zavitkovski and Ferrell

1970, Kramer and Kozlowski 1979). Plants rewatered after water

stress may begin transpiring before photosynthesizing, indicating

a period when the photosynthetic mechanism is being repaired

(Kramer and Kozlowski 1979).

The ability to tolerate water stress is an important com-

ponent of the adaptability of a species. A species may be lim-

ited to moist sites by sensitivity of stomata to water stress and

poor recovery of leaf gas exchange following rewatering (Davies

and Kozlowski 1977). While slash pine is a wet-site species

(Fisher 1981), it possesses the xerophytic adaptations of a heavy

cuticle and sunken stomates characteristic of the genus. More-

over, it can be found growing naturally alongside longleaf pine

(Pinus palustris L.) on drier sites.

Water stress has been shown to cause specific changes in the

carbohydrate physiology of tree seedlings. Decreases in starch

simultaneous to increases in sugars during drought stress have

been reported for sugar maple (Acer saccharum Marsh.) (Parker

1970) and black oak (Quercus velutina Lam.)(Parker and Patton

1975). In the case of sugar maple, the total reserves were not

affected, indicating a transfer of carbohydrate from the storage

form (starch) to sugars. Similar changes occur in the inner bark

of loblolly pine (Hodges and Lorio 1969) and the tops of cotton

plants (Eaton and Ergle 1948).











The increase in sugar concentrations resulting from water

stress may serve a number of purposes, of which two are of prime

consideration. First, an increase in free sugars increases the

levels of readily available substrate for respiration. Brix

(1962) found that when loblolly pine seedlings were subjected to

increasing levels of water stress, respiration first decreased,

then dramatically increased. At one point (-2.8MPa plant water

potential), respiration was 140% of the rate at soil field

capacity.

A second effect of increased free sugars during water stress

is to lower the osmotic potential of cell solution and therefore

maintain cell turgidity as soil water becomes unavailable. Car-

bohydrates, organic acids, and inorganic ions can be used for

osmotic regulation (Kramer 1983). That solute accumulation oc-

curs in woody plants subjected to moisture stress was demonstra-

ted by Osonubi and Davies (1978). The authors stressed young

seedlings of English oak (Quercus robur L.) and silver birch

(Betula verrucosa Ehrl.) and observed a more negative leaf solute

potential as soil water content decreased. The efficiency of

osmotic regulation varied between species. The oak was able to

lower solute potentials and thus maintain turgor pressure and

keep stomates open at lower soil water potentials. Unfortun-

ately, the constituents of the solutes were not assessed in this

study, and it is not known whether organic or inorganic ions were

the principal influences upon changes in osmotic potential.











Root pruning (undercutting) is a part of the standard

lifting process. This results in substantial loss of the root

system, especially non-suberized root tips as well as damage to

the entire root system when the soil is lifted as the blade

passes underneath. Undercutting would therefore induce seedling

moisture stress.

By successively root pruning over a period of time well

before lifting, seedlings are rendered more drought tolerant.

Root "wrenching" is a standard cultural technique in the pro-

duction of radiata pine in New Zealand (Van Dorsser and Rook

1972), and Caribbean pine (Pinus caribaea Mor.) in Australia

(Bacon and Hawkins 1979). These studies and others (Shoulders

1963, Tanaka et al. 1976) have found that the systematic stres-

sing of seedlings by root pruning and/or wrenching in the nursery

improves field performance.

The reason for the improved performance is not clear but may

be related to morphological changes resulting from undercutting.

Root pruning results in a more fibrous root system, smaller

foliage volume, and a reduced height, which makes for a more

favorable root/shoot ratio (Rook 1971). Physiological changes in

radiata pine include increasing amounts of photosynthate sent to

the pruned roots, which causes higher concentrations of reducing

sugars and starch (Rook 1971).











Bacon and Bachelard (1978) found that root wrenching

Caribbean pine changed not only seedling morphology but also the

physiological response of seedlings to water stress after

outplanting. Seedlings that had been intensively wrenched kept

stomates open upon potting, whereas control seedlings did not.

Control seedlings stopped transpiring within two days after

potting even though soil moisture was adequate. On the other

hand, while wrenched seedlings declined in transpiration rate

after potting, they did not completely stop. Furthermore,

wrenched seedlings showed the highest net photosynthetic rate

during the 15-day measurement period after potting. The results

indicate that the seedling's ability to photosynthesize after

transplanting is related to the level of conditioning in the

nursery.

Because seedlings are normally lifted in the winter, they

may be exposed to low soil temperatures after outplanting.

Nambiar et al. (1979, p.1119) found that low soil temperature

slowed the initiation and elongation of new roots of radiata

pine. The study indicated that the water stress which occurred

during the first several weeks after planting was "due primarily

to the suppressive effect of low soil temperature on root

regeneration." The authors maintained the soil temperature of

potted seedlings at 5, 10, or 200C. The seedling stress levels

varied among these soil temperatures even though the pots were

watered twice weekly.











Undoubtedly, some water was absorbed through suberized roots

(Kramer 1946, Chung and Kramer 1975), but the growth of new roots

was necessary to establish an adequate internal water balance.



Root Regeneration in Southern Pines



Wakeley (1949) was one of the earliest investigators to re-

late carbohydrate levels to planting stock quality of southern

pines. He found that either shading or defoliating longleaf

(Pinus palustris Mill.) and slash pine seedlings in the nursery

over a period of 3 to 12 weeks affected the survival after

outplanting. These experiments were repeated over four succes-

sive years. The more extreme treatments reduced survival by 45

to 97% in each of the 4 years. Staining with potassium iodide

indicated that both severely shaded and defoliated seedlings con-

tained little or no starch.

Gilmore (1961) was able to demonstrate that shading reduced

carbohydrate reserves of loblolly pine, and correlated survival

of the shaded seedlings with their root carbohydrate reserves.

In later publications (1964, 1965), he reported no correlation

between carbohydrate reserves and survival or root growth after

outplanting normal unshadedd) nursery-run seedlings. He con-

cluded that a "growth promoting substance" produced in the shoots

provide roots with a necessary stimulus for root growth.











Hay and Woods (1968, 1975, 1978) examined the interaction

between root deformation and root regeneration in loblolly pine.

When seedlings were planted in "deformed" positions, such as "J

rooting," free sugars tended to concentrate at the point of

curvature. Translocation was therefore impeded, with the root

deformation acting as a phloem girdle. Lateral root development

was most visible at the bottom of the "J" and in association with

the increased carbohydrate concentrations.

The only recent work concerned with carbohydrate reserves in

slash pine seedlings is that of Barnard et al. (1981). The au-

thors compared root starch concentration with survival after out-

planting for nursery-run seedlings from each of four nurseries in

northern Florida. Low survival appeared to be associated with

low root starch concentrations. The authors concluded that the

relationship of low starch with low survival was consistent and

warranted further investigation.















