AN ABSTRACT OF THE THESIS OF
Glen L. Creasy
Horticulture
for the degree of
presented on
Master of Science
in
August 5, 1991 .
Title: Xylem Discontinuity in Vitis vinifera L. Berries.
Abstract approved:
Dr. Porter B. Lombard
Several effects of xylem discontinuity in Pinot noir
and Merlot grape berries were studied.
There was a
reduction in the amount of apoplastic dye (Eosin Y or
azosulfamide) uptake through cut pedicels into soft
versus firm berries, suggesting a reduction in maximal
xylem flow at that time.
Both greenhouse and field grown
Pinot noir berries took up less dye after softening.
Merlot berries, collected on one date from the field and
separated by hand into four developmental categories,
took up different amounts of azosulfamide dye according
to category.
Soft green and just colored berries took up
3 0%, and fully colored berries 70%, less dye than firm
green berries.
The reduction in xylem conductivity was
related to the developmental stage of each individual
berry and not the cluster as a whole.
The accumulation of K+ (primarily a phloem
transported element) and Ca2+ (primarily a xylem
transported element) differed in field grown Pinot noir
berries during maturation.
On a per berry basis K+
increased after veraison, suggesting greater phloem
activity, but Ca
content remained stable after
veraison, suggesting little xylem activity.
Berry diameters on pre-veraison clusters on well
watered vines increased slightly; those on unwatered
vines decreased, losing 0.87 mm in diameter during day 3
of the experiment.
Pre-veraison berry deformabilities
were 380% higher in unwatered versus watered vines on day
4.
Bagging pre-veraison clusters to slow transpiration
reduced the rate of berry diameter loss and softness only
slightly.
Pre-veraison berry shrivelling occurred before
vine wilting.
Post-veraison Pinot noir berry diameters
and deformabilities in the greenhouse were not
significantly affected by short term vine water stress.
Heat girdling cluster peduncles to block phloem flow
reduced pre-veraison berry growth rates to near zero and
increased the rate of diameter loss significantly in
post-veraison berries.
Girdling had little effect on
pre-veraison berry deformabilities, but a large effect on
those of post-veraison berries.
The different berry responses to vine water stress
and peduncle girdling before and after veraison suggested
a change in xylem activity.
Prior to veraison there was
rapid water loss from the berry to a stressed vine, but
after veraison berries were more isolated, showing little
response to vine water stress.
Blocking phloem transport
in the cluster peduncle prior to veraison reduced berry
growth to almost zero but affected deformability little,
which suggested that xylem was maintaining berry size and
firmness.
Under the same conditions post-veraison
berries lost both size and firmness rapidly, which
suggested little xylem activity.
Xylem Discontinuity in Vitis vinifera L. Berries
by
Glen L. Creasy
A THESIS
submitted to
Oregon State University
in partial fulfillment of
the requirements for the
degree of
Master of Science
Completed August 5, 1991
Commencement June 1992
APPROVED:
Professor of Horticulture in charge of major
Head of Department of Horticulture
-y-
Dean of Grad/date School
Date thesis is presented:
August 5, 1991
Typed by Glen L. Creasy for
Glen L. Creasy
For my parents,
to whom I owe everything
ACKNOWLE DGEMENTS
The culmination of this Masters degree program could
not have come about without friendship, help, and support
from a myriad of sources. Thanks go to my committee
members, Drs. Porter Lombard, Mina McDaniel, Pat Breen,
and Don Grabe, for their efforts in the production and
review of this manuscript. I am also grateful for
financial support from the Oregon wine industry and
especially the Weatherspoon Viticulture Fellowship.
Their generous contributions have allowed me to explore
and experience Oregon and the PNW in all its splendor.
My thanks go to the downstairs morning coffee group
for their lively discussions, whose subjects ranged from
sublime to scientific to just plain silly. Many thanks
also to the past and present office and accounting staff
in the department, who always have a cheery "hello" to
spare.
I thank especially Porter, Pat, and Steve Price for
friendship, guidance, and discussion.
To Cathy, Cheryl, Kirk, Nance, Bernadine, and all
the rest (too numerous to mention), thanks for the
laughs, good times, and moral support, without which I
most certainly would have been reduced to a mental
vegetable by now.
I could not possibly omit thanking my parents, to
whom this thesis is dedicated. Without their
understanding, encouragement, guidance, and financial
support, I could never have come this far.
TABLE OF CONTENTS
Page
Chapter 1.
Introduction
1
Chapter 2.
Literature Review
4
Grape Berry Development
Berry Structure
Berry Growth
Berry Expansion and Skin
Extensibility
Chemical Changes
4
5
6
9
9
Veraison
10
Sequences of Events Near Veraison .... 11
Berry Softening
12
Vascular Changes
13
Xylem: Structure and Function
16
Contribution to Wine Quality
18
Flow Reversal - Physical Evidence .... 19
Discontinuity in Grape
20
Discontinuities in Other Fruits
20
Xylem and Water Relations in Vitis
spp
24
Xylem Discontinuity and Sugar
Accumulation
25
Xylem Reversal Revisited
26
Berry Cuticle, Transpiration, and
Mineral Content
Waterberry (implications of recent
findings)
27
29
Chapter 3.
Xylem Discontinuity In Pinot noir and
Merlot Grapes: Effects on Dye Uptake
and Mineral Composition During Berry
Maturation
32
Abstract
Introduction
Materials and Methods
Results
32
33
36
41
Discussion
Literature Cited
44
59
Heat Girdling and Vine Water Stress
Affects Pre- and Post-veraison Grape
Berry Growth and Deformability
61
Abstract
Introduction
Materials and Methods
Results
Discussion
Literature Cited
61
63
65
68
70
78
Chapter 4.
Bibliography
81
LIST OF FIGURES
Figure
Page
Chapter 3
3.1
Time series for perfusion of 5% (w/v)
aqueous Eosin Y into pre-veraison Pinot
noir berries collected from the
greenhouse. Regression equation (and r2
value) for 0.5 to 6 hours is given in the
figure. See text for calculation of
extracted dye absorbance.
(Expt. 1)
49
3.2
Dye extracted from greenhouse grown Pinot
noir berries after perfusion in 5% (w/v)
aqueous Eosin Y as a function of fruit
development. Dye uptake was through cut
berry pedicels for 6 hr. Berry softening
occurred 56 days from first bloom and
veraison on day 60 (arrows). See text for
calculation of extracted dye absorbance.
Bars indicate SE.
(Expt. 1)
50
3.3
Pre-softening Pinot noir berries perfusing
1% (w/v) aqueous azosulfamide solution.
Berries collected from the greenhouse.
51
3.4
Post-softening Pinot noir berries perfused
in 1% (w/v) aqueous azosulfamide solution
for five hours and cut longitudinally to
show the uneven dye uptake. Berries
collected from the greenhouse.
52
3.5
Pre- (left) and post- (right) softening
Pinot noir berries perfused in 1% (w/v)
aqueous azosulfamide solution and cut to
show differences in dye uptake. Berries
collected from the greenhouse.
53
3.6
Diameters of eight Pinot noir berries
grown at Woodhall III vineyard (Alpine,
Oregon) in 1990. Softening for all but
one occurred before or coincidentally with
the start of Stage III.
(Expt. 2)
54
3.7
Time series for perfusion of 1% (w/v)
aqueous azosulfamide into pre-veraison
Pinot noir berries collected from Woodhall
III vineyard in 1990. Regression equation
55
(and r^ value) for 1 to 12 hours is given
in the figure. See text for calculation
of relative absorbance.
(Expt. 2)
3.8
Relative absorbance of dye extracted from
perfused Pinot noir berries collected from
Woodhall III vineyard. Softening occurred
68 days from first bloom (upward pointing
arrow), and veraison on day 71 (downward
pointing arrow). Berries perfused in 1%
(w/v) aqueous azosulfamide dye for five
hours. See text for calculation of
relative absorbance. Bars indicate SE.
(Expt. 2)
56
3.9
Berry deformability and relative
absorbance of dye extracted from Merlot
berries separated into four developmental
stages and perfused in 1% (w/v) aqueous
azosulfamide for five hours. Berries
collected from Woodhall III vineyard on 18
Sept., 1990. See text for calculation of.
relative absorbance. Bars indicate SE.
(Expt. 3)
57
3.10
Developmental changes in Pinot noir K+ and
Ca
content per berry in 1989 (a) and
1990 (b). Veraison in the field occurred
68 days from first bloom in 1989 and 71
days from first bloom in 1990 (arrows).
Berries were collected from Woodhall III
vineyard.
(Expt. 4)
58
Chapter 4
4.1
Leaf water potentials of watered and
unwatered potted Pinot noir vines during a
water stress experiment. Means of three
to five measurements. Bars indicate SE.
74
4.2
Berry growth rates on pre- (a) and post(b) veraison greenhouse Pinot noir watered
or unwatered vines. Means of 9 berries.
Bars indicate SE.
75
4.3
Berry deformabilities on pre- (a) and
post- (b) veraison greenhouse Pinot noir
watered or unwatered vines. Points for
unwatered, bagged pre-veraison clusters
76
are means of 6 berries; all others are
means of 9 berries. Bars indicate SE.
4.4
Growth rates (a) and deformabilities (b)
of pre- and post-veraison berries on
ungirdled, girdled, and detached
greenhouse Pinot noir clusters. Means:
pre-veraison ungirdled = mean of 90
measurements on 30 berries; pre-veraison
girdled = 60 on 15; post-veraison
ungirdled = 90 on 15; post-veraison
girdled = 90 on 15; post-veraison detached
= 25 on 5. Bars indicate SE.
77
Xylem Discontinuity in Vitis vinifera L. Berries
Introduction
Chapter 1
Winegrapes are one of the most widely grown fruit
crops in the world, and thus are of great economic
consequence (Winkler et al., 1974).
Research concerning
viticulture is done in an effort to better understand the
intricacies of grape production.
In Oregon there is a small but growing winegrape
industry producing about 3.2 metric tons of fruit
annually.
The industry is concentrated in western
Oregon's Willamette Valley, which is between the Coastal
and Cascade mountain ranges.
Pinot noir, Chardonnay, and
Riesling are the primary grape varieties planted (Miles,
1987), but fruit maturation in these varieties is often
jeopardized by rainfall near harvest.
Grape berry maturation is one of the more intensely
studied aspects of grape physiology because of its
complexity.
Grape berries are unusual in that they can
accumulate high concentrations of sugars, and do so in a
relatively short period of time (Winkler et al., 1974).
They also begin to ripen earlier in the growing season
than other fruits.
The initiation of ripening is near
veraison, a time during which there are major chemical
and physical changes in the berry (Coombe, 1976).
One of
the more recently discovered physical changes is in the
berry vasculature.
Two research groups (During et al.,
1987; Findlay et al., 1987) have found that flow through
pedicel xylem into berry decreases sharply near veraison.
In these studies with Riesling and Muscat Gordo Blanco
berries, physical breaks in xylem vessels of the berry
vasculature were evident post- but not pre-veraison.
A reduction in xylem contribution to fruit
development at this time indicates that the berry depends
primarily on phloem inputs during ripening.
Indeed,
blockage of the xylem pathway between the vine and berry
may contribute to the fruit achieving such high sugar
concentrations at maturity by redirecting photoassimilate
flow and preventing water from moving into or out of the
berry (Lang and Thorpe, 1989).
If a xylem discontinuity
exists, berry responses to vine water stress will be
different in pre- versus post-veraison berries, since the
xylem is a largely unregulated pathway for water
movement.
Also, because there is less xylem flow from
the vine to berry after veraison, and a corresponding
decrease in delivery of mineral elements carried in the
xylem, certain post-veraison disorders, such as
waterberry, may occur (Christensen and Boggero, 1985;
Morrison and lodi, 1990).
In order to clarify the effects of xylem
discontinuity on berry maturation, the objectives of this
research were: 1) determine if a decrease in dye uptake
occurs in Pinot noir and Merlot grapes and if so, 2)
determine when during berry development the decrease
takes place in relation to external, measurable factor(s)
(e.g. berry diameter change, color change, and
softening); 3) measure berry mineral compositional
changes before and after veraison; and 4) measure preand post-veraison berry responses to water stress and
blockage of phloem transport.
Data gathered should be
consistent with the hypothesized reduction in xylem flow
after veraison.
Literature Review
Chapter 2
Grape Berry Development
Grape berry maturation is a much studied topic, with
veraison (Fr. veraison, meaning first color change and
berry softening (Winkler et al., 1974)) being the most
interesting aspect, physiologically.
The grape berry has
a double-sigmoidal growth curve (Considine and Knox,
1979; Coombe, 1976; Harris et al., 1968; Nii and Coombe,
1983; Winkler et al., 1974) and is non-climacteric
(Biale, 1960; Kader et al., 1985; Peynaud and RibereauGayon, 1971).
There are three commonly described phases
of growth: anthesis and first stage of rapid growth (cell
divisions, enlargement, and pedicel growth), a period of
no or slow growth (seed maturation and testa hardening),
and a final rapid growth (cell enlargement), which are
commonly described as Stage I, Stage II, and Stage III,
respectively (Nakagawa and Nanjo, 1965; 1966; Winkler and
Williams, 1935; Winkler et al., 1974).
An over-ripe
stage can also be described, where sugars stop
accumulating, but acids continue to decline (Peynaud and
Ribereau-Gayon, 1971; Winkler et al., 1974).
Veraison is
associated with the start of Stage III, and the time of
transition from Stage II to Stage III is of most interest
in this study.
Within a period of a few days, a berry goes from a
stage of quiescence (lag phase = Stage II) to one of
rapid growth, sugar accumulation, acid metabolism, and,
for a colored variety like Pinot noir, anthocyanin
synthesis.
Varieties without anthocyanins (e.g.
Chardonnay) become translucent, losing green color
through chlorophyll degradation (Harris et al., 1968).
Berry Structure
The fruit of Vitis spp. is a true berry: a multiseeded fruit derived from a single ovary.
The grape
ovary has four ovules, and thus a maximum of four seeds
(Nelson, 1985; Winkler and Williams, 1935; Winkler et
al., 1974).
The majority of the flesh of a mature fruit
is made up of mesocarp (Harris et al., 1968; Nakagawa and
Nanjo, 1965) .
There are two categories of vascular bundles in the
berry: peripheral (= dorsal), which lie below the subepidermis, and axial (= ventral), which pass through the
axis of the berry, branch off to the seed(s), and
eventually spread out at the stylar end of the berry
(Nelson, 1985).
The pedicel attaches the berry to the cluster
framework, the rachis, and enlarges at its junction with
the berry, a region called the torus (Nelson, 1985).
The
normal abscission zone between a berry and pedicel forms
in the torus.
If a berry is pulled from its pedicel
before the abscission layer forms, the berry brush
(portions of the vascular strands and pulp from inside
the berry) remains attached to the pedicel (Coombe, 1987;
Findlay et al., 1987; Nelson, 1985).
The outer surface of the berry has important
function to berry development.
The first six to eight
layers of epidermal cells contain most of the chlorophyll
and anthocyanins and is covered by the cuticle, a thin
wax-like layer on the epidermis that protects the berry
against water loss and pathogen attack (Nelson, 1985;
Peynaud and Ribereau-Gayon, 1971; Radler, 1965a; Winkler
et al., 1974).
Berries chemically stripped of cuticular
wax have a significantly higher rate of water loss than
untreated berries, while berries dipped in liquid
paraffin (to reduce transpiration) lose water at a rate
similar to that of untreated berries (During and
Oggionni, 1986; Radler, 1965a).
Thus, the natural berry
coating is very effective at insulating the berry from
desiccation.
Berry Growth
During Stage I, berry growth is due to both cell
divisions and cell enlargement (Coombe, 1960; Harris et
al., 1968; Nakagawa and Nanjo, 1965).
The embryo
increases in size and fresh weight during this stage, but
may continue to develop throughout berry maturation
(Coombe, 1960; Nakagawa and Nanjo, 1965; 1966)
In
general, the inner cells of the berry pericarp stop
dividing before the outer cells (Nakagawa and Nanjo,
1965).
The duration of Stage I seems to be similar for
all grape varieties (Coombe, 197 6; Nakagawa and Nanjo,
1966).
Stage II is a period of reduced or no growth, the
length of which is highly variable (Coombe, 1973; 1976;
Nakagawa and Nanjo, 1966).
The slowing of growth may be
caused by competition for photoassimilates between the
seed and berry flesh, since during this time period the
testa hardens and the seed becomes mature and viable
(Cawthon and Morris, 1982; Nakagawa and Nanjo, 1965).
However, arguments that neither fruit to vine nor seeds
to flesh competition is the cause of periodicity in berry
growth have been presented (Matthews et al., 1987;
Winkler and Williams, 1935).
Berries from seedless
varieties usually have a shorter Stage II and weaker
double sigmoid shape in their growth curves, perhaps due
to the lack of competition between seed and fruit
(Coombe, 1960; Winkler and Williams, 1935).
Still,
several researchers have found that seeds are always
mature and viable before berries enter Stage III, so some
interaction is likely (Cawthon and Morris, 1982; Winkler
and Williams, 1935).
Stage III starts near veraison and is a period of
rapid cell expansion (Harris et al., 1968; Winkler et
al., 1974).
Harris et al. (1968) reported an apparent
decrease in berry cell number during the latter part of
Stage III, but dismissed it as a procedural artifact.
Jona et al. (1983) reported similar results, but
suggested that this may have been the result of cell
fusion or compaction rather than an artifact.
Berry size is influenced by seed number, with a
greater number of seeds associated with larger berries
(Cawthon and Morris, 1982; Winkler and Williams, 1935).
Placental tissue produced to replace the volume that a
seed would normally take up may be insufficient to do so,
resulting in a smaller berry (Nakagawa and Nanjo, 1965).
Cheng (1985) reports that seeds influence berry growth
through hormonal action, rather than competition.
Jona et al. (1983) found that cell number in two
cultivars was the same, yet berry sizes were different.
They then concluded that berry size was determined by
cell volume and not cell number.
Certainly, cell
expansion has a great influence on final berry size
(Coombe, 1976; Harris et al., 1968).
Berry Expansion and Skin Extensibility
Berry expansion results from a net flow of materials
into the fruit (Lang and Thorpe, 1988) .
Due to
modification of pectocellulose in cell walls, berries can
expand to accommodate volume increases (Peynaud and
Ribereau-Gayon, 1971).
Berry skin extensibility may be
one measure of the ability of a berry to increase in
volume, since the expansion rate may be limited by the
skin (Considine and Kriedemann, 1972; Matthews et al.,
1987) .
As expected, in early Stage III skin
extensibility increases greatly, but however logical it
might be for increased pressure to cause berry expansion,
Matthews et al.
(1987) found low berry turgor associated
with rapid expansion.
The rate of assimilate flow into
the berry may also be a factor determining the rate of
expansion.
Chemical Changes
Berry pH tends to rise after veraison (Al-Kaisy et
al., 1981; Carroll and Marcy, 1982; Hrazdina et al.,
1984) .
Berry acidity, which contributes to berry pH, is
mainly influenced by tartaric and malic acid content
(Hrazdina et al., 1984; Kliewer, 1966; Winkler et al.,
1974) .
When expressed on a percent basis, both rise
during Stage I (malic much more so) and fall during Stage
III (Coombe, 1987; Hrazdina et al., 1984; Winkler et al..
10
1974) .
Expressed on a per berry basis after veraison,
malic acid content falls while that of tartaric remains
constant (Carroll and Marcy, 1982).
Glucose and fructose are the predominant sugars,
with their ratio being close to one for most of berry
development (Hrazdina et al., 1984; Kliewer, 1966;
Winkler et al., 1974).
More fructose than glucose is
present in over-ripe fruit (Winkler et al., 1974).
Sucrose in Vj. vinifera berries is present in only small
amounts, though it does accumulate to a limited extent
late in Stage III ((Coombe, 1987; Hrazdina et al., 1984;
Kliewer, 1966; Nelson, 1985).
However, in V.
rotundifolia (the muscadine grape) fruit has a
significant amount of sucrose (Carroll and Marcy, 1982).
Starch is virtually non-existant in mature grape berries,
like most fleshy fruits (Coombe, 1976; Nelson, 1985).
Veraison
Originally, French viticulturalists used the term
veraison to denote first color change in clusters
(Winkler et al., 1974).
For some, the term has come to
describe the entire initial process of berry ripening,
which occurs near the beginning of Stage III (Coombe,
1973; 1980; Coombe and Bishop, 1980; Findlay et al.,
1987; Nil and Coombe, 1983).
This includes color change,
sugar accumulation, acid metabolism, decreased
11
respiration, berry softening, and rapid berry expansion
(Coombe, 1973; 1980; Coombe and Bishop, 1980; Harris et
al., 1968; Peynaud and Ribereau-Gayon, 1971; Winkler et
al., 1974).
For the purposes of this discussion,
veraison will refer to the time of first color change.
Sequences of Events Near Veraison
The initiation of different events associated with
veraison is not always consistent, and may or may not be
cultivar dependent.
Findlay et al.
(1987) found that
berry volume increases about one week after the inception
of rapid sugar accumulation, softening, and color
development, and Considine and Knox (1979) observed that
sugar increase occurs before diameter increase.
Coombe
(1960; 1980) and Winkler and Williams (1935) found
diameter and sugar concentration changes to occur
coincidentally.
Hrazdina et al.
(1984) found that sugar
accumulation starts before color development.
Coombe and
Bishop (1980) found that berry deformability usually, but
not always, increases before sugar accumulation and the
second rapid diameter increase characteristic of Stage
III.
Berry expansion is an integral part of sugar
accumulation.
Berries kept from expanding by placing
them in plastic enclosures did not accumulate sugar, but
when released from confinement expanded rapidly and
12
imported sugar (Coombe, 1973).
Similarly, berry skin may
the primary factor limiting berry expansion, since peeled
berries absorb much more water without cracking than
unpeeled berries (Considine and Kriedemann, 1972).
However, Matthews et al.
(1987) found little evidence
that skin extensibility limits berry expansion.
The sudden weight increase after veraison has been
attributed to water and not accumulated sugars (i.e.
fresh weight increased before dry weight)
Coombe and Bishop, 1980).
(Cheng, 1985;
This suggests that the
vascular tissue brings in larger amounts of water than
sugar.
However, over the course of Stage III total berry
water content decreases (Carroll and Marcy, 1982; Harris
et al., 1968).
Sugar accumulation can be a result of both transport
into and synthesis within the berry (Coombe, 1987), but
concentration is also affected by dehydration and
dilution (Coombe, 1976).
Peynaud and Ribereau-Gayon
(1971) report that in situ sugar synthesis is not the
cause of rapid solute rise in grape berries at veraison,
thus import must be the major cause.
Berry Softening
Fruit softening is used in many fruit crops to
determine picking dates (Kader et al., 1985; Westwood,
1988) , but not in grape, since it occurs many weeks
13
before harvest.
However, softening can be used as an
indicator of imminent sugar and color accumulation, and
also diameter increase.
Softening may also be implicated
in the inception of sugar accumulation in the berry
(Coombe and Phillips, 1982).
Berry firmness may decline as a result of a
reduction in cell turgor (Matthews et al., 1987) caused
by water stress or physiological tissue changes, and is
linearly and positively related to (artificially induced)
turgor pressure (Bernstein and Lustig, 1985).
Temperature may also affect berry pressure (and thus
firmness) significantly (Lang and During, 1990).
Vascular Changes
Findlay et al.
(1987) and During et al.
(1987)
discovered independently that major changes occur in the
apoplastic pathway entering the berry near veraison.
The
amount of apoplastic dyes that were taken up through cut
berry pedicels or cluster peduncles decreased greatly at
that stage of development.
Anatomical studies of berry
tissue showed that mechanical breaks in xylem vessels at
the basal end of the berry (near the berry brush) were
much more prevalent in post-veraison berries, and an
increase in their freguency corresponded with the
decrease in dye uptake and other factors associated with
veraison.
Both groups of authors postulated that breaks
14
in the xylem were caused by rapid berry expansion
following veraison.
The relatively inextensible xylem
vessels in the berry brush were stretched beyond the
breaking point, while the more extensible phloem elements
accommodated the change (During et al., 1987; Findlay et
al., 1987).
Peduncle, rachis, or pedicel vascular tissue have
ceased growth by veraison, so vascular strands in these
organs should not have been stressed enough to break at
veraison (Nii and Coombe, 1983; Theiler and Coombe, 1985;
Winkler et al., 1974).
Therefore, vascular tissue inside
the berry must be the site of the discontinuities, since
only the berry is rapidly growing. After veraison berries
tend to grow faster in girth than length, in opposition
to pre-veraison growth (Harris et al., 1968).
Thus the
peripheral strands experience different stresses after
veraison, which may contribute to breakage.
Contrary to the findings of Findlay et al.
and During et al.
(1987)
(1987), Kriedemann (1969) found that
H-glucose applied through Stage III cut Thompson
Seedless berry pedicels was distributed in both
peripheral and axial vascular bundles.
(1987) reported similar results using
Findlay et al.
14
C-sucrose
perfused through cut pedicels into pre-veraison, but not
post-veraison berries.
In these Stage III berries most
of the radioactivity was found in the brush region.
15
With regard to the
3
H-glucose uptake through post-
veraison berry pedicels (Kriedemann, 1969), it seems
unlikely that phloem could have transported so much
material into the berry so quickly, since by 1 hour
radioactivity was found throughout the berry. A possible
explanation for the seemingly high amount of flow through
the xylem of these post-veraison berries was that there
was rapid movement of water from the bathing solution
through the pedicel apoplast (excluding the xylem) and
into the berry, bringing labelled glucose with it.
However, in pedicels dipped in dye solutions for less
than 2 hours, only the vascular bundles were stained,
suggesting that materials must have moved within the
pedicel xylem to enter the berry (personal observation),
and that the pedicel apoplast (excluding the xylem) was a
poor pathway for water movement.
A post-veraison xylem
discontinuity was not consistent with the findings of
Kriedemann (1969).
Because seedless grape berries have
slightly less pronounced double sigmoid growth curve
shape than seeded berries (Coombe, 1960; Winkler and
Williams, 1935), xylem elements may not have been
stretched to the point of breaking during growth after
Stage II.
In this case, the Stage III berries of
Thompson Seedless used by Kriedemann may have had
functional xylem.