CHAPTER I
CHANGES IN MORPHOLOGY AND CARBOHYDRATE RESERVES
DURING THE NURSERY SEASON






Introduction



In order to relate the field performance of slash pine

seedlings to their carbohydrate reserves, some basic information

on seedling carbohydrate physiology is required. For example,

what are "normal" levels of carbohydrate reserves for slash pine

seedlings? Are these reserves found in amounts sufficient to

contribute to survival after outplanting? Also, is there an ap-

preciable seasonal variation in carbohydrate reserves? If so,

this would suggest variability in the potential contribution of

such reserves to field performance.

A study to follow morphological and total non-structural

carbohydrate (TNC) development over time was established in an

industrial forest nursery near Archer, Florida. The quantity of

reserves, their changes over time, and their general relation to

the morphological characteristics of the plant were followed.













Materials and Methods

Field Procedure

The nursery selected for this study has produced around 20

million bare-root loblolly and slash pine seedlings per year

since beginning production in 1971. The area receives an annual

average of 1370 mm rainfall, evenly distributed except for a late

fall- early winter dry period, with November averaging less than

50 mm (Dohrenwend 1978). Mean January and July temperatures are

14.4 and 27.3C, respectively. The soil is a moderately well

drained Grossarenic Paleudult, which has been graded to improve

drainage. A clay layer is found from 100 to 120 cm depth in the

area of the study.

The sampling area was managed according to standard pro-

cedures of this nursery. The seed was from a single open-

pollinated seed orchard of improved slash pine. After soil fumi-

gation, seed were sown mechanically at a rate that would produce

about 30 seedlings per square foot of bed space. Fertilization

consisted of a plowdown application of 0-20-10 at 672 kg/ha, and

subsequent applications of 10-5-5 liquid fertilizer three times

during the summer. The liquid applications were made through the

sprinkler system with an objective of 16.8 kg/ha of elemental

nitrogen at each application. Bayleton (triademiton) systemic

fungicide was sprayed on May 5 and 20 and on June 10.











Seedlings were sampled throughout the entire growing season,

beginning 4 weeks after sowing on April 28, 1982, and terminating

on March 8, 1983. The experimental area consisted of two nursery

beds 173.7 m (570') long by 1.2 m (4') wide, with eight seedling

rows per bed. Eight 9.1 m (30') long replicate plots were evenly

spaced along these two beds (Figure 1-1). Every 2 weeks, at

least 20 seedlings were lifted from each replication. As there

were eight replications and eight rows per bed, a different row

was sampled from each replication at each time; thus the entire

bed width was represented. Lifting was done with a shovel, at-

tempting to get the entire root system.



Laboratory Procedure

After lifting, the seedlings were placed in plastic bags and

covered with ice in a cooler. They were then brought to the lab-

oratory and stored frozen (-20 to -100C) until measured.

Upon removal from the freezer, the seedlings were immersed

in water for thawing and washing (3-5 minutes). Several morpho-

logical characteristics were then measured on 20 seedlings from

each replication. These characteristics included shoot length

from root collar (height), diameter at root collar (DRC), length

of the primary root from the root collar (measured only when the

primary root was intact), and presence or absence of a dormant

(non-active) bud in seedlings sampled after October. Ten of























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these seedlings were blotted dry and both green and oven dry

(1050C) weight determined. The remaining ten were used for

carbohydrate determination. Root and shoot were separated and

dried at 1050C for 1 hour followed by 70C for 23 hours.

Thereafter, the tissue was milled to pass 40 mesh and stored in

air-tight vials. A total of 336 carbohydrate analyses was

required for analyses of roots and shoots of eight replications

and 21 sample dates.

The analytical procedure followed that of Rowe (1981). Free

sugars were extracted from a 100 mg sample with 80% ethanol. The

sample was then subjected to enzymatic hydrolysis of the starch

for 2 hours using amyloglucosidase (Sigma, from Rhizopus). Both

the ethanol solution of free sugars and the enzyme-produced glu-

cose solution were analyzed colorimetr.ically using anthrone re-

agent dissolved in 80% concentrated sulfuric acid. Their sum is

the total non-structural carbohydrate (TNC) of the sample).


Statistical Analysis

Values of the eight replications on each sampling date were

averaged and expressed on a per seedling basis. The averaged

values were graphed to follow parameter changes over time. Also,

average seedling values of root and shoot dry weight were used

for allometric analysis, which plots the logarithm of shoot dry

weight against the logarithm of root dry weight. The MEANS and











General Linear Model procedures of the Statistical Analysis

System (SAS) were used for data analysis.



Because of a freezer malfunction, stored samples taken on

November 2, December 1, and January 10 did not produce acceptable

carbohydrate data. Morphological data from these samples were

utilized, however.



Results



Morphological Characteristics

There were three distinct phases of height growth: an init-

ial period of slow growth, rapid summer growth, then a decreasing

growth rate in fall and winter (Figure 1-2). In the 6 weeks from

July 8 to August 19, the seedlings grew 13.2 cm, or 44% of the

total height accrued during the entire study. A second period of

rapid height increase began in late February.

From first sampling on May 27 to July 8, a period when shoot

growth was relatively slow, the primary root was rapidly elonga-

ting. By July 8, primary root length was 30.5 cm, or 86% of the

total length at the termination of the study. Primary root elon-

gation slowed during July and was generally associated with a

decreased length in the zone of elongation and increased degree

of suberization. This did not appear to be related to any speci-

fic soil changes.










40


30


20


PRIMARY ROOT LENGTH


-"... -*.------- ._._ .


JULY 1


SEPT 1


NOV 1


JAN 1


MAR 1


Figure 1-2. Average shoot
represents 160 seedlings.
for height and 2.2 cm for


height and primary root length. Each point
Maximum S- for any single point is .54 cm
primary root depth.


SHOOT HEIGHT




/
0*0"j


(Il


101


30 L


I I I

















5



4 ,*

/* \0/

,, 3

-- /0


/
2/









JULY 1 SEPT 1 NOV 1 JAN 1 MAR 1






Figure 1-3. Average root collar diameter. Each point represents 160
seedlings. Bar represents the maximum S- for any single point.
Jx











While top height and primary root length remained fairly

stable after a certain period, seedling root collar diameter

(DRC) continued to increase throughout the study period (Figure

1-3). Although the increase is approximately linear indicating

that diameter growth occurred at a steady pace, there was a

slight decrease during the winter.

Like DRC, seedling dry weight increased throughout the en-

tire study period (Figure 1-4). Increases of root and shoot var-

ied by season. There was an early development of root mass

followed by the summer and fall shoot increase, which overlapped

the subsequent period of fall and winter root growth. Mean shoot

dry weight increased from 2.3 g on November 16 to 2.7 g on Feb-

ruary 24, an increase of 18%, whereas root weight went from .62 g

to 1.22 g, an increase of 97%. Changes in relative growth rate

of the two organs caused a strong U-shape in the root/shoot ratio

over time (Figure 1-5).