16
There is additional evidence for a drop in xylem
conductivity in that accumulation of xylem borne elements
(e.g. Ca2+) tends to level off following veraison,
whereas phloem transported elements (e.g. K+) continue to
accumulate (During and Oggionni, 1986; Hrazdina et al.,
1984; Lang and Thorpe, 1989; Morrison and lodi, 1990;
Possner and Kliewer, 1985).
Evidence supporting the hypothesis that the cause of
the xylem discontinuity is physical rather than
physiological, is that mechanical shocks (such as
dropping a berry a short distance) decreases xylem
conductivity as measured by dye uptake (Findlay et al.,
1987) .
Large water potential gradients found between the
leaves and fruit of grapevine may also suggest an
increased resistance to flow into the berry (Cheng, 1985;
Greenspan and Matthews, 1991; van Zyl, 1987).
Xylem:
Structure and Function
The main pathway for water movement in a plant is
via the xylem, in which water and dissolved materials may
pass with little resistance (Galston et al., 1980).
In
general, xylem sap moves passively from regions of less
negative water potential to those of more negative water
potential (i.e. down the water potential gradient)
(Canny, 1990; Galston et al., 1980).
17
A majority of the water supplied to the fruit passes
through the xylem.
These supplies may be incorporated
into the fruit, lost through transpiration, or shunted
back into the parent plant by reverse flow (Jones and
Higgs, 1982).
With the water carried to the fruit are
minerals necessary for cell growth (Tromp and Oele,
1972) .
Very little Ca
is carried in the phloem, thus
xylem is the primary source of this element (Hanger,
1979; Kirkby and Pilbeam, 1984; Pate, 1975; Tromp and
Oele, 1972; Wiersum, 1966).
The calcium ion is required for cell wall, middle
lamella, and pectin production and maintenance (Hrazdina
et al., 1984; Poovaiah, 1979; Poovaiah et al., 1988).
A
change in the K+/Ca2+ ratio during maturation may affect
cell membrane permeability and be involved with the
ripening process (Dvorak and Cernohorska, 1972; Poovaiah,
1979; Sacher, 1973).
A change in K+/Ca2+ ratio may also
be associated with berry softening (Lang and Thorpe,
1989; Poovaiah et al., 1988).
Positive xylem pressures may be important in
delivering Ca
to plant organs with low transpiration
rates (Guttridge et al., 1981; Tachibana, 1991).
In
strawberry and tomato, Ca2+ import into developing leaves
and fruits, respectively, is favored during the night
cycle, when xylem pressure is positive (Guttridge et al.,
1981; Tachibana, 1991).
Grape berries have a very low
18
transpiration rate due to the effectiveness of the
cuticle (During and Ogginni, 1986), but water potential
gradients within grapevines are not conducive to
producing positive root pressures during Stage III
(Zinunermann, 1983).
Ca
In opposition to inward movement,
may move out of organs through the xylem in reverse-
flow, as dictated by water potential gradients (Hanger,
1979) .
Calcium may also get into organs with low
transpiration rates through exchange movement, where
there is slow transfer of calcium ions along the
negatively charged xylem vessel walls (Armstrong and
Kirkby, 1979)•
At least one researcher (Hanger, 1979)
states that all Ca
movement is the result of exchange
reactions, and not mass flow.
But if this is true there
would be little relationship between transpiration and
fruit Ca2+ content.
In grape, there is a positive
correlation between berry transpiration rate and Ca
accumulation within it (During and Oggionni, 1986).
Contribution to Wine Quality
Grape berry mineral composition is an important wine
making consideration.
High K+ concentrations in the must
are associated with high wine pH, resulting in color and
storage problems (Cox, 1988; Hand et al., 1988; Winkler
et al., 1974).
A high juice pH can also result in
19
spoilage and fermentation problems (Winkler et al.,
1974).
Both K+ and Ca2+ can form salts of malate and
tartrate that can lower wine titratible acidity;
something that may or may not be desirable (Winkler et
al., 1974).
Both K+ and Ca2+ have been implicated in the
development of waterberry, an affliction that results in
flaccid, low sugar fruit (Branas, 1974; Fregoni et al.,
1979; Ureta et al., 1981).
Flow Reversal - Physical Evidence
There are measurable variations in both fruit and
stem diameters that may be related to xylem flow
reversal.
Apple tree stems and fruits, grape berries,
pears, and lemons have all been found to have daily
changes in volume or diameter, depending on light, heat,
or degree of water stress (Coombe and Bishop, 1980;
Huguet, 1985; Jones and Higgs, 1982; Klepper, 1968; Lang,
1990; Shimomura, 1967; Tukey, 1964).
Klepper (1968)
suggested that fruit could be used as a xylem water
potential gauge, since fruit size changes depending on
the water status of the tree.
Ca2+ has also been used to follow the movement of
water in and out of fruits.
Ca
Tachibana (1991) found that
moved diurnally in tomato fruit, and Wilkinson
(1968) found evidence that Ca2+ moved out of apple
fruits.
20
Discontinuity in Grape
Breaks in xylem vessels renders each such vessel
inactive, since a column of tension can no longer be
maintained (Findlay et al., 1987; Galston et al., 1980;
Zimmermann, 1983).
This change in vine to berry
connections has major implications on our understanding
of the maturation process (Lang and Thorpe, 1989).
Despite a major pathway for ions and water being cut off,
sugar is transported into the berry at a rapid pace;
fruit soluble solids can increase as much as 0.8% per day
in hotter areas (Al-Kaisy et al., 1981).
The rate of
sugar accumulation in a berry may be affected by vacuolar
solute concentration inside a cell, and since solute
concentration is partially regulated by the amount of
water entering the berry, a xylem discontinuity can
affect sugar accumulation (Coombe, 1980; Coombe and
Phillips, 1982).
The inability of xylem to serve as a direct, largely
unregulated pathway from the vine to the berry means
water relations between the vine and the fruit change
after the discontinuity occurs.
Discontinuities in Other Fruits
In wheat and other grasses, xylem continuity is
interrupted by modified tracheary elements, leading to
21
reduced apoplastic flow to the grain (Zee and O'Brien,
1970) .
Zee and O'Brien (1970) speculate that this may be
a device to aid solute transfer to the phloem.
However,
despite this, Martin (1982) found that both xylem mobile
only and phloem mobile only elements minerals were
translocated from senescing leaves to the grain.
In some legumes, xylem has been demonstrated to be a
two-way conduit for transport of materials to and from
fruit (Clements, 1940; Pate et al., 1977; 1985; Peoples
et al., 1985).
In most model systems, more water enters
via the phloem than is required for transpiration or
expansion, with the excess leaving through the xylem
(Pate et al., 1985; Peoples et al., 1985).
This has been
suggested for other fruits as well (Clements, 1940;
Ziegler, 1963).
Others speculate that phloem supplies
the balance of water that is not supplied by the xylem
but is necessary to meet the needs of the fruit (Pate et
al., 1977; van Die and Willemse, 1980).
Pate et al.
(1985) suggests a slightly more detailed
model in which flow through the xylem is toward the fruit
at night, when the phloem cannot supply enough water via
mass flow to satisfy
fruit transpirational requirements,
and outward during the day, when water supplied by mass
flow exceeds that needed for fruit transpiration.
Peoples et al.
(1985) proposes that xylem undergoes
periodic flow reversals, with up to 70% of the total
22
water brought in by both the xylem and phloem being
recycled back to the plant through the xylem.
All xylem
strands may reverse flow for a time, or some strands my
continue inward flow while others flow out.
Apoplastic movement of assimilates is not always
under strict plant control.
found that
1
Hamilton and Davies (1988a)
C-sucrose was exported out of heat girdled
pea peduncles.
Subsequently (1988b) they found that
material was exported through xylem in response to water
potential gradients established from rapid leaf
transpiration.
Thus, the phenomenon is a physical
response that non-selectively distributes materials.
The changing role of xylem is likely one way for a
plant to balance the water economy of a developing fruit
(Pate, 1975).
Pate et al.
(1977) working with lupin
fruit found that xylem contributes less and less to fruit
development as the fruit becomes a stronger sink.
tomato, Ho et al.
In
(1987) found that xylem contribution to
fruit development gradually falls from three weeks after
pollination to maturity.
Ninety percent of the water for
fruit growth comes from the phloem, possibly due to the
low transpiration rate of the tomato fruit.
These
researchers also speculate that reverse flow out of
tomato fruit via the xylem may have been restricted, an
idea similar to that previously suggested by Jones and
Higgs (1982) for apple fruits.
Neither group of
23
researchers proposed a mechanism for the increased xylem
resistance.
Diurnal changes in fruit volume are well documented,
and may suggest liquid flow out of fruits (Coombe and
Bishop, 1980; Greenspan and Matthews, 1991; Jones and
Higgs, 1982; Klepper, 1968; Lang, 1990; Shimomura, 1967;
Tukey, 19 64).
In apple, Lang (199 0) found that xylem
flows to the fruit reverses, especially during times of
high plant transpirational stress, suggesting that this
is a part of the tree's overall water economy.
The fruit
acts as a water buffer from which the parent plant draws
water during times of high transpirational stress: an
idea supported by the data of Huguet (1985), Greenspan
and Matthews (1991), Klepper (1968), Shimomura (1967),
Tukey (1964), and van Zyl (1987).
Lang (1990) also found
that xylem contributes less and less material as an apple
fruit matures, resulting in most Ca
early stages of fruit growth.
reported similar Ca
accumulating in the
Other researchers have
accumulation patterns (Wiersum,
1966; Wilkinson, 1968).
These factors combined with
xylem flow reversal, which may result in solute export
(Lang, 1990), make up the mineral and water economy of
the apple fruit.
24
Xylem and Water Relations in Vitis spp.
According to Lang and Thorpe (1988) there are four
water flows in a "nearly mature" grape berry that is no
longer enlarging: 1) out through the xylem, 2) in through
the phloem, 3) out via transpiration, and 4) in via
osmotic absorption.
During rapid sugar accumulation in
the berry, Lang and Thorpe (1989) speculate that xylem
backflow may be necessary to balance excess water brought
in with sugar by the phloem but not lost through
transpiration.
However, Greenspan and Matthews (1991)
find that while xylem is an important contributor to preveraison grape berry development, it is not during postveraison berry maturation.
In fact water delivery
through the xylem after veraison may not be necessary:
Findlay et al.
(1987) calculated that phloem could supply
all of the berry water requirements.
However, the
researchers did not speculate as to whether excess water
would be brought in, or where it would go if it was.
van Zyl (1987), on research on vine and cluster
water potentials, postulated that berries act as a water
reservoir, supplying water to the vine when the leaves
were under high transpirational stress and being
recharged from the soil at night.
Greenspan and Matthews
(1991) and Shimomura (1967) found that diurnal changes in
berry size were exacerbated under higher vine water
stress, but that post-veraison berries has a lesser
25
response to the same conditions, supporting the idea of
increased resistance to xylem flow after veraison.
Xylem Discontinuity and Sugar Accumulation
Xylem discontinuity may enhance sugar unloading in
the berry.
Lang and Thorpe (1986) and Lang et al.
(1986)
theorize that translocation favors regions within the
plant that have a more negative water potential.
Solute
storage in the berry apoplast (as Coombe (1976) suggests)
would contribute toward decreasing the water potential
inside the berry.
Reduced xylem flow would also
contribute to a more negative water potential because
transpirational water loss from the berry will increase
the solute concentration (Coombe, 1976).
(1989) and Matthews et al.
Lang and Thorpe
(1987) found significantly
lower water potentials in Stage III than in Stage I or II
berries.
Smith and Milburn (1980) found in Ricinus
communis that as xylem water potential (which
approximates that in the apoplast) decreases, solute flux
through the phloem increases.
Earlier theories concerning the rapid increase in
fruit soluble solids after veraison included those of
Coombe (1960), who suggested that sugars were imported
into the berry first, which then attracted water into the
berry osmotically.
Later (1980) Coombe speculated that
the sugar accumulation rate was influenced by solute
26
concentration in cell vacuoles, since an unloading
mechanism based on concentration would explain the
similar sugar accumulation rates in berries of different
sizes.
However, Coombe (1976; 1980) also observed that
berries continued to accumulate sugar on a per berry
basis after volume increases had stopped, so size
increases were not linked exclusively to sugar
accumulation.
Since berry water potentials were found to
be very negative after veraison (Cheng, 1985; Matthews et
al., 1987), in order for phloem sap to be translocated
into that sink, the sap must have a higher solute
concentration than that already present in the fruit
(Lang and Thorpe, 1989).
The high potential fruit
pressures that would be associated with high osmotic
potentials could be compensated for in grape through the
restriction of water flow to and from the fruit (Cheng,
1985; Considine and Brown, 1981).
Xylem Reversal Revisited
The reduction in xylem conductivity may also prevent
the movement of berry solutes, including sugar, out of
the berry (Findlay et al., 1987; Lang and Thorpe, 1989).
Lang and Thorpe (1989) found that exudate forced from
intact post-veraison berry pedicels was identical to
expressed berry juice, supporting their idea that a post-
27
veraison grape berry is better described as a "bag of
sugary water" than as an organized plant tissue.
Cell wall degradation at veraison (Considine and
Knox, 1979) coincides with berry softening and sudden
loss in turgor; a disruption in membrane function may be
partly responsible for these happenings (Lang and Thorpe,
1989).
These drops in berry turgor have been associated
with rapid berry expansion during Stage III (Matthews et
al., 1991).
The "breakdown of apoplastrsymplast
compartmentation" (proposed by Lang and Thorpe (1989))
allows a large negative osmotic potential difference to
build up between the vine and the berry: a potential
gradient that may help direct assimilate partitioning
(Lang and Thorpe, 1986; 1989; Lang et al., 1986; Lee,
1986).
This forms the causal link between softening and
sugar import (Lang and Thorpe, 1989), but still does not
explain what triggers the process.
Berry Cuticle, Transpiration, and Mineral Content
Grape berries have a surface coating ten times as
thick as that on a leaf, and are thus more resistant to
water loss (Radler, 1965b).
Most grape berry
transpiration occurs during the day, and as berries
mature they have significantly lower transpiration rates
(Lang and Thorpe, 1989; During and Oggionni, 1986).
However, net cuticle wax per unit area remains relatively
28
constant after veraison (Considine and Knox, 1979;
Radler, 1965b; Rosenquist and Morrison, 1988).
Berry
transpiration rate is affected by turgor, with higher
turgor associated with more water loss (Lang and Thorpe,
1988).
Post-veraison. berries, however, have low turgors
(Matthews et al., 1987), possibly explaining the
transpiration rate decrease.
Pathways for direct water loss through the berry
skin are limited, increasing the fruit's resistance to
desiccation.
Grape berries may have stomata (Peynaud and
Ribereau-Gayon, 1971) or may not (Nakagawa and Nanjo,
1965; 1966; Pratt, 1971).
Lenticels have also been
reported.to occur on grape berries (Nelson, 1985; Pratt,
1971).
A decrease in transpirationally driven xylem flow
after veraison should preclude rapid and large changes in
berry solutes normally associated with that flow (such as
Ca2+).
This view is supported by the frequent
observation that Ca
does not accumulate significantly
after veraison (During and Oggionni, 1986; Hrazdina et
al., 1984; Lang and Thorpe, 1989; Morrison and lodi,
1990; Possner and Kliewer, 1985).
However, in one experiment (During and Oggionni,
1986) cuticle-stripped grape berries accumulated Ca
after veraison in much the same pattern as that of K , a
finding that runs contrary to most.
No explanation for
29
this observation has been offered.
Nevertheless, with
few discrepancies, this trend of decreased Ca2+
accumulation following veraison is well established for
healthy grape berries.
The Ca2+ accumulation rates in tomato and apple
fruits also decrease during maturation, but Ca
does
move into these fruits later in maturation (Ho et al.,
1987; Tromp and Oele, 1972).
Lang (1990) reasons that
the reduced role of xylem later in apple maturity causes
fruit Ca
accumulation to stop.
In apple, this
phenomenon is unrelated to transpiration, since
transpirational water loss remains relatively constant
throughout fruit development.
Alternatively, the decrease in Ca
accumulation
rate in ripening berries could be due to the increased
phloem flux to the berry.
Ca
levels may fail to rise
simply because increased phloem activity decreases the
relative xylem flux, irrespective of any possible
increase in resistance to flow through the xylem (Van de
Geijn and Smeulders, 1981).
Waterberry (implications of recent findings)
Waterberry (also known as Stiellahme, dessechement
de la rafle, palo negro, bunch stem necrosis, and
shanking) is a serious problem in many viticultural areas
(Christensen and Boggero, 1985), and a recent study has
30
elucidated the differences between affected and normal
berries (Morrison and lodi, 1990).
Affected clusters
will have dark or necrotic spots on the peduncle, rachis,
and/or pedicel at any time after veraison, which, in
effect girdle the portion apical to the necrosis.
Affected berries are "watery, soft and flabby," and their
colors will be "metallic, opaque, or dull green in white
grapes and red to dark blue in black grapes" (Branas,
1974; Fregoni et al., 1979; Ureta et al., 1981; Winkler
et al., 1974).
Morrison and lodi (1990) speculate that the phloem
activity in affected berries slows, resulting in greatly
reduced berry growth at veraison.
Compared to unaffected
berries, waterberry weight increases much more slowly
after veraison.
Ca
(expressed on a concentration and
per berry basis) continues to increase in waterberries,
unlike in healthy berries, whose rapid size increase is
associated with no net Ca2+ gain.
To explain this the
authors speculate that xylem connections between the
waterberry and vine remain continuous, unlike in normal
berries, where xylem interruptions are postulated to
occur (During et al., 1987; Findlay et al., 1987).
Thus
the berries may be growing under pre-veraison xylem and
phloem inputs, which are responsible for slow and
incomplete berry maturation.
Morrison and lodi (1990)
suggest waterberry results in a modified ripening pattern
31
rather than the absence of ripening, since affected
berries still soften and their malic acid content falls.
Kasimatis (1957) reported that tyloses were plugging
xylem in pedicels of waterberries, restricting solute
movement to and from the fruit, and causing the disorder.
However, recent work by Morrison and lodi (1990) suggests
that the xylem is still functioning in affected berries.
32
Xylem Discontinuity in Pinot noir and Merlot Grapes:
Effect on Dye Uptake and Mineral Composition
During Berry Maturation
Chapter 3
Abstract
To estimate changes in xylem conductivity during
greenhouse and field grape berry maturation, apoplastic
dye (aqueous Eosin Y or azosulfamide) was allowed to
perfuse through cut pedicel ends for a fixed amount of
time.
Dye uptake in soft but not yet colored Pinot noir
berries was significantly less than that in firm green
berries.
Merlot clusters, collected from the field near
veraison and the berries separated by deformability and
color, showed a similar decrease in dye uptake.
Pinot
noir and Merlot berry deformability increased sharply
before veraison (3.5 ±1.3 SD days before for Pinot noir),
and preceded the second rapid berry expansion.
In Pinot
noir, total K+ per berry increased rapidly after
veraison, but total Ca2+ per berry did not. The decrease
in dye uptake and lack of Ca
accumulation after
veraison suggest a xylem discontinuity and coincide with
berry softening and initiation of rapid growth.
33
Introduction
Grape berries are one of the few fruits to
accumulate sugars to such high concentrations, >25%
soluble solids (SS), and with such rapidity, >0.8% SS per
day in hotter areas (Al-Kaisy et al., 1981).
Much
research has focused on berry maturation in an attempt to
understand why berries are such strong assimilate sinks
after veraison, yet many aspects of the ripening process
are still not well understood.
That xylem connections between the berry and the
vine are severed near veraison further complicates our
understanding of the ripening process.
There is direct
evidence for xylem flow interruption in that physical
gaps are found in post- but not pre-veraison berry brush
xylem, and that apoplastic dyes do not traverse this area
in post-veraison berries as readily (During et al., 1987;
Findlay et al., 1987).
Indirect evidence for a change in xylem conductivity
can be seen in berry to vine water relations.
Pre-
veraison berries are much more susceptible to drought
stress than post-veraison berries, showing greater
diurnal diameter changes, or shrinking under more severe
stress (Greenspan and Matthews, 1991; Shimomura, 1967).
After veraison, diurnal berry diameter changes are less
apparent, possibly due to restricted water flow to and
34
from the fruit (Coombe and Bishop, 1980; Greenspan and
Matthews, 1991).
A decrease in calcium accumulation after veraison
also suggests a change in xylem conductivity.
of Ca
Movement
into berries is a passive process, being brought
in with the transpiration stream, and has been used as an
indicator of cumulative xylem flow (During and Oggionni,
1986; Hrazdina et al., 1984; Lang and Thorpe, 1989;
Morrison and lodi, 1990; Possner and Kliewer, 1985).
Xylem discontinuity may also affect photoassimilate
partitioning within the vine.
A low sink water potential
may increase assimilate partitioning to that sink (Lang
and Thorpe, 1986; 1989; Lang et al., 1986; Lee, 1986).
Xylem discontinuity can lower berry water potential
indirectly, because the primary route for supplying water
for transpiration is cut off.
The berry will lose water
through the skin, concentrating the solutes inside,
decreasing the fruit water potential, and causing more
assimilates to be directed to that sink.
Thus, the
discovery of a major drop in xylem conductivity may be an
important key to understanding grape berry ripening.
While a reduction in dye uptake and physical breaks
in the berry xylem have been reported in Riesling and
Muscat Gordo Blanco berries near veraison (During et al.,
1987; Findlay et al., 1987), the exact timing of this
phenomenon in relation to other berry attributes has not
35
been reported.
Determining the relative timing of xylem
discontinuity and quantifying related effects in two red
wine grape cultivars are the objectives of this study.
Decreases in apoplastic dye conductivity into berries
were compared with changes in berry %SS, size,
deformability (a measure of berry softness), color, and
K+ and Ca2+ content.
36
Materials and Methods
Greenhouse Pinot noir Perfusion (Expt. 1):
Established
greenhouse Pinot noir vines in 3 liter pots were trained
to one upright shoot and fertilized with half-strength
Hoagland's solution twice weekly.
One or two clusters
were permitted to develop on each shoot, and berries
thinned from them to reduce cluster compactness at
maturity and allow access for diameter measurements.
First bloom date for each cluster was recorded and used
to calculate approximate age when perfused.
Diameters of
selected berries were measured with a micrometer and
recorded as a reference for comparing dye uptake and
berry growth stage.
Data were collected between 10 May
and 11 July, 1990; greenhouse temperatures varied from
170C to 260C during this time.
One or two clusters of the same age were collected
at 3 to 4 day intervals throughout berry development.
All clusters were harvested before 0900 to minimize
temperature and transpiration effects on vines.
Fifteen
berries from these clusters were used in each dye uptake
experiment.
If most berries on a cluster were soft (as
determined by increased berry deformability and flesh
translucency), only soft berries were used in that
perfusion experiment.
This same selection procedure was
followed for berries at veraison.
Berry deformability
37
was defined as the difference between berry diameter with
and without external pressure.
A micrometer supplied the
compression force through its friction stop, which was
modified to reduce pressure applied at the contact
points.
Pedicels with berries still attached were cut from
the rachis with a razor blade under distilled water.
Pedicel ends were suspended in either a 5% (w/v) aqueous
Eosin Y (an apoplastic dye (Findlay et al., 1987))
solution (ten berries - perfused treatment), or distilled
water (five berries - controls) for 6 hr.
Total time for
dye uptake (= perfusion) was determined by running an
experiment on extracted dye versus time perfused.
After perfusion, berries were rinsed with distilled
water, blotted dry, their pedicels removed, and weights
recorded.
Berries were ground with a mortar and pestle
and the dye extracted with 2.0 ml 3 0% glacial acetic acid
in a centrifuge tube.
Ground berries and solvent were
agitated for 10 sec, then centrifuged at 2400 rpm for 10
minutes to settle particulates.
Part of the supernatent
(1.6 ml) was pipetted into test tubes containing 20 ^1
30% H2O2 to bleach anthocyanins.
After 30 minutes 1.5 ml
of the extract was pipetted into semi-micro disposable
polystyrene cuvettes and absorbance at 520 nm (peak dye
absorbance) and 580 nm (background absorbance) determined
with a Shimadzu UV-160 spectrophotometer.
Absorbance
38
values for control berries were averaged, and data from
perfused berries expressed as extracted dye absorbance:
(perfused A520 - A580) - (control A520 - A580) .
Field Pinot noir Perfusion (Expt. 2):
Mature Pinot noir
vines used in this study were located in the southern
Willamette Valley (Woodhall III vineyard, Alpine,
Oregon).
Five primary clusters were sampled from one
vineyard block at 2 to 5 day intervals (starting at 49
days from first bloom) during the 1990 growing season.
Diameters of eight berries on thinned clusters in the
block were measured before 0800 at 2 to 3 day intervals
throughout the season.
Softening date (determined as in
Expt. 1) and first coloring for each of these berries was
also noted.
From the five clusters collected, 15 berries
were used for perfusion.
An experiment was run on field collected berries to
determine an appropriate length of time for perfusion in
1% (w/v) aqueous azosulfamide solution (4-sulfanyl
phenyl-2-azo-7-acetamido-l-hydroxy naphthalene 3,6disulfonic acid, disodium salt; an apoplastic dye
(Ashworth, 1982)).
Berries were perfused in azosulfamide
or water for 5 hr and the dye extracted in the same
manner as in Expt. 1.
Absorbance of extracted dye was
read at 505 nm (peak dye absorbance) and 700 nm
(background absorbance), and the data expressed on a
39
relative absorbance (RA) basis (data from control berries
averaged first),
[(perfused A505 - A700) - (control A505 - A700) ]
RA=
,
(control Asos - Am)
where "A505 - A700" values were calculated from raw
absorbance data first, and the result expressed on a per
gram fresh weight basis.
Field Merlot Perfusion (Expt. 3):
Merlot clusters from
Woodhall III vineyard were collected on one date near
veraison (18 Sept., 1990) and berries separated by degree
of softness (as determined by touch) and color into four
categories (firm-green, soft-green, just colored, and
fully colored).
Ten berries were treated as controls and
20 perfused with dye for 5 hr within each group.
Berry
deformability and weight without pedicel were measured
after perfusion but before storage at -80"C.
Approximately 3 weeks after collection berries were
removed from storage and ground frozen with a mortar and
pestle.
After the tissue was ground, %SS of each control
berry was measured with a hand-held refractometer.
was extracted and its amount measured following the
procedures described in Expt. 2.