Allometric analysis is often used to analyze the relative

growth rates of plant organs (Leopold and Kreidiman 1975). A

regression of log shoot dry weight with log root dry weight has

an R2 of .94 (Figure 1-6). Despite the high correlation, the

line is obviously not linear, especially at the uper end.



Carbohydrates

Free sugar concentrations of both roots and shoots were

higher during the winter than in the summer by about 20 mg/g (2%)


















TOTAL


SHOOT


JULY 1 SEPT 1 NOV 1 JAN 1 MAR 1


Figure 1-4. Average root, shoot, and
point represents 160 seedlings. Bars
point.


total dry weight per seedling. Each
are the maximum S- for any single
x












.5




.4




.3




.2




.1






JULY 1 SEPT 1 NOV 1 JAN 1 MAR 1



Figure 1-5. Average root/shoot ratio per seedling (dry weight basis). Each
point represents 160 seedlings. Bar represents the maximum S- for any single
point. x


















3.5-


3



I-
I-I
"'2.5


O-


C.
-r
1 2





1.5


0*/

0/
*6/


/0


/0


1.5


3.5


LOG (ROOT DRY WEIGHT)


Figure 1-6. The relationship of log shoot dry weight to log root dry weight
for biweekly samples. (The linear regression is y = 0.51 + 1.01x, R2 .94.)














on a dry weight basis (Figure 1-7), with concentrations beginning

to increase in October and peaking on January 24. In general,

free sugars in the shoots and roots remained around 5% and 3%,

respectively, during the summer and then increased to 7% and 5%

during the winter.

The considerable fluctuation of sugar concentrations between

sampling dates in early summer may be related to seedling size.

For example, on June 11 the seedlings were only 6.5 cm tall and

had a proportionately large photosynthetic area. The diurnal

variation of photosynthesis would result in higher sugar con-

centrations if sampled in midafternoon as opposed to early

morning (Kramer and Kozlowski 1979). As seedlings get larger,

these daily concentration changes are buffered by the propor-

tionally greater amount of woody tissue.

The late fall,and winter increases in starch concentration

were much larger (Figure 1-8). Root starch concentration in-

creased from an average of 1% in late October to 12.8% on Feb-

ruary 28, whereas shoot concentration rose, from around 1% to

4.3% in the same period.

The relative contribution of starch and sugar concentrations

to seedling TNC becomes clearer when placed in the perspective of

the whole plant (Figure 1-9). Sugar concentrations of the whole

plant are higher than starch concentrations until late winter

(February 10). Because sugars are concentrated in the shoot,

which is the larger fraction of total seedling weight, their

contribution to the total seedling is proportionately large.











100



80

S-


60




E 40

o SHOOTS


20 ROOTS



-I I I I
JULY 1 SEPT 1 NOV 1 JAN 1 MAR 1






Figure 1-7. Average seedling sugar concentrations. Each point is an average
of eight composite samples of 10 seedlings. Bars represent the maximum S- for
any single point.











140



120
ROOTS -o------o-

SHOOTS I
100


E
S80



60


40


I
20


1 1 I i I

JULY 1 SEPT 1 NOV 1 JAN 1 MAR 1





Figure 1-8. Root and shoot starch concentrations. Each point is an average
of eight composite samples of 10 seedlings. Bars represent the maximum S-
for any single point. x







































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00 ko 1-z CM.


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ia C-











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600



500


100



0






Figure 1-10.
Each point is
represent the


TOTAL
SHOOTS
ROOTS


JULY 1 SEPT 1 NOV 1 JAN 1 MAR 1


Absolute amount of non-structural carbohydrate per seedling.
an average of eight composite samples of 10 seedlings. Bars
maximum S- for any single point.
x\











The mass of total non-structural carbohydrate per seedling

(Figure 1-10) increased over the entire nursery season. This is

primarily a function of increasing seedling mass. Sugars compose

89% of the total amount of carbohydrate on October 19, and starch

only 11%. This difference gradually diminishes until on February

24 when 48% of the total reserves are sugars and 52% are

starches.



Proportionality of Dry Weight Increases

Seasonal variability in photosynthate allocation is indica-

ted in Table 1-1. The nursery season has been divided into four

equal periods of 5 weeks each. Dry weight increases of shoots

were largest in the late summer, while the roots added more dry

weight in the winter. The reduced rate of growth during the per-

iod of October 19 to December 15 is most likely related to the

low rainfall, normal for this time of year (Dohrenwend 1978),

coupled with the practice of withholding irrigation at this time.

Seasonal variability of TNC increase per seedling followed

that of dry weight. The greatest increase in weight and carbo-

hydrate reserves of the shoots took place between July 22 and

October 4 (Table 1-1), whereas the greatest increase in root

weight and carbohydrate content occurred between December 15 and

February 24.






















Table 1-1. Periodic changes in dry weight and total non-structural
carbohydrate (TNC) per seedling (R = roots, S = shoots, T = total).

Sample period Dry weight increase TNC increase ATNC/A dry weight

R S T R S T R S T


(mg) (mg) (mg)
May 27 July 22 129 497 626 5 27 32 4 5 5
July 22 Oct 4 329 1523 1852 27 165 192 8 11 10
Oct 4 Dec 15 198 530 728 45 114 159 23 22 22
Dec 15 Feb 24 562 181 743 135 66 201 24 36 27











The percentage of total dry weight increase attributable to

non-structural carbohydrates increased from 5% in the early sum-

mer to 27% during the winter. Thus, during the late fall and

throughout winter, the seedlings were allocating ever-increasing

amounts of photosynthate to storage instead of growth. Regard-

less of season, the proportion of mass increase attributable to

reserve carbohydrates is roughly similar for roots and shoots,

with a maximum difference in the final period (24% vs. 36%).

This is unexpected as the root is normally thought of as a stor-

age organ that would accumulate carbohydrates at a faster rate

than it would form new cell wall material.



Discussion



The results of morphological analysis demonstrate that the

relative growth rates of roots and shoots changed according to

season. Fertilization undoubtedly influenced the patterns of

growth by stimulating shoot development during the summer. The

rapid summer growth differred from Huberman's (1940) early find-

ing with slash pine in which seedlings increased in height at a

more or less constant rate through the summer and fall and then

leveled off at 25 to 30 cm in November and December. The present

study also found height to level off at 25 to 30 cm, but this oc-

curred in September. Presumably, these results are influenced by











the fertilization practices not employed when Huberman did his

work. Climate may have been a factor also, as the former study

was done in central Louisiana.

The summer applications of nitrogen increased top growth

more than root growth. The regression coefficient of 1.01 from

the allometric analysis, also known as the coefficient of allo-

metric growth (Leopold and Kreideman 1975), is high when compared

to a previously reported value of .75 for potted loblolly pine

(Ledig et al. 1970), indicating a greater shoot weight accumu-

lation relative to roots during the June through October period.