Dye
40
Field Pinot noir Mineral Analysis (Expt. 4):
During the
1989 and 1990 growing seasons random samples of 50
primary clusters were taken (from the same block used in
Expt. 2) during fruit development and frozen at -80°C.
A
subsample of 200 berries from both years was hand sorted
to remove seedless and multi-seeded berries, weighed, and
then ground to a frozen powder using liquid nitrogen and
a Waring blender.
A 0.5 g to 0.9 g portion of the
resulting powder was mixed with 1.0 ml 10% HNO3 and ashed
at 500"C for 8 hr.
The ashed samples were diluted with
10 ml 5% HNO3 one day before K+ and Ca2+ content was
determined by atomic absorption spectrophotometry.
were expressed on a per berry basis.
Data
41
Results
Greenhouse Pinot noir Perfusion (Expt. 1):
Greenhouse
berry growth curves were double sigmoidal in shape, with
softening just after the lag phase, 56 days from first
bloom, and veraison 4 days later (data not shown).
The absorbance of Eosin Y extracted from preveraison berries in the time series experiment is shown
in Fig. 3.1.
Dye uptake was linear for up to 6 hr and
resulted in sufficiently high absorbance values after
extraction.
Pre-softening berries took up a relatively large,
but variable, amount of Eosin Y (Fig. 3.2).
There was a
large drop in the amount of dye extracted from soft
versus firm berries, and another drop in that extracted
from berries at veraison.
Pre-softening berries in all
experiments took dye up evenly in their peripheral
vascular strands (Fig. 3.3), while post-softening berries
(both uncolored and at veraison) did not (Fig. 3.4).
Dye
distribution within the berry also differed, with less
dye visible in the axial and peripheral vascular strands
of a post-softening berry as compared to a pre-softening
one (Fig. 3.5).
42
Field Pinot noir Perfusion (Expt. 2):
Seasonal changes
in berry diameters were double sigmoidal in shape.
Growth curves of eight Pinot noir berries are shown in
Fig. 3.6.
Seven of these berries softened before or on
the day that rapid berry growth resumed; veraison for all
berries occurred after the start of Stage III.
On
average, Pinot noir berries softened 3.5 (±1.3 SD) days
prior to veraison.
Azosulfamide dye uptake through cut pre-veraison
berry pedicels was approximately linear for up to 12 hr
(Fig. 3.7), but to avoid problems with pedicel end
browning and berry softening that occurred during longer
perfusion times, berries in future experiments were
perfused for only 5 hr.
Dye uptake before veraison was variable, but
relative uptake dropped significantly at veraison (Fig.
3.8).
There was no significant difference between dye
uptake in soft but uncolored berries (days 68 and 70) and
dye uptake in earlier samples.
However, there was a
visible difference in dye distribution within the berry,
as noted in Expt. 1.
Field Merlot Perfusion (Expt. 3):
Merlot berries that
were soft or colored took up significantly less dye than
firm green berries (Fig. 3.9).
The pattern of dye uptake
in pre- and post-softening berries was similar to that in
43
Pinot noir.
Berry deformability, which was higher in
more developed berry categories, was inversely related to
dye uptake .
Berry %SS was lowest in pre-softening
berries (5.5%) and highest in fully colored ones (11%),
but there was no difference in the amount of %SS rise
between the categories.
There was also no significant
difference in berry weights between categories.
Field Pinot noir Mineral Analysis (Expt. 4):
Per berry
.
...
Ca 9+ content increased
only slightly
during the time
surveyed in 1989, but per berry K+ levels rose rapidly
post-veraison (68 days from first bloom) (Fig. 3.10a).
similar post-veraison pattern of mineral accumulation
occurred in berries sampled from the same block in 1990,
where veraison occurred 71 days from first bloom (Fig.
3.10b).
Ca^
There was no clear trend for accumulation of
per berry before veraison in the 1990 data.
A
44
Discussion
Dye conductivity through the berry xylem decreased
before veraison in all perfusion experiments.
Data from
the greenhouse (Expt. 1) and Merlot (Expt. 3) studies
suggest that this decrease occurs as the berries soften,
but data from the field Pinot noir study (Expt. 2)
suggest that the decrease occurs later, at veraison.
However, a distinct change in dye distribution within the
berry after softening occurred in all the perfusion
experiments.
There is a possible explanation for the high dye
uptake in soft field Pinot noir berries, Expt 2.
3.8).
(Fig
A newly mixed batch of azosulfamide dye was used
on day 65 and later perfusions, and the fresh dye may
have perfused into berries more rapidly than the older
dye.
After repeated use during the season, the dye
solution would develop darker gel-like aggregations,
which would settle out upon standing.
If the dye were
polymerizing, the resulting larger molecules may not have
entered or moved through the xylem as readily
(Zimmermann, 1983).
If the older dye solution did not
perfuse in the same manner as that of freshly mixed dye,
the unexpectedly high amounts of dye uptake at berry
softening could be explained.
If the relative absorbance
on day 65 is taken as a new average for pre-softening
45
berries, then there is a significant drop in relative
absorbance between day 65 and 68, and another significant
drop when berries colored on day 71 (Tukey HSD, p=0.01).
Peripheral strands are outlined by dye more
completely in pre- versus post-softening berries.
The
pattern of dye visible in the peripheral strands of postsoftening berries is less regular than that of presoftening berries.
Although dye enters post-softening
berries, fewer peripheral xylem strands are visibly
stained, resulting in a more uneven distribution pattern.
Also, dye in peripheral strands of post-softening berries
diffused outward, away from the bundle, more than dye in
pre-softening berries (see Fig. 3.4), suggesting cellular
breakdown.
Consistent with this, Lang and Thorpe (1989)
theorize that during softening cell membranes are
disrupted within the berry, leading to a mixing of the
contents of the apoplast and symplast.
Axial vascular strands also stained less predictably
after softening.
Unlike in firm berries, dye in soft
ones rarely reached the stylar end (see Fig. 3.5).
Contrary to the findings of Findlay et al.
(1987), there
was no evidence of more dye accumulating in berry
pedicels before versus after veraison; the amounts of dye
in both categories of tissue did not differ
significantly.
46
The Merlot experiment, in which fruit was harvested
on the same day, demonstrated that the stage of berry
development, and not that of the cluster as a whole, was
the critical factor in the dye uptake decrease.
There
was significantly higher %SS in soft-green versus firmgreen berries, which suggested that sugar accumulation
coincided with the drop in dye uptake.
Both berry
softening and sugar accumulation occurred before
veraison, as has been reported by other researchers
(Coombe and Bishop, 1980; Hrazdina et al., 1984).
Per berry Ca
content has been reported to remain
relatively constant after veraison (During and Oggionni,
1986; Hrazdina et al., 1984; Lang and Thorpe, 1989;
Morrison and lodi, 1990; Possner and Kliewer, 1985).
However, a pre-veraison rise in per berry Ca
content
was not found here, unlike in other reports (Lang and
Thorpe, 1989; Possner and Kliewer, 1985).
This result
may have been due to the berry sorting procedure before
mineral analysis.
There was very poor Pinot noir set in
1990, which resulted in high numbers of seedless berries.
To reduce the effect of berry size and seed number on the
analysis, berries from both years were visually sorted by
size to retain only one-seeded berries.
This may have
artificially selected larger pre-veraison berries,
leading to higher Ca
levels per berry.
The average
berry weight curve for samples used in mineral analysis
47
was double sigmoidal, but it was possible that preveraison berry weights were still unrepresentatively
high.
The trend toward gradual post-veraison Ca^
accumulation in 1989 and 1990, as seen in Fig. 3.10 and
in the research of During and Oggionni (1986) may be due
to cation exchange movement along xylem vessel wall
surfaces (Armstrong and Kirkby, 1979).
Ca
The amount of
brought into berries by exchange before veraison
would be masked by the relatively massive transport of
Ca
via the transpirational stream.
Alternatively, the
limited xylem connection that exists between the vine and
fruit after veraison may still supply small amounts of
water (and thus Ca2+) to the berry.
One so far unrepeated result published by During and
Oggionni (1986) occurred in post-veraison berries treated
with an emulsion to increase their transpiration rate.
Despite normal post-veraison expansion of emulsion
treated berries, Ca
in them accumulated rapidly, and m
a pattern very similar to that of K+.
This seems to be
contrary to findings here and elsewhere that report a
reduction in xylem flow near the initiation of rapid
berry expansion (During et al., 1987; Findlay et al.,
1987).
Any increase in water flux through the berry skin
should not result in significant amounts of Ca2+ being
delivered to the fruit if the xylem flow capacity is
48
reduced.
Phloem sap contains little or no Ca2+, and is
not considered to be a medium for Ca
movement (Hanger,
1979; Kirkby and Pilbeam, 1984; Pate, 1975).
An interesting connection between xylem
discontinuity and waterberry has been published recently
(Morrison and lodi, 1990), and supports the hypothesis
that rapid berry expansion causes xylem discontinuities
in the berry.
These researchers found that waterberries
expanded at a much slower rate than unaffected berries,
and that significant amounts of Ca
berries after veraison.
accumulated m these
The accumulation of Ca
that is
associated with reduced growth in waterberries after
veraison gives strong proof that xylem connections
between the affected berry and vine are still continuous.
Follow-up research on Ca
accumulation into berries
treated to increase transpiration, further study of
waterberries, and experiments dealing with the
restriction of berry growth during Stage III and
extensibility of berry xylem could do much to elucidate
the causes of xylem discontinuity and its effects on
berry maturation.
49
0.6
0.5
g
o
o
o
8
o
Abs = -0.02 + 0.07(hrs)
r2 = 0.96
H—10
15
20
Hours of Perfusion in 5% Eosin Y
«—i—i—i—|—i-
5
25
Fig. 3.1 Time series for perfusion of 5% (w/v) aqueous
Eosin Y into pre-veraison Pinot noir berries collected
from the greenhouse. Regression equation (and r2 value)
for 0.5 to 6 hours is given in the figure. See text for
calculation of extracted dye absorbance.
(Expt. 1)
50
0.40- O
C
o
-Q
i_
0.25-
Q
0.20- -
X)
(U
o
o
X
Ld
o
0.30- -
O
tn
-O
<
0
0.35-
o
T
T
o1
I
O^
v
?
o
Jl
1
?TT
g
T
6
T
0
o
1
oT
o
0.15- 0.10- -
6
0.05- -
toO
0.00-
11111111111
15
i 111111111 111 11 11 t 1 M M M 1 M 1 1 1 1 1 1 1 1 1 HH-HII
25
35
45
55
Days from First Bloom
o
1 1 1 1 1 1 1 1 1 I
65
75
Fig. 3.2 Dye extracted from greenhouse grown Pinot noir
berries after perfusion in 5% (w/v) aqueous Eosin Y as a
function of fruit development. Dye uptake was through
cut berry pedicels for 6 hr. Berry softening occurred 56
days from first bloom and veraison on day 60 (arrows).
See text for calculation of extracted dye absorbance.
Bars indicate SE.
(Expt. 1)
51
Fig. 3.3 Pre-softening Pinot noir berries perfusing 1%
(w/v) aqueous azosulfamide solution. Berries collected
from the greenhouse.
52
Fig. 3.4 Post-softening Pinot noir berries perfused in
1% (w/v) aqueous azosulfamide solution for five hours and
cut longitudinally to show the uneven dye uptake.
Berries collected from the greenhouse.
53
Fig. 3.5 Pre- (left) and post- (right) softening Pinot
noir berries perfused in 1% (w/v) aqueous azosulfamide
solution and cut to show differences in dye uptake.
Berries collected from the greenhouse.
54
E
i_
0)
to
■*-•
E
D
1111111111111111111111 ti 1111111111111111111111111111111111111111111111111111111
45
55
65
75
85
95
Days from First Bloom
105
115
125
Fig. 3.6 Diameters of eight Pinot noir berries grown at
Woodhall III vineyard (Alpine, Oregon) in 1990.
Softening for all but one occurred before or
coincidentally with the start of Stage III.
(Expt. 2)
55
vJ -
o
2<u
o
c
o
'
JQ
9- -
o
m
o
8
r
<r
<
> 6-
Q
/*
o
_o
<u
3-
00
o
o
o
o
1
i—i—
RA= 1.47 + 0.71 (hra)
r2 = 0.76
•—i—•—i—i—i—i—i—i—i—i—i—i—i—i—i—\—i—i—i—i—
5
10
15
20
25
Hours of Perfusion in 1 % Azosulfamide
Fig. 3.7 Time series for perfusion of 1% (w/v) aqueous
azosulfamide into pre-veraison Pinot noir berries
collected from Woodhall III vineyard in 1990. Regression
equation (and r2 value) for 1 to 12 hours is given in the
figure. See text for calculation of relative absorbance.
(Expt. 2)
56
12- -
i
nJO" .
<U
u
c
o
-e 8- -
}
o
{
CO
jQ
<
(U
>
6- -
I
6
§
-§ 4-
[
(»
oc:
*
*
t
1, *
2- -
o
045
—i—i—i—i—|—i
50
1—1
-h—
60
65
70
Days from First Bloom
1—\—(1—1—4—1
55
1
1
1
1
1-
1
1—H-|—1—1—(—i—1—1—t—1—1—
75
80
Fig. 3.8 Relative absorbance of dye extracted from
perfused Pinot noir berries collected from Woodhall III
vineyard. Softening occurred 68 days from first bloom
(upward pointing arrow), and veraison on day 71 (downward
pointing arrow). Berries perfused in 1% (w/v) aqueous
azosulfamide dye for five hours. See text for
calculation of relative absorbance. Bars indicate SE.