The present study, as well as others (Huberman 1940, Perry

1971), revealed substantial dry weight increases, presumably due

to photosynthesis, during the winter for southern pines. Furth-

ermore, 70 to 80% of the seedlings sampled for this study in De-

cember, January, and February had non-dormant terminal buds, with

a minimum of 57% on February 10. The lack of a resting terminal

bud, however, has been shown not to be detrimental to outplanting

survival (Wakeley 1954).

The higher sugar concentrations during fall and winter pre-

sumably relate to cold weather protection (Aronsson and Ingestad

1976, Kaurin et al. 1981). Increased sugar concentration, pri-

marily sucrose, in response to lower temperatures, is one of the

most important physiological changes occurring during the devel-

opment of cold hardiness (Kramer and Kozlowski 1979).











Root starch concentrations began to increase in November and

continued to increase throughout the winter, with no certain peak

(Figure 1-8). This is in contrast to the winter increases of

seedling root starch concentrations reported for Douglas-fir

(Krueger and Trappe 1967), which increased from 0.5% in late Feb-

ruary to 8% in middle March. Thereafter, root starch gradually

declined through the growing season. In the present study, slash

pine seedlings began starch accumulation earlier and attained

considerably higher concentrations. Without samples later in

March, it cannot be determined whether the small decline from

February 24 to March 8 was the beginning of a downward trend.

The winter starch concentrations reported here are similar

to those of Barnard et al. (1981). These authors had sampled the

same nursery 4 years earlier and found average root starch con-

centrations of 7.6% in December and 14.5% in February. Samples

from other Florida nurseries indicated similar increases over

winter. One nursery had a much smaller increase, from 5.4% in

December to only 7.6% in February, whereas three other nurseries

averaged about 12.5% in February.

While caution is advisable when only 1 year's data are

available, the results of the present study suggest that in terms

of carbohydrate reserves, seedling quality increases throughout

the lifting season. This fact may have a direct effect upon the

storability of seedlings as the season progresses, inasmuch as

several authors have shown that carbohydrate reserves decrease











during storage (Hellmers 1962, Ronco 1973, McCracken 1979,

Ritchie 1982). If slash pine seedlings are lifted in late

January or February, their ability to survive long-term storage

should be better than if lifted in November or December.

How the increased reserves would relate to root growth ca-

pacity upon outplanting is difficult to ascertain. If the winter

reserves can be utilized for root regeneration, then obviously

RGC will be enhanced. Root regeneration is a complex phenomenon,

however, and is influenced by the physiological controls assoc-

iated with winter dormancy. For example, Ritchie (1982) found

RGC of Douglas-fir seedlings independent of carbohydrate concen-

tration. Physiological controls such as chilling requirements

and photoperiod response are involved (Lavender 1964, Stone and

Jenkinson 1971). Therefore, even though carbohydrate may be

available for root regeneration, hormonal controls may not allow

the formation and/or expansion of primordia until specific en-

vironmental conditions are met.

Slash pine does not show the same degree of temperature and

photoperiod control (Kaufmann 1977, Fisher 1981) as compared with

more temperate species, such as Douglas-fir. The mild winters of

its native range presumably preclude the necessity for such con-

trols, and the species can physiologically respond to the sunny

days which frequent northern Florida in December through Febru-

ary. This was demonstrated by the sustained dry weight increases

throughout the winter and the lack of a well formed dormant

terminal bud.






43



The data reveal intense root activity during the winter

(Table 1-1). Therefore, no physiological "switch" needs to be

turned on to initiate root growth after outplanting. It is

plausible that root growth immediatley after planting is sup-

ported and strengthened by reserves. The amount of this support

required probably depends upon how quickly the seedling can re-

establish normal water relations and the photosynthetic process.















CHAPTER II
CHANGES IN SEEDLING MORPHOLOGY AND CARBOHYDRATE
RESERVES AFTER OUTPLANTING





Introduction



Non-structural carbohydrates contribute to plant growth via

three basic metabolic functions: (1) as a substrate for respi-

ration; (2) as the basic component of structural polysaccharides;

and (3) as carbon skeletons for other essential organic compounds

(Priestly 1962). The ultimate source of carbohydrates is photo-

synthesis, but, in the case of outplanted seedlings, the photo-

synthetic process is interrupted by root loss resulting from lift-

ing and trimming, perhaps by cold storage, by transport, handling

and planting, and finally, by an almost inevitable period of

post-planting moisture stress. Furthermore, the effects of out-

planting on photosynthetic production would be further prolonged

and intensified by dry field conditions.

Under these circumstances, reserve carbohydrates may be used

for basic metabolism, particularly respiration and cell wall for-

mation. Stored reserves can therefore directly support the root











extension necessary for survival after outplanting. If it could

be shown that seedling TNC decreases substantially upon outplant-

ing, then a strong case would be made for their contribution to

survival.

A study to follow the changes in seedling morphology and TNC

after outplanting was combined with a study of undercutting in

the nursery. Seedlings subjected to different intensities of

undercutting in the nursery have been shown to have significant

morphological (Rook 1971, Tanaka et al. 1976) and physiological

differences (Rook 1971) when compared to intact controls.

Moreover, the differences resulting from undercutting have been

associated with improved seedling performance upon outplanting

(Shoulders 1963, Rook 1971, Tanaka et al. 1976). Thus, under-

cutting was expected to produce seedlings with different morpho-

logical and physiological qualities in which the contribution of

TNC to performance after outplanting could be examined.





Materials And Methods



Nursery

The nursery phase of the experiment was carried out in the

same nursery described in Chapter I. Seed source, location, and











cultural treatments were also identical. The nursery experi-

mental design was a randomized complete block with four

treatments replicated four times. The four treatments were as

follows:

Treatment C- Control, no undercutting prior to lifting;

Treatment 1- undercut once, 2 weeks prior to lifting;

Treatment 2- undercut twice, at 6 and 2 weeks before

lifting;

Treatment 3- undercut four times, at 8, 6, 4, and 2

weeks before lifting.



Undercutting was done with a tractor-drawn bar that cut

roots about 15 cm below the bed surface. The bar lifted the bed

about 2 cm as it worked and therefore had a "wrenching" effect.

Each treatment plot was 19.8 m long and one bed wide (Figure

2-1).

The first undercutting was on November 9, 1982, 8 weeks be-

fore lifting on January 4, 1983. All treatments were undercut at

the time of lifting in accordance with standard nursery practice.

This made the total number of undercutting to be 1, 2, 3, and 5

times, for the control and treatments 1, 2, and 3, respectively.

At this time, each treatment plot was sampled by hand, pull-

ing 30 seedlings from each of four equally spaced subplots

(Figure 2-1). Each subplot covered a different two rows of the













R4 5- 65'- R3

2 1 3 C 1 3 C 2


1 3 C 2 3 2 1 C

R1 R2


-row 1
L j -2
3


-7


sub plot


Figure 2-1. Archer nursery root pruning study
treatment, and sample plot location


C -
1 -
2-
3-


indicating replication,


Control, no root pruning;
Root pruned 2 weeks prior to lifting;
Root pruned 6 and 2 weeks prior to lifting;
Root pruned 8, 6, 4, and 2 weeks prior to lifting.











bed and seedlings were sampled from both rows. Thus, there were

120 seedlings per treatment plot, representing all rows. The

four subplot samples were mixed. Twenty samples were separated

for laboratory analysis. The remaining 100 trees from each plot

were placed in KP bags with hydromulch and stored at 50C until

outplanting occurred 2 days later.



Field

The field experiment was hand planted with dibbles on

January 6. The design was a randomized complete block with four

replications. Nursery replications were continued in the field.

Each plot was a row of 80 seedlings spaced 0.6 m (2 feet) within

rows and 3.6 m (12 feet) between rows. The soil was a moderately

deep, well-drained loamy sand of the Jonesville series. The soil

was moist at the time of planting and the weather cool, partly

cloudy, and windy. The site had been a mature slash pine plan-

tation which had been harvested in 1981 then drum chopped, raked,

burned and bedded during the summer of 1982. Weed control was

generally good.

After outplanting, five seedlings per row were sampled every

2 weeks for 12 weeks. The seedlings were carefully excavated,

attempting to leave the root system intact. Each five tree sample

was used for laboratory analysis. After the 12 week period, the

remaining 50 trees per row were tallied for survival at 13 weeks

and survival and height at 1 year. Daily rainfall was measured at

a fire tower located 2 miles east of the planting site.











Laboratory

Laboratory procedure for all samples was identical to that

of Chapter I, with two exceptions. At each sampling time, the

unsuberized roots of each seedling were cut off to obtain a total

dry weight of new roots. Also, the percent moisture of all sam-

ples was determined after drying for 1 hour at 70C, and 23 hours

at 1050C. Root/shoot ratio was calculated on a dry weight basis.



Statistical Analysis

The General Linear Model procedure of the Statistical Analy-

sis System (SAS) (Freund and Littel 1981) was employed for com-

parisons between replications, treatments, and sample times. A

split plot design was used to compare times, treatments, and

times x treatment interaction during the 12 week sampling period

after outplanting (Table 2-1). A randomized complete block de-

sign was used for comparing treatments at the time of lifting.

The "Contrast" statement of SAS was used to make orthogonal com-

parisons between treatment means and time means for linear, quad-

ratic, and cubic models.

Post-outplanting field samples of the undercutting experi-

ment were being taken simultaneously with those of the seedling

development study presented in Chapter I. Therefore, the control

treatment of the undercutting experiment was directly comparable

to the results presented in Chapter I inasmuch as they were from

adjacent nursery areas and received identical nursery care up to
















Table 2-1. Analysis of variance design.


A. For time of lifting

Source of variation df

Replications 3
Treatments 3
Error 9

Total 15


B. For all sampling times

Source of variation df


Replication 3
Treatments 3
Error A 9
Time 6
Time x treatment 18
Error B 72


Total 111


111


Total











the time of lifting. Both morphological and carbohydrate values

of the two experiments were compared for the period of time when

they were sampled simultaneously.

Furthermore, when there were no statistical differences

among undercutting treatments, all treatments were averaged to

increase the number of individuals used to compare outplanted

seedlings to nursery seedlings.

Due to a freezer malfunction, samples taken at the time of

lifting were unavailable for the purpose of carbohydrate deter-

mination. The carbohydrate levels at lifting time can be esti-

mated by using data from the December 28 and January 24 values

presented in Chapter I. These values were not used, however, in

any statistical analysis of the present experiment.



Results



Morphological Characteristics at Lifting

Height, diameter at root collar (DRC), and root/shoot

ratio--characteristics often used to assess seedling quality--

were not statistically different among any of the treatments at

the time of lifting (Table 2-2).

The control seedlings were significantly lower in both fresh

and dry weight than undercut seedlings. Treatment 1 seedlings

averaged 599 mg more dry weight than the controls. This is un-

expected inasmuch as treatment 1 was root pruned only one time, 2










52






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weeks prior to lifting. The effect was consistent for all four

replications.

No statistically significant difference in root dry weight

was found among treatments and the control. There was, however,

a significant linear increase in root fresh weight and the number

of unsuberized roots with increasing intensity of undercutting

(Table 2-2). The undercutting resulted in a profusion of new

roots. The most intensively pruned treatment had six times the

dry weight of new roots as the control (14.9 mg vs. 2.5 mg).



Morphological Changes after Outplanting

There were no significant interactions between treatment and

time after outplanting with the exception of new roots. This

fact allows comparison among treatments for combined time aver-

ages for all varieties except new roots. For example, the height

measurements for all four treatments were combined to increase

the sample size for each sampling time after outplanting.

The seedlings increased significantly in height, DRC, and

dry weight in the 12 weeks after outplanting (Table 2-3). Al-

though the data for dry weight increase fit a linear model over

the 12 weeks (Table 2-3), the actual increase was negligible up

to 6 weeks followed by an abrupt surge. The largest increase in

height and terminal bud activity began after the eighth week.

























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The dry weight accumulation of outplanted seedlings over the

12 week period was smaller than that of seedlings remaining in

the nursery (Figure 2-2). Shoot dry weights of nursery and out-

planted seedlings were essentially the same during the 12 week

study period. Root dry weight differed considerably between nur-

sery and outplanted seedlings due to an approximate 30% reduction

in root dry weight upon lifting and outplanting. As a result,

total dry weight of outplanted seedlings was below that of seed-

lings remaining in the nursery. This difference had disappeared

by 8 weeks after planting, indicating that outplanted seedling

growth rate was faster than seedlings remaining in the nursery.

While there was no dry weight loss after outplanting, seed-

lings lost water in both roots and shoots during the first 8

weeks (Figure 2-3). After 8 weeks, average percent moisture in-

creased substantially in shoots and only slightly in roots.

Analysis revealed significant differences in moisture content

among sample dates using a quadratic model for roots and shoots

(Table 2-3).

Seedlings developed significant differences among treatments

during the 12 week period after outplanting for several morph-

ological characteristics (Table 2-4). For the most part, this

was related to root growth. Nursery undercutting significantly

increased root fresh and dry weight, root/shoot ratio, number of

new roots, and root moisture content. Root growth increased lin-

early with increasing intensity of nursery root pruning.







































Figure 2-2. Average root, shoot, and total dry weight per seedling after
outplanting compared to seedlings remaining in the nursery. Values for
nursery seedlings were given in Chapter I. Bars indicate + S- Shaded
Sx
bars in the bottom indicate rainfall at the planting site for specific
dates.