(Expt. 2)
57
Firm Green Soft Green
1st color
full color
Fig. 3.9 Berry deformability and relative absorbance of
dye extracted from Merlot berries separated into four
developmental stages and perfused in 1% (w/v) aqueous
azosulfamide for five hours. Berries collected from
Woodhall III vineyard on 18 Sept., 1990. See text for
calculation of relative absorbance. Bars indicate SE.
(Expt. 3)
58
70
60
35
Fig. 3.10
45
80
55
90
100
65
75
85
95
Days from First Bloom
110
105
120
115
125
Developmental changes in Pinot noir K+ and
Ca2+ content per berry in 1989 (a) and 1990 (b).
Veraison in the field occurred 68 days from first bloom
in 1989 and 71 days from first bloom in 1990 (arrows).
Berries were collected from Woodhall III vineyard.
(Expt. 4)
59
Literature Cited
Al-Kaisy, A.M., A.G. Sachde, H.A. Ghalib, and S.M. Hamel.
1981. Physical and chemical changes during ripening
of some grape varieties grown in Basrah. Amer. J.
Enol. Vitic.
32:268-271.
Armstrong, M.J. and E.A. Kirkby.
1979. The influence of
humidity on the mineral composition of tomato plants
with special reference to calcium distribution.
Plant Soil.
52:427-435.
Ashworth, E.N.
1982. Properties of peach flower buds
which facilitate supercooling. Plant Physiol.
70:1475-1479.
Coombe, B.G. and G.R. Bishop.
1980. Development of the
grape berry. II Changes in diameter and
deformability during veraison. Aust. J. Agric. Res.
31:499-509.
During, H. and F. Oggionni.
1986. Transpiration und
Mineralstoffeinlagerung der Weinbeere. Vitis.
25:59-66.
During, H., A. Lang, and F. Oggionni.
1987. Patterns of
water flow in Riesling berries in relation to
developmental changes in xylem morphology. Vitis.
26:123-131.
Findlay, N., K.J. Oliver, N. Nii, and B.G. Coombe.
1987.
Solute accumulation by grape pericarp cells. IV.
Perfusion of pericarp apoplast via the pedicel and
evidence for xylem malfunction in ripening berries.
J. Expt. Bot.
38:668-679.
Greenspan, M.D. and M.A. Matthews.
1991. Developmental
control of the diurnal water budget of the grape
berry.
Presented at the 42nd Annual Meeting of the
American Society for Enology and Viticulture,
Seattle, WA.
Hanger, B.C.
1979. The movement of calcium in plants.
Comm. Soil Sci. PI. Anal.
10:171-193.
Hrazdina, G., G.F. Parsons, and L.R. Mattick.
1984.
Physiological and biochemical events during
development and maturation of grape berries. Amer.
J. Enol. Vitic.
35:220-227.
60
Kirkby, E.A. and D.J. Pilbeam.
1984. Calcium as a plant
nutrient. Plant Cell Environ. 7:397-405.
Lang, A. and M.R. Thorpe.
1986. Water potential,
translocation and assimilate partitioning. J. Expt.
Bot.
37:495-503.
Lang, A. and M.R. Thorpe.
1989. Xylem, phloem and
transpiration flows in a grape: application of a
technique for measuring the volume of attached
fruits to high resolution using Archimedes'
principle. J. Expt. Bot. 40:1069-1078.
Lang, A., M.R. Thorpe, and W.R.N. Edwards.
1986. Plant
water potential and translocation. p.193-194. In
J. Cronshaw, W.J. Lucas, and R.T. Giaquinta (eds.).
Plant Biology, Vol. 1. Phloem Transport. Alan R.
Liss, Inc. New York, New York.
Lee, D.R.
1986. Water in plant storage tissues. p.327328.
In J. Cronshaw, W.J. Lucas, and R.T. Giaquinta
(eds.). Plant Biology, Vol. 1. Phloem Transport.
Alan R. Liss, Inc. New York, New York.
Morrison, J.C. and M. lodi.
1990. The influence of
waterberry on the development and composition of
Thompson Seedless grapes. Amer. J. Enol. Vitic.
41(4):301-305.
Pate, J.S.
1975. Exchange of solutes between phloem and
xylem and circulation in the whole plant, p.451473. In A. Pirson, M.H. Zimmermann (eds.). Transport
in Plants I.
Phloem Transport. Encyclopedia of
Plant Physiology, New Series Volume 1. SpringerVerlag, Berlin.
Possner, D.R.E. and W.M. Kliewer.
1985. The
localisation of acids, sugars, potassium and calcium
in developing grape berries. Vitis.
24:229-240.
Shimomura, K.
1967. Effects of soil moisture on the
growth and nutrient absorption of grapes. Acta
Agron. Academ. Scien. Hung. Tomus.
16:2 09-216.
Zimmermann, M.H.
1983. Xylem Structure and the Ascent
of Sap. Springer-Verlag, New York.
61
Heat Girdling and Vine Water Stress Affects Pre- and
Post-Veraison Grape Berry Growth and Deformability
Chapter 4
Abstract
To elucidate the effects of developmental xylem
discontinuity in post-veraison berries, greenhouse grown
Pinot noir vines were subjected to water stress and
peduncle girdling or detaching.
Berry growth rate
(change in diameter per day) and deformability were
measured as an indicator of berry water status.
Berry
diameters on pre-veraison clusters on well watered vines
increased slightly; those on unwatered vines decreased,
losing diameter at a rate of 0.87 mm/day during day 3.
Pre-veraison berry deformabilities were 380% higher in
unwatered versus watered vines on day 4.
Bagging pre-
veraison clusters to slow transpiration had little effect
on berry growth rates and deformabilities.
Post-veraison
berry growth rates and deformabilities were not affected
significantly by vine water stress.
Girdling cluster peduncles reduced pre-veraison
berry growth (in diameter) from 0.12 to 0.03 mm/day, but
had a negligible effect on berry deformabilities.
Girdling increased post-veraison berry diameter loss from
62
0.04 to 0.09 mm/day, and increased berry deformabilities
by 50%.
The rate of berry shrinkage on a detached
cluster was almost three times higher, and berry
deformability 20% higher, than the same measurements on a
girdled cluster.
A xylem discontinuity in the berry near
veraison is consistent with these findings.
63
Introduction
The growth curve of a grape berry has a double
sigmoid shape (Coombe, 1976; Coombe and Bishop, 1980).
Much research, including that on berry water relations,
has focused on the last period of growth (Stage III),
where there is rapid sugar accumulation inside an
expanding berry (for example: Coombe, 198 0; 1987; Coombe
and Hand, 1986; During et al., 1987; Findlay et al.
1987; Lang and Thorpe, 1988; 1989).
Less research has
concentrated on pre-veraison (Stage I and II) grape
development, especially with reference to fruit water
economy (Greenspan and Matthews, 1991; Matthews et al.,
1987) .
Reports that xylem flow between the berry and the
vine is interrupted prior to veraison (During et al.,
1987; Findlay et al., 1987) challenge conventional ideas
about vine to berry relationships during ripening (Lang
and Thorpe, 1989).
The grape berry, like many other
fruits, acts as a water reservoir for the vine.
A more
negative water potential in leaves than in clusters
during the day draws water out of berries, and at night a
reversed potential gradient replenishes the water
(Huguet, 1985; Klepper, 1968; van Zyl, 1987).
Diurnal
changes in fruit diameter are thought to be a
manifestation of this phenomenon, which occurs in many
64
fruits (Huguet, 1985; Jones and Higgs, 1982; Klepper,
1968; Lang, 1990; Tukey, 1964).
Pre-veraison grape
berries also exhibit this pattern of diameter change,
which is exacerbated by decreasing vine water status
(Greenspan and Matthews, 1991; Shimomura, 1967).
Post-veraison grape berries were thought to serve a
similar role in acting as a reservoir, with the water
coming in through the phloem and exiting via the xylem
(Lang and Thorpe, 1988).
However, in comparison to pre-
veraison grape berries, post-veraison berries show lesser
or no diurnal diameter changes under normal and low vine
water status, suggesting limited water flow between the
vine and fruit (Coombe and Bishop, 1980; Greenspan and
Matthews, 1991; Shimomura, 1967).
A xylem discontinuity
near veraison would explain the pre- versus post-veraison
change in diameter fluctuations.
To investigate the effects of xylem discontinuity on
pre- and post-veraison berries several treatments were
imposed on greenhouse grown Pinot noir vines and
clusters.
Vines were subjected to water stress, or
cluster peduncles girdled or detached.
Berry growth rate
(change in diameter per day)and deformability, measured
before and after application of treatments, were used as
a measure of berry water status.
65
Materials and Methods
Greenhouse grown Pinot noir vines in 3 liter pots
were trained to one shoot with one or two clusters per
shoot.
Shoot height was maintained at 1.2 m by repeated
trimming.
Berries were thinned after fruit set to reduce
cluster compactness at maturity and to allow access for
diameter measurements.
Vines were fertilized with one-
half strength Hoagland's solution twice weekly; powdery
mildew was controlled with alternating wettable sulfur
and Bayleton sprays.
Greenhouse temperatures ranged from
17 "C (night) to 30 "C (day).
Diameter and deformability were measured between
0700 and 0800 using a 25mm micrometer with its friction
stop modified to apply less pressure during deformability
measurements.
Berry deformability was defined as the
difference between berry diameter with and without
pressure (as applied by the micrometer's friction stop).
Water Stress Effect on Berry Growth and Deformability:
Water stress was applied by withholding water, starting
on day 0.
The small pot volume meant that water
availability was extremely limited; after day 4 the vines
were visibly wilted and the experiment stopped.
Unstressed vines were watered two or three times daily to
66
ensure adequate moisture.
Vine leaf water potential was
measured with a pressure bomb on days 0, 2, and 4.
Three berries on each of three clusters were
measured in each treatment.
On unwatered vines, an
additional two clusters on one vine were enclosed with a
clear plastic bag containing a damp paper towel to
increase humidity and reduce cluster transpiration.
Heat Girdling Effect on Berry Growth and Deformability:
Berry growth rate and deformability measurements were
taken from six pre-veraison clusters on three vines.
Five berries per cluster were monitored for 3 days, after
which the peduncle of one cluster on each vine was heat
girdled.
Data were taken on all clusters for an
additional 4 days.
Cluster peduncle heat girdling was accomplished with
a bent nail and alcohol burner, a method similar to that
used by Dewey et al.
(1988).
(1987) and Hamilton and Davies
The nail was heated to a dull reddish color,
then held close to the peduncle for 10 sec.
The lower
and upper sides of the peduncle were treated in this
manner two times each.
Metal foil was wrapped around the
cluster and shoot to protect them from possible heat
damage.
Post-veraison treatments were similar.
Berries were
in late Stage III and already losing diameter.
Berry
67
growth and deformability were measured on five berries on
each of three vines for 6 days.
After the day 6
measurements, all cluster peduncles were girdled and one
cluster detached.
The cut end of the detached cluster
was covered with Parafilm and hung in the canopy.
measurements were taken for an additional 7 days.
Berry
68
Results
Water Stress Effect on Berry Growth and Deformability:
Pre-veraison berries showed a dramatic response to water
stress.
By day 2 berries on unwatered vines were visibly
shrivelled, but vines were not visibly wilted until day
4.
Water potentials of leaves on unwatered vines did not
decrease during the experiment, but those on watered
vines became less negative (Fig. 4.1).
Pre-veraison berry growth rates on watered vines
were positive throughout the experiment (Fig. 4.2a).
Pre-veraison berry deformabilities on watered vines were
low, and did not differ significantly during this time
(Fig. 4.3a).
Water stressed pre-veraison berry growth rates
decreased rapidly after cessation of watering (Fig.
4.2a).
Bagging pre-veraison clusters resulted in
slightly less negative berry diameter changes (Fig. 4.2a)
and slightly lower deformabilities by day 3 (Fig. 4.3a).
By day 4 pre-veraison berries on all unwatered vines were
extremely shrivelled, leading to very high deformability
measurements.
Growth rates of post-veraison berries on both
watered and unwatered vines decreased slightly during the
experiment (Fig 4.2b).
Post-veraison berries showed
little deformability change during the experiment, with a
69
slight decrease in watered vines and a slight increase in
unwatered vines (Fig 4.3b).
On watered vines post-
veraison berry deformabilities were about twice those of
pre-veraison berries.
Heat Girdling Effect on Berry Growth and Deformability:
Immediately following girdling, peduncle tissue became a
lighter green and by 8 hr the tissue had collapsed and
browned: observations similar to that of Dewey et al.
(1987).
Heat girdled peduncles retained rigidity and
were able to support the weight of the cluster easily.
Girdling pre-veraison cluster peduncles slowed berry
growth by 75%, to near zero (4.4a) but deformability
changed only slightly (Fig 4.4b).
Post-veraison berries
on girdled clusters lost 0.05 mm in diameter per day more
than those on ungirdled clusters (Fig. 4.4a).
Post-
veraison berry deformabilities were 50% higher on girdled
versus ungirdled clusters (Fig. 4.4b).
Girdling had a
much larger effect on berry deformabilities post-veraison
than pre-veraison.