4 TOTAL


SHOOT


Outplanted



ROOT

I--I I
t---i-- i-- ^


Rainfall


II 1 1


2 4 6 8


WEEKS AFTER PLANTING


Nursery


-O---o-


(Jan 6)


(Mar 31)



































Figure 2-3.
compared to
for nursery


Root and shoot moisture content of outplanted seedlings as
those remaining in the nursery. Bars indicate + S-. Values
seedlings were presented in Chapter I.










SHOOTS


Nursery o-o
Outplanted *-*


ROOTS



-KI


Nurseryo-o
Outplanted -- *


WEEKS AFTER OUTPLANTING


240,


200 L


160


120


240


0 2 4 6 8 10


12

(Mar 31)


I I


(Jan 6)






















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0 2 4 6 8 10 12

(Jan 6) WEEKS AFTER PLANTING (Mar 31)



Figure 2-4. Average dry weight of new roots per seedling after outplanting.
Bars indicate the largest S- for any one sample date.











The only treatment x time interaction concerned new root

production. Treatment 2 and 3 added new roots faster and earlier

than either treatment 1 or the control (Figure 2-4). The growth

of new roots began after four weeks in the field. This is im-

mediately before the surge in total dry weight occurring after 6

weeks (Figure 2-2). Also, new root growth occurred even while the

overall root moisture content declined (Figure 2-3).



Changes in Carbohydrate Reserves after Planting

Analysis revealed no interaction between time and treatment

for any carbohydrate variable, either concentrations or amounts.

This allowed comparison among times using combined treatment

averages and comparisons among treatments using combined time

averages.

Although mean total carbohydrate content differed apprec-

iably with treatment, only one of the differences in carbohydrate

concentration or content was statistically significant (Table

2-5). This may be a result of the relatively small sampling unit

(five seedlings) or the relatively variable field environment.

Only root starch mass was found to vary significantly by treat-

ment. This relationship was linear, with amounts of root starch

increasing with intensity of nursery undercutting.

Carbohydrate reserves changed significantly with sampling

date (Table 2-6). From January 20 (2 weeks after planting) to

March 31 (12 weeks after planting), sugar concentrations
























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decreased while starch concentrations increased. The only excep-

tion to this trend was shoot sugar concentrations which increased

from the second to fourth week after outplanting (January 20 to

February 3), then began to decline.

The values determined for root and shoot starch concentra-

tions at time zero are unreliable due to sample thawing after a

freezer malfunction. These values are shown in Table 2-6, how-

ever, because they are similar to those of nursery samples taken

near the same time (Figure 2-6). If they are accurate, they

would indicate a 44% decrease in root starch concentrations dur-

ing the first 2 weeks after outplanting.

The changes over time in carbohydrate concentrations after

outplanting were remarkably similar to the changes in nursery

seedlings during the same period (Figure 2-5, 2-6, 2-7). Starch

concentration increased and sugar concentration decreased in

both. Generally, reserve concentrations of outplanted seedlings

followed the same seasonal trends as were encountered in seed-

lings remaining in the nursery.

There were, however, two important differences between the

data sets. First, starch concentrations of outplanted seedlings

were lower than those of nursery seedlings during the first 8

weeks, whereas sugar concentrations were higher (Figure 2-5).

































Figure 2-5. Average sugar and starch concentration per seedling after
outplanting, compared to those of seedlings remaining in the nursery.
Bars represent + S- Values for nursery seedlings were presented in
Chapter I; lines here have been extended to values for December 28.
The value for starch concentration of Time 0 is actual analysis result,
but may be erroneous (see text).


~











Nursery o0-

Outplanted *- *


STARCH


I I


WEEKS AFTER OUTPLANTING


SUGARS


100



90



80


70


60


50





100



80


(Jan 6)


(Mar 31)




















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Figure 2-7. Average carbohydrate concentrations and absolute amounts
for whole seedlings after outplanting compared to those of seedlings re-
maining in the nursery. Bars represent + S- Values for nursery sam-
x
ples were presented in Chapter I; lines here have been extended to
values for December 28.














700





.600
r-o

U,
0,
E
500
,)




400


MASS


CONCENTRATION


I I


WEEKS AFTER OUTPLANTING


Nursery -o---o

Outplanted


300


150o


130

cn
E


o
p 110 -

I-
Lii
0
<_


0
(Jan 6)


(Mar 31)











Second, total carbohydrate reserves of outplanted seedlings de-

clined in both concentration and amount (Figure 2-7) during the

first 2 weeks after planting. As noted before, a 44% decrease in

root starch concentrations may have occurred in the roots during

the first 2 weeks after planting. The loss in reserves was in

part related to the 30% reduction in root mass upon lifting. The

field-planted seedlings caught up to and surpassed nursery seed-

lings with respect to total reserves after 8 weeks.



Survival and Height at 1 Year

Height growth and survival after a year in the field dif-

fered significantly among treatments. Survival of seedlings from

treatment 3 was 96% versus 72% for the controls. The relation-

ship between survival and the number of nursery undercutting was

curvilinear, with an R2 of .56 (Figure 2-8). An arc sine trans-

formation of survival percent increased the R2 to .59. Thus,

increasing the frequency of nursery root pruning significantly

increased field survival.

As noted, seedling height at the time of lifting did not

vary among treatments (Table 2-2). Yet after 1 year in the field,

control seedlings averaged 33.8 cm height and treatment 3 aver-

aged 39.7 cm height. Frequency of root pruning also improved
























































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first year height increment, R2 = .39 (Figure 2-9). The low R2

indicates considerable variability in treatment response,

however.

Correlations between field performance and morphological or

carbohydrate reserve variables were tested using a large number

of linear models. Table 2-7 presents only the models which test-

ed significantly, i.e., whether individual model coefficients are
equal to zero (Freund and Littel 1981). Even so, the 2 values

resulting from these models were all low. The maximum amount of

variability accounted for was 48% in the case of survival vs. new

root mass (Figure 2-10) and 50% in the case of new root mass vs.

total carbohydrate concentration (Figure 2-11). The scatter of

points is considerable in both cases even though a definite

linear response is discernable.

Figure 2-11 indicates a 1.1% increase in survival per ad-

ditional milligram of new roots in the 1-12 week period after

outplanting. The amount of new roots per seedling is, in turn,

related to the amount of carbohydrate reserves (Figure 2-11).

Morphological characteristics normally used as seedling quality

indicators, such as height, DRC, and root/shoot ratio, were poor

predictors of field performance.
















Table 2-7. Linear Regression Models for which the test B f 0
proved significant at alpha = .05 (Freund and Littel 1981).


1. Survival =
where x =


2. Survival =
where x =



3. Survival =
where x =


1.12 (x) + 75.8 R2 = .25
mg of new roots per seedling at lifting.


1.1 (x) + 61.2 R2 = .48
average mg of new roots per seedling
2-12 weeks after outplanting.


.51 (x) + 18.1 R2 = .28
average total carbohydrate concentration per
seedling 2-12 weeks after outplanting.


4. Increment = .28 (x) + 5.5 R2 = .31
where increment is the first year's height growth,
and x = average mg of new roots per seedling 2-12 weeks
after outplanting.


5. New Roots = .43 (x) 35 R2 = .50
where new roots = average mg of new roots per seedling
2-12 weeks after outplanting
and x = average total carbohydrate concentration per
seedling 2-12 weeks after outplanting.










100



90



Z 80


0 0


y = 1.lx + 61.2


60 r


NEW ROOTS mg/seedling dry wt



Figure 2-10. The relationship of survival at 1 year after outplanting to
average mass of new roots per seedling during the first 12 weeks after
outplanting. Each point is an average of 24 composite samples of five
seedlings and includes six sampling dates.




















































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Discussion



The lack of morphological variation between treatments at

the time of lifting was due to two factors: (1) the treatments

were applied in November-December when top growth had slowed

(Chapter I); and (2) the 8 week time lapse was not sufficient

enough for statistically significant differences to develop.

Studies that have found significant morphological variation due

to undercutting involved actively growing seedlings and treat-

ments applied over a several month period (Rook 1971, Tanaka et

al. 1976). In fact, radiata pine seedlings must be root wrenched

when they are actively growing to achieve the desired morpholo-

gical and physiological modifications (Rook 1971).

Root dry weight was unaffected by the intensity of root

pruning (Table 2-2), which concurs with studies of radiata pine

(Rook 1971) and loblolly pine (Tanaka et al. 1976). In the

present study, the form of roots was modified rather than total

weight. November and December is a period when seedlings were

actively adding root weight (Chapter I). In undercut seedlings

this increase occurred on 70% of the original root mass, inasmuch

as 30% had been lost at the first undercutting (Figure 2-2).

The more intensive root pruning treatment distributed this in-

crease among a larger number of new roots (Table 2-2). The

result of the various undercutting intensities was to produce

seedlings of similar dry weight but different root morphologies.











Nursery root pruning caused a profusion of new roots. Root

auxin and cytokinin activity are increased by root pruning (Carl-

son and Larson 1977). It is the specific balance of these two

hormones that controls root primordia initiation (Altman and

Wareing 1975). Undercutting produced more new roots and root

primordia which could expand upon outplanting. The effect of

nursery undercutting continued in the field where intensively

undercut seedlings produced new roots faster and in greater quan-

tities than controls (Figure 2-4).

Morphological and physiological development after outplant-

ing closely followed that of undisturbed seedlings in the nur-

sery. Apparently, the degree of "transplant shock" experienced

by the seedlings in this experiment was not great. The results

clearly show that growth can begin shortly after planting. The

large dry weight increases between the fourth and sixth week

after planting (Figure 2-2) indicate that photosyntehtic capacity

must have been restored. In fact, increases in TNC indicate that

net photosynthesis began soon after the second week in the field.

There was only 2.5 mm of rainfall during the first 2 weeks after

planting. The increase in TNC after 2 weeks in the field

coincided with several good rains (Figure 2-7).

Caution is adviseable when interpreting results from a sin-

gle year study. Lower rainfall or antecedent soil moisture might

have limited the ability of seedlings to re-initiate carbon as-

similation. Furthermore, lower soil temperature might well have

slowed root expansion (Nambiar 1979).











The seedlings did not put out roots at the expense of

shoots. Both roots and shoots added dry weight at the same rate

they would have in the nursery. One might expect the plant to

direct photoassimilates to restore root systems damaged as a re-

sult of the lifting and planting process. It has been shown that

undercut seedlings may translocate a larger proportion of photo-

assimilated 140C to their roots than untreated controls (Rook

1971, Bacon and Bachelard 1978). Such redirection to roots did

not occur after outplanting in the present study. In fact, when

dry weight of outplanted seedlings equaled that of nursery seed-

lings at about 8 weeks after planting, it was due to the large

increases in shoot mass, inasmuch as root mass was still less

than that of nursery seedlings (Figure 2-2). During the 12 week

period of this study, root dry weight increased by 0.39 g, while

shoot dry weight increased by 0.70 g (Table 2-3).

The results of this study concur with the hypothesis that

root growth potential is a good indicator of field performance

(Sutton 1980) and is a better indicator than the morphological

characteristics generally used for grading seedlings. The amount

of new roots found on seedlings during the 2 to 12 week period

after outplanting correlated with both survival and height incre-

ment (Table 2-7). No morphological variable, including root/-

shoot ratio, showed any correlation with performance.











A certain amount of the unaccounted variability in the

regression of first year height increment and new roots (R =

.48), is a result of site conditions. The experimental area

covered 0.17 ha (.44 ac) and subjected seedlings to a range of

microsites, arising from varying degrees of vegetative com-

petition, logging debris, and water retention. Furthermore, the

soil is sandy and well-drained. Bedding was a result of standard

operating procedure rather than a site specific recommendation.

Bedding dry sites may in fact decrease first year height incre-

ment (Broerman et al. 1981). Water stress, especially during the

late summer, could have caused the relatively small seedling

height increment during the first year.

It cannot be concluded from this study that RGP is dependent

upon carbohydrate reserves. Although the amount of new roots was

positively correlated to the total carbohydrate concentrations

averaged over the 2 to 12 week period after outplanting (Figure

2-11), this is not to say that one causes the other.

There was no indication that carbohydrate reserves were used

to support root or shoot growth for more than a very brief period

after planting. Although TNC decreased during the first two

weeks after planting, it increased thereafter. The seedling re-

established its normal wintertime pattern of carbohydrate ac-

cumulation (Figure 1-9) before substantial growth occurred. As

outplanted seedlings caught up and then surpassed nursery











seedlings in total dry weight, they also surpassed them in total

reserves. The uncrowded, sunny conditions of field planting ap-

parently became beneficial after an initial phase of reduced dry

weight accumulation. Seedlings remaining in the nursery at a

density of approximately 30 per square foot were individually

exposed to less light and intense competition for soil moisture

and nutrients.

The decline in TNC in the first few weeks after planting was

relatively small, from 370 mg of non-structural carbohydrate at

the time of lifting (based on Chapter I results) to 317 mg after

2 weeks, a decline of only 15%. Moreover, part of the decrease

is due to the loss of 30% of seedling root mass and its included

reserves during lifting and outplanting. Nevertheless, an im-

portant part of the decline must have been due to utilization by

the seedling, as is evidenced by a 19% decrease in TNC concen-

trations (Figure 2-7).

The loss of carbohydrate concentration involved only the

starch fraction, specifically root starch. The simultaneous

increase in seedling sugar concentrations indicates that starch

was being transformed to sugar (Figure 2-5). The reason for the

increase in sugar concentrations is unclear.