Berry deformabilities on post-
veraison ungirdled clusters were 170% of those on preveraison clusters.
Berry diameter loss on a detached post-veraison
cluster was almost three times as high and berry
deformabilities 20% higher (Fig. 4.4) than that on a
girdled cluster.
70
Discussion
Berry growth rates and deformabilities were much
more sensitive to vine water status before veraison than
after veraison.
Pre-veraison berries showed signs of
water stress (e.g. reduced growth rates and increased
deformabilities) earlier than post-veraison berries, and
long before vines began to wilt.
Post-veraison berries
showed little signs of stress in all watered and
unwatered treatments.
Similar results were reported by
Greenspan and Matthews (1991), where diurnal changes in
berry diameter were measured.
They showed that diurnal
berry contractions were smaller in magnitude on postversus pre-veraison clusters in both potted and field
vines.
Bagging pre-veraison clusters to reduce
transpiration did little to prevent berries from
shrivelling on unwatered vines, suggesting that water
movement from the cluster to the vine, not transpiration
from the berries, is the primary mode of water loss.
Greenspan and Matthews (1991) found that bagging preveraison clusters reduces the magnitude of diurnal berry
contractions only 3 0%, while bagging whole shoots reduces
the contractions by 70%.
This suggests that shoot
transpiration is partly responsible for the diurnal
change in berry diameter.
For the pre-veraison grape,
71
the water demands of the vine appear to take precedence
over those of the berries.
Pre-veraison grapes are
highly dependent on the xylem to support growth
(Greenspan and Matthews, 1991), so factors that affect
the availability of xylem water in the vine will affect
berry water status.
Post-veraison grapes, however, are
more isolated from the vine.
In potted and field grown
vines bearing post-veraison clusters, diurnal berry
diameter changes were not affected by bagging shoots to
slow vine transpiration, suggesting that shoot
transpiration does not contribute to diurnal berry
contractions (Greenspan and Matthews, 1991).
For a berry to accumulate high concentrations of
sugars isolation from the water demands of the vegetative
organs is almost a necessity (Lang and Thorpe, 1989).
Cell wall degradation in the grape berry near veraison
results in a breakdown between apoplast and symplast
compartmentation (Considine and Knox, 1979; Lang and
Thorpe, 1989).
The resulting sugary sap that fills the
berry could be drawn out in response to vine water stress
unless there is a xylem interruption (Findlay et al.,
1987; Lang and Thorpe, 1989).
Also, if xylem flow is not
restricted, water could move from the vine to the berry
in response to water and hydrostatic pressure gradients,
thus diluting the sap (Lang and Thorpe, 1989; Lang et
al., 1986; Lee, 1986).
There has been shown to be a
72
favorable shoot to cluster water potential gradient in
ripening berries on both well watered and water stressed
vines (Greenspan and Matthews, 1991), but despite this
water does not move in and dilute berry contents.
In
terms of water flow through the xylem, post-veraison
berries seem to be isolated from the vine and no longer
serve as a water reservoir for the vine.
Berry growth
during the ripening stage is highly dependent on the
phloem and not the xylem (Greenspan and Matthews, 1991;
Lang and Thorpe, 1989), so factors affecting phloem
transport should have a significant impact on berry
development.
Heat girdling has been used extensively to block
phloem transport in plants (Clements, 1940; Dewey et al.,
1987; Hamilton and Davies, 1988; Lang, 1990; Lang and
Thorpe, 1989; Martin, 1982).
In grape a heat girdle has
two different effects, depending on at what stage the
berries are when the girdle is made.
This is because the
xylem in post-veraison berries is largely nonfunctioning.
In a berry on a girdled pre-veraison
cluster the xylem can supply enough water to support
limited growth and turgor maintenance (which is
responsible, in part, for berry firmness (Matthews et
al., 1987)).
In a post-veraison berry xylem conductivity
is much lower (Creasy, Chap. 3; During et al., 1987;
Findlay et al., 1987), so when the phloem is blocked the
73
berry shrinks and becomes softer.
Water lost through
berry transpiration cannot be replaced.
However, berries on a detached cluster shrank faster
and had higher deformabilities than berries on girdled
clusters, suggesting that berries on girdled clusters
still had xylem connections to the vine.
Studies using
dye taken up into berries show that xylem conductivity
near veraison decreases but does not stop completely,
supporting these findings (Creasy, Chap. 3; During et
al., 1987; Findlay et al., 1987).
The significant
differences in growth rate and deformability between
post-veraison berries on girdled and detached clusters
may also be due to the extra transpirational surface on
the detached cluster.
The rachis of the girdled clusters
still had a direct xylem connection to the vine, and thus
a pathway through which to satisfy its transpirational
water requirements.
The rachis on the detached vine did
not, and so may have drawn water out of the attached
berries via the phloem or remaining xylem connections,
resulting in rapid berry decline.
Cluster rachises and
berry pedicels should be coated with an anti-transpirant
to reduce this effect, or berries removed from the vine
individually and measured.
74
-0.45
O
O watered vines
• — • unwatered vines
o
a.
I
-O
1
-0.55
:s
"5
■*->
C-
-0.65
-f-*
o
CL
i-
03
*J
U
-0.75
-0.85
Day 0
Day 2
Day 4
Fig. 4.1 Leaf water potentials of watered and unwatered
potted Pinot noir vines during a water stress experiment.
Means of three to five measurements. Bars indicate SE.
75
D
X)
0.15--0.10--
E
a
X)
< -0.35--
Pre—veraison
£ -0.60
O—O watered
A — A unwatered
▲
A unwatered. bagged
o
o -0.85
CD
-1.10
o
0.15
■o
a
V——-.
o
E
^E -0.10E
"—V
-
s
-^ --
1
¥
■-
< -0.35o
DC
-0.60
o
o -0.85
CD
•1.10
—— ~ A
b
Post-veraison
O
V
O watered
V unwatered
1
1
1
1
1
2
Day of Experiment
Fig. 4.2 Berry growth rates on pre- (a) and post- (b)
veraison greenhouse Pinot noir watered or unwatered
vines. Means of 9 berries. Bars indicate SE.
76
1.5
Pre-veroison
1.2--
O
A
O wotered
A unwatared
A unwotered. bogged
A
E
E
a
0.9JQ
O
§ 0.6
o
<u
Q
0.3--
0.0
1.5
Post—veraison
1.2
O
V
O watered
V unwatensd
E
E
0.9
o
§ 0.6o
0.3-
0.0
-\
1
1
2
13
Day of Experiment
Fig. 4.3 Berry deformabilities on pre- (a) and post- (b)
veraison greenhouse Pinot noir watered or unwatered
vines. Points for unwatered, bagged pre-veraison
clusters are means of 6 berries; all others are means of
9 berries. Bars indicate SE.
77
o
0.15---
E
J,
E
T
0.05 -
1
o
I
1
T
X)
< -0.05<0
01
1
a
-0.15
*-*
s
o
6 -0.25
CD
-0.35
1
1
1
1
1
I—I—
1
^ 1.25
E
E
^ 1.00
1
1.50
-*J
I 0.75
1
!5
I
o
*
1
0.50
1
Q
0.25
1
CD
i_
CD
0.00
—I—
—I—
—I—
—I—
—I—
Ungirdled
Girdled
Ungirdled
Girdled
Detached
Pre-veraison
Fig. 4.4
Post-veraison
Growth rates (a) and deformabilities (b) of
pre- and post-veraison berries on ungirdled, girdled, and
detached greenhouse Pinot noir clusters.
Means: pre-
veraison ungirdled = mean of 90 measurements on 30
berries; pre-veraison girdled = 60 on 15; post-veraison
ungirdled = 90 on 15; post-veraison girdled = 90 on 15;
post-veraison detached = 25 on 5.
Bars indicate SE.
78
Literature Cited
Clements, H.F.
1940. Movement of organic solutes in the
Sausage Tree, Kicrella africana. Plant Physiol.
15:689-700.
Considine, J.A. and R.B. Knox.
1979.
Development and
histochemistry of the cells, cell walls, and cuticle
of the dermal system of fruit of the grape Vitis
vinifera L. Protoplasma. 99:347-365.
Coombe, B.C.
1976. The development of fleshy fruits.
Ann. Rev. Plant Physiol.
27:507-528.
Coombe, B.G.
1980.
Development of the grape berry. I
Effects of time of flowering and competition. Aust.
J. Agric. Res.
31:125-131.
Coombe, B.G.
1987.
Distribution of solutes within the
developing grape berry in relation to its
morphology. Amer. J. Enol. Vitic.
38:120-127.
Coombe, B.G. and G.R. Bishop.
1980. Development of the
grape berry. II. Changes in diameter and
deformability during veraison. Aust. J. Agric. Res.
31:499-509.
Coombe, B.G. and P.G. Hand.
1986. Grape berry
development. p.50-54. Proceedings Sixth Aust. Wine
Indust. Tech. Conf. University of California,
Davis, California.
Dewey, S.A., D.O. Chilcote, and A.P. Appleby.
1987. A
hot-wire technique for girdling stems and petioles
of herbaceous plants. Agron. J. 79:587-590.
During, H., A. Lang, and F. Oggionni.
1987. Patterns of
water flow in Riesling berries in relation to
developmental changes in xylem morphology. Vitis.
26:123-131.
Findlay, N., K.J. Oliver, N. Nii, and B.G. Coombe.
1987.
Solute accumulation by grape pericarp cells. IV.
Perfusion of pericarp apoplast via the pedicel and
evidence for xylem malfunction in ripening berries.
J. Expt. Bot.
38:668-679.
79
Greenspan, M.D. and M.A. Matthews.
1991. Developmental
control of the diurnal water budget of the grape
berry. Presented at the 42nd Annual Meeting of the
American Society for Enology and Viticulture,
Seattle, WA.
Hamilton, D.A. and P.J. Davies.
1988. Export of organic
materials from developing fruits of pea and its
possible relation to apical senescence. Plant
Physiol.
86:951-955.
Huguet, J-G.
1985. Appreciation de 1'etat hydrique
d'une plante a partir des variations micrometriques
de la dimension des fruits ou des tiges au cours de
la journee. Agronomie.
5:733-741.
Jones, H.G. and K.H. Higgs.
1982. Surface conductance
and water balance of developing apple (Malus pumila
Mill.) fruits. J. Expt. Bot.
33:67-77.
Klepper, B.
1968.
woody plants.
Diurnal pattern of water potential in
Plant Physiol. 43:1931-1934.
Lang, A.
1990. Xylem, phloem and transpiration flows in
developing apple fruits. J. Expt. Bot.
41:645-651.
Lang, A. and M.R. Thorpe.
1988. Why do grape berries
split? p. 69-71. Proc. 2nd Intl. Cool Climate
Vitic. Oenology Symp. Auckland, New Zealand.
Lang, A. and M.R. Thorpe.
1989. Xylem, phloem and
transpiration flows in a grape: application of a
technique for measuring the volume of attached
fruits to high resolution using Archimedes'
principle. J. Expt. Bot. 40:1069-1078.
Lang, A., M.R. Thorpe, and W.R.N. Edwards.
1986. Plant
water potential and translocation. p.193-194.
In
J. Cronshaw, W.J. Lucas, and R.T. Giaquinta (eds.).
Plant Biology, Vol. 1. Phloem Transport. Alan R.
Liss, Inc. New York, New York.
Lee, D.R.
1986. Water in plant storage tissues. p.327328.
In J. Cronshaw, W.J. Lucas, and R.T. Giaquinta
(eds.). Plant Biology, Vol. 1. Phloem Transport.
Alan R. Liss, Inc. New York, New York.
Matthews, M.A., G. Cheng, and S.A. Weinbaum.
1987.
Changes in water potential and dermal extensibility
during grape berry development. J. Amer. Hort. Sci.
112:314-319.
80
Martin, P.
1982. Stem xylem as a possible pathway for
mineral retranslocation from senescing leaves to the
ear in wheat. Aust. J. Plant Physiol. 9:197-207.
Shimomura, K.
1967. Effects of soil moisture on the
growth and nutrient absorption of grapes. Acta
Agron. Academ. Scien. Hung. Tomus.
16:209-216.
Tukey, L.D.
1964. A linear electronic device for
continuous measurement and recording of fruit
enlargement and contraction. Proc. Amer. Soc. Hort.
Sci. 84:653-660.
van Zyl, J.L.
1987. Diurnal variation in grapevine
water stress as a function of changing soil water
status and meteorological conditions. S. Afr. J.
Enol. Vitic. 8:45-52.
81
Bibliography
Al-Kaisy, A.M., A.G. Sachde, H.A. Ghalib, and S.M. Hamel.
1981. Physical and chemical changes during ripening
of some grape varieties grown in Basrah. Amer. J.
Enol. Vitic.
32:268-271.
Armstrong, M.J. and E.A. Kirkby.
1979. The influence of
humidity on the mineral composition of tomato plants
with special reference to calcium distribution.
Plant Soil.
52:427-435.
Ashworth, E.N.
1982. Properties of peach flower buds
which facilitate supercooling. Plant Physiol.
70:1475-1479.
Bernstein, Z. and I. Lustig.
1985. Hydrostatic methods
of measurement of firmness and turgor pressure of
grape berries (Vitis vinifera L.). Sci. Hort.
25:129-136.
Biale, J.B.