The transformation of starch to sugar may be related to the

internal water balance of the seedling. Water loss or drought is

generally regarded as the most serious cause of seedling mor-

tality (Wakeley 1954, Kozlowski 1979). A substantial part of the











absorbing root surface is lost during lifting. Storage, trans-

port, and planting may further desiccate roots. Hence, control

of subsequent water loss is essential for seedling survival.

During the first 8 weeks after outplanting, the moisture content

of seedlings declined (Figure 2-3). The simultaneous increase in

sugar concentrations suggested the possibility of free sugars as

osmoregulators to check water loss and maintain a positive turgor

potential. The role of free sugars as osmoregulators has been

discussed by Hsiao et al. (1976) and Turner and Jones (1980).

A capacity for osmotic adjustment would be highly advanta-

geous for outplanted seedlings. By lowering cell osmotic

potential, turgor and turgor-dependent processes can be main-

tained. The result is continued cell enlargement and growth,

open stomata, and photosynthesis at water potentials which would

otherwise by limiting (Kramer 1983). As noted previously, net

photosynthesis presumably began between the second and fourth

week after outplanting in a period when the seedlings were still

losing water.

The results suggest the sequence of events after outplanting

to be: (1) an immediate transformation of starch to sugars used

for respiration, metabolism, and possibly osmoregulation; (2) re-

initiation of photosynthesis when the internal moisture condition

is favorable; (3) re-establishment of the normal seasonal carbo-

hydrate accumulaton; (4) expansion of new roots from existing

root tips and meristems in various stages of development, both











more abundant in seedlings previously root pruned; and (5) shoot

expansion. The critical role of carbohydrate reserves is during

the first step when an unfavorable water balance prohibits photo-

synthesis. By their osmoregulatory ability, carbohydrates can

contribute to a favorable internal water potential. The extent

of this contribution would depend upon the severity and extent of

water deficits.

Nursery undercutting improved seedling performance by

affecting one or more of these steps. Upon undercutting, a large

portion of the seedling root absorption system is lost. The re-

sulting water deficits would initiate the conversion of starch to

sugar for the osmoregulatory process. Repeated undercutting

could result in stable high cell osmotic potentials. These seed-

lings would be more resistent to water loss when outplanted. A

second effect is the increased number of root tips and more fi-

brous root system of undercut seedlings. This would facilitate

water uptake after planting (Rook 1971, Tanaka et al. 1976, Bacon

and Bachelard 1978). Nursery undercutting therefore has the po-

tential to increase seedling performance by increasing water ab-

sorbing surface area, and by changing seedling physiology so as

to decrease water loss due to desiccation.














CHAPTER III
THE EFFECT OF WATER STRESS ON SEEDLING MORPHOLOGY
AND CARBOHYDRATE PHYSIOLOGY



Introduction


Desiccation after outplanting is the principal cause of

initial seedling mortality during plantation regeneration

(Wakeley 1954, Kozlowski 1979, Xydias et al. 1981). The defin-

ition of a quality seedling necessarily includes the ability to

successfully withstand drought conditions that frequently occur

after outplanting. Furthermore, lifting, storage, transport, and

planting may also contribute to seedling desiccation.

Seedling morphology is important to drought resistance. For

example, low root/shoot ratios are considered to indicate a low

ratio of absorbing surface to transpiring surface. But morpho-

logical characteristics do not necessarily indicate potential

seedling performance after outplanting (Wakeley 1949, Stone 1955,

Blair and Cech 1974), and it is generally accepted that seedling

quality is a function of physiological condition (Brissette et

al. 1981).

Reserve carbohydrate, particularly root starch, has been

proposed as an important determinant of physiological quality

(Wakeley 1949, Hellmers 1962, Ronco 1973, Barnard et al. 1981).











These reserves would be essential to sustain metabolism such as

respiration in the case where post-outplanting moisture stress is

sufficient to close stomates and reduce carbon fixation. It is

also possible that sugars act as osmoregulators to maintain tur-

gor potential and allow water absorption at lower soil mositure

contents (Hsiao et al. 1976, Osonubi and Davies 1978, Kramer

1983).

The present study followed the effect of different levels of

water stress on the development of morphological characteristics

and carbohydrate reserves of slash pine seedlings in the nursery.

The objective was to determine changes in morphological and car-

bohydrate variables and their possible correlation with seedling

quality as determined by survival and growth after outplanting.





Materials And Methods

Nursery

The experiment was executed in the area of the Archer,

Florida, nursery previously described in Chapter I (see also

Figure 1-1). The experiment required complete control of the

amount of water received; therefore, the entire experiment was

sheltered by a frame covered with 6 mil (0.006 inch) clear

polyethylene plastic. The structure covered three beds, with











only the middle one used for the experiment (Figure 3-1). It was

designed so that air movement was not restricted and temperatures

at the seedling level were as close to those outside the struc-

ture as possible.

The experimental design was a randomized complete block rep-

licated three times (Figure 3-2). Individual plots were 90 cm

long separated by a vertical sheet of plastic extending from 2 cm

above the bed surface to 30 cm below. This barrier served to

restrict the lateral movement of water between plots.

The treatments were defined according to the frequency of

watering as well as the total amount received during the 8 week

experiment period: December 7, 1982, to January 31, 1983. The

five treatments were as follows:


Treatment

Treatment

Treatment


Treatment 4-



Treatment 5-


watered once at 8 weeks before lifting;

watered twice, at 8 and 4 weeks before lifting;

watered three times, at 8, 5 and 1 week before

lifting;

watered four times, at 8, 6, 4, and 2 weeks

before lifting;

watered each week.


Each application was equivalent to a 2.5 cm rain (28

L/plot). The water was applied using four buckets which slowly

dripped out the measured quantity of water over a period of 20 to






























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30 minutes while suspended over a single plot. Because of this

long application period and the porous nature of the soil, vir-

tually no water was lost due to surface runoff. The total am-

ounts of water applied were equivalent to 25 mm, 50 mm, 75 mm,

100 mm, and 200 mm rainfall for treatments 1-5, respectively.

A single porous ceramic tensiometer was placed in each plot

at 15, 30, or 45 cm depth. Each treatment had a tensiometer at

each depth randomly assigned to one of the three replications

(Figure 3-3). Tensiometers were installed on December 15 and

read weekly thereafter immediately before the scheduled water

applications.

Total stem water potentials were determined using the pres-

sure bomb. Whole seedlings were used and all determinations were

pre-dawn. A number of measurements were made over the duration

of the experiment for different treatments at different times.

At the time of lifting, 2.5 cm diameter soil cores were ta-

ken at the specific depth of each tensiometer cup and enclosed in

double plastic bags until gravimetric determination of moisture

content.

On January 31, twenty seedlings were lifted from rows 3, 4,

5, and 6 in each plot using a shovel. The trees were lifted by

cutting laterally through the plot at 17-20 cm depth to simulate

the undercutting of normal nursery lifting procedures. Five

trees in each row were taken for laboratory analysis (20 total).