1960. The postharvest biochemistry of
tropical and subtropical fruits. Adv. Fd. Res.
10:293-354.
Branas, J.
1974. Viticulture,
Dehan, Montpellier.
p.615-618.
Imprimerie
Canny, M.J.
1990. Fine veins of dicotyledon leaves as
sites for enrichment of solutes of the xylem sap.
New Phytol.
115:511-516.
Carroll, D.E. and J.E. Marcy.
1982. Chemical and
physical changes during maturation of muscadine
grapes (Vitis rotundifolia). Amer. J. Enol. Vitic.
33:168-172.
Cawthon, D.L. and J.R. Morris.
1982. Relationship of
seed number and maturity to berry development, fruit
maturation, hormonal changes, and uneven ripening of
'Concord' (Vitis labrusca L.) grapes. J. Amer. Soc.
Hort. Sci.
107(6):1097-1104.
Cheng, G.
1985. Water relations of fruit growth in
Vitis vinifera. MS thesis, University California,
Davis, California.
Christensen, L.P. and J.D. Boggero.
1985. A study of
mineral nutrition relationships of waterberry in
Thompson Seedless. Amer. J. Enol. Vitic.
36(1):5764.
82
Clements, H.F.
1940. Movement of organic solutes in the
Sausage Tree, Kigella africana.
Plant Physiol.
15:689-700.
Considine, J. and K. Brown.
1981.
fruit growth. Plant Physiol.
Physical aspects of
68:371-376.
Considine, J.A. and R.B. Knox.
1979. Development and
histochemistry of the cells, cell walls, and cuticle
of the dermal system of fruit of the grape Vitis
vinifera L. Protoplasma. 99:347-365.
Considine, J.A. and P.E. Kriedemann.
1972.
Fruit
splitting in grapes: determination of the critical
turgor pressure. Aust. J. Agric. Res.
23:17-24.
Coombe, B.G.
1960. Relationship of growth and
development to changes in sugars, auxins, and
gibberellins in fruit of seeded and seedless
varieties of Vitis vinifera. Plant Physiol.
35:241-250.
Coombe, B.G.
1973. The regulation of set and
development of the grape berry. Acta Hort.
273.
34:261-
Coombe, B.G.
1976. The development of fleshy fruits.
Ann. Rev. Plant Physiol.
27:507-528.
Coombe, B.G.
1980.
Development of the grape berry. I
Effects of time of flowering and competition. Aust.
J. Agric. Res.
31:125-131.
Coombe, B.G.
1987. Distribution of solutes within the
developing grape berry in relation to its
morphology. Amer. J. Enol. Vitic.
38:120-127.
Coombe, B.G. and G.R. Bishop.
1980. Development of the
grape berry. II. Changes in diameter and
deformability during veraison. Aust. J. Agric. Res.
31:499-509.
Coombe, B.G. and P.G. Hand.
1986. Grape berry
development. p.50-54.
Proceedings Sixth Aust. Wine
Indust. Tech. Conf. University of California,
Davis, California.
Coombe, B.G. and P.E. Phillips.
1982. Development of
the grape berry.
III. Compositional changes during
veraison measured by sequential hypodermic sampling,
p.132-136. Proc. Intl. Sym. Grapes and Wine.
University of California, Davis, California.
83
Cox, J.
1988. From Vines to Wines. Harper and Row
Publishers, Inc., New York, New York.
Dewey, S.A., D.O. Chilcote, and A.P. Appleby.
1987. A
hot-wire technique for girdling stems and petioles
of herbaceous plants. Agron. J.
79:587-590.
During, H. and F. Oggionni.
1986. Transpiration und
Mineralstoffeinlagerung der Weinbeere. Vitis.
25:59-66.
During, H., A. Lang, and F. Oggionni.
1987. Patterns of
water flow in Riesling berries in relation to
developmental changes in xylem morphology. Vitis.
26:123-131.
Dvorak, M. and J. Cernohorska.
1972. Comparison of
effects of calcium deficiency and IAA on the pumpkin
plant (Cucurbita pepo L.). Biol. Plant.
14:28-38.
Findlay, N., K.J. Oliver, N. Nii, and B.C. Coombe.
1987.
Solute accumulation by grape pericarp cells. IV.
Perfusion of pericarp apoplast via the pedicel and
evidence for xylem malfunction in ripening berries.
J. Expt. Bot.
38:668-679.
Fregoni, M., A. Scienza, M. Bosselli, R. Miravalle, and
M. Zamboni.
1979. L'appassimento del grappolo
della vite causato dal disseccamento del rachid:
nuovi contributi terapeutici. Vignevini.
6:43-46.
Galston, A.W., P.J. Davies, and R.L. Satter.
1980. The
Life of The Green Plant. Third ed. Prentice-Hall
Inc., Englewood Cliffs, New Jersey.
Greenspan, M.D. and M.A. Matthews.
1991. Developmental
control of the diurnal water budget of the grape
berry. Presented at the 42nd Annual Meeting of the
American Society for Enology and Viticulture,
Seattle, WA.
Guttridge, C.G., E.G. Bradfield, and R. Holder.
1981.
Dependence of calcium transport into strawberry
leaves on positive pressure in xylem. Ann. Bot.
48:473-480.
Hamilton, D.A. and P.J. Davies.
1988a Export of organic
materials from developing fruits of pea and its
possible relation to apical senescence. Plant
Physiol.
86:951-955.
84
Hamilton, D.A. and P.J. Davies.
1988b Mechanism of
export of organic material from the developing
fruits of pea. Plant Physiol. 86:956-959.
Hanger, B.C.
1979. The movement of calcium in plants.
Comm. Soil Sci. Plant Anal.
10:171-193.
Harris, J.M., P.E. Kriedemann, and J.V. Possingham.
1968. Anatomical aspects of grape berry
development. Vitis. 7:106-119.
Ho, L.C., R.I. Grange, and A.J. Picken.
1987. An
analysis of the accumulation of water and dry matter
in tomato fruit. Plant Cell Environ.
10:157-162.
Hrazdina, G., G.F. Parsons, and L.R. Mattick.
1984.
Physiological and biochemical events during
development and maturation of grape berries. Amer.
J. Enol. Vitic.
35:220-227.
Huguet, J-G.
1985. Appreciation de I'etat hydrique
d'une plante a partir des variations micrometriques
de la dimension des fruits ou des tiges au cours de
la journee. Agronomie.
5:733-741.
Hand, P.G., A.J.W. Ewart, D.G.R. Bruer, and C. Hartner.
1988. Effects of skin contact, pH, and SO2 on young
red wine composition. p.225-226. Proc. Second
International Cool Climate Vitic. and Oenol. Symp.,
Aukland, New Zealand.
Jona, R., R Vallania, and C. Rosa.
1983. Cell wall
development in the berries of two grapevines. Sci.
Hort.
20:169-178.
Jones, H.G. and K.H. Higgs.
1982. Surface conductance
and water balance of developing apple (Malus pumila
Mill.) fruits. J. Expt. Bot.
33:67-77.
Kader, A.A., R.F. Kasmire, F.G. Mitchell, M.S. Reid, N.F.
Sommer, and J.F. Thompson.
1985. Postharvest
technology of horticultural crops. Univ. Calif.
Coop. Exten. Special Publication 3311.
Kasimatis, A.N.
1957. Some factors influencing the
development of water berries in Thompson Seedless
grown for table use. MS thesis, University of
California, Davis, California.
Kirkby, E.A. and D.J. Pilbeam.
1984. Calcium as a plant
nutrient. Plant Cell Environ. 7:397-405.
85
Klepper, B.
1968.
woody plants.
Diurnal pattern of water potential in
Plant Physiol. 43:1931-1934.
Kliewer, W.M.
1966. Sugars and organic acids of Vitis
vinifera. Plant Physiol. 41:923-931.
Kriedemann, P.E.
1969. Sugar uptake by the grape berry:
a note on the absorption pathway. Planta. 85: Hill?.
Lang, A.
1990. Xylem, phloem and transpiration flows in
developing apple fruits. J. Expt. Bot. 41:645-651.
Lang, A. and H. During.
1990. Grape berry splitting and
some mechanical properties of the skin. Vitis.
29:61-70.
Lang, A. and M.R. Thorpe.
1986. Water potential,
translocation and assimilate partitioning. J. Expt.
Bot.
37:495-503.
Lang, A. and M.R. Thorpe.
1988. Why do grape berries
split? p. 69-71. Proc. 2nd Intl. Cool Climate
Vitic. Oenology Symp. Auckland, New Zealand.
Lang, A. and M.R. Thorpe.
1989. Xylem, phloem and
transpiration flows in a grape: application of a
technique for measuring the volume of attached
fruits to high resolution using Archimedes'
principle. J. Expt. Bot. 40:1069-1078.
Lang, A., M.R. Thorpe, and W.R.N. Edwards.
1986. Plant
water potential and translocation. p.193-194. In
J. Cronshaw, W.J. Lucas, and R.T. Giaquinta (eds.).
Plant Biology, Vol. 1. Phloem Transport. Alan R.
Liss, Inc. New York, New York.
Lee, D.R.
1986. Water in plant storage tissues. p.327328.
In J. Cronshaw, W.J. Lucas, and R.T. Giaquinta
(eds.).
Plant Biology, Vol. 1. Phloem Transport.
Alan R. Liss, Inc. New York, New York.
Martin, P.
1982. Stem xylem as a possible pathway for
mineral retranslocation from senescing leaves to the
ear in wheat. Aust. J. Plant Physiol.
9:197-207.
Matthews, M.A., G. Cheng, and S.A. Weinbaum.
1987.
Changes in water potential and dermal extensibility
during grape berry development. J. Amer. Hort. Sci.
112:314-319.
Miles, S.D.
1987. Vineyard acreage in Oregon 1986.
Wine Advisory Board Research report.
5:7-13.
The
86
Morrison, J.C. and M. lodi.
1990. The influence of
waterberry on the development and composition of
Thompson Seedless grapes. Amer. J. Enol. Vitic.
41(4):301-305.
Nakagawa, S. and Y. Nanjo.
1965. A morphological study
of Delaware grape berries. J. Jpn. Soc. Hort. Sci.
34:85-95.
Nakagawa, S. and Y. Nanjo.
1966. Comparative morphology
of the grape berry in three cultivars. J. Jpn. Soc.
Hort. Sci.
35:117-126.
Nelson, K.E.
1985. Harvest and handling California
table grapes for market. Univ. Calif. Agr. Expt.
Sta. Bui. 1913.
Nii, N. and B.C. Coombe.
1983. Structure and
development of the berry and pedicel of the grape
Vitis vinifera L. Acta Hort.
139:129-140.
Pate, J.S.
1975. Exchange of solutes between phloem and
xylem and circulation in the whole plant, p.451473.
In A. Pirson, M.H. Zimmermann (eds.).
Transport in Plants I. Phloem Transport.
Encyclopedia of Plant Physiology, New Series Volume
1. Springer-Verlag, New York.
Pate, J.S., P.J. Sharkey, and C.A. Atkins.
1977.
Nutrition of a developing legume fruit. Plant
Physiol.
59:506-510.
Pate, J.S., M.B. Peoples, A.J.E. van Bel, J. Kuo, and
C.A. Atkins.
1985. Diurnal water balance of the
cowpea fruit. Plant Physiol. 77:148-156.
Peoples, M.B., J.S. Pate, C.A. Atkins, and D.R. Murray.
1985. Economy of water, carbon, and nitrogen in the
developing Cowpea fruit. Plant Physiol. 77:142147.
Peynaud, E. and P. Ribereau-Gayon.
1971. The Grape, p.
171-205.
In A.C. Hulme (ed.). The Biochemistry of
Fruits and Their Products. Vol. 2. Academic Press,
New York, New York.
Poovaiah, B.W.
1979. Role of calcium in ripening and
senescence. Comm. Soil Sci. Plant Anal.
10:83-88.
87
Poovaiah, B.W., G.M. Glenn, and A.S.N. Reddy.
1988.
Calcium and fruit softening: physiology and
biochemistry. p.107-152. In J. Janick (ed.).
Horticultural Reviews Vol. 10. Timber Press,
Portland, Oregon.
Possner, D.R.E. and W.M. Kliewer.
1985. The
localisation of acids, sugars, potassium and calcium
in developing grape berries. Vitis. 24:229-240.
Pratt. C.
1971. Reproductive anatomy in cultivated
grapes - a review. Amer. J. Enol. Vitic. 22:92109.
Radler, F.
1965a. Reduction of loss of moisture by the
cuticle wax components of grapes. Nature.
207:1002- 1003.
Radler, F.
1965b. The surface waxes of the Sultana vine
(Vitis vinifera cv. Thompson Seedless). Aust. J.
Biol. Sci.
18:1045-1056.
Rosenquist, J.K. and J.C. Morrison.
1988. The
development of the cuticle and epicuticular wax of
the grape berry. Vitis.
27:63-70.
Sacher, J.A.
1973. Senescence and postharvest
physiology. Ann. Rev. Plant Physiol.
24:197-224.
Shimomura, K.
1967. Effects of soil moisture on the
growth and nutrient absorption of grapes. Acta
Agron. Academ. Scien. Hung. Tomus.
16:209-216.
Smith, J.A.C. and J.A. Milburn.
1980. Osmoregulation
and the control of phloem-sap composition in Ricinus
communis L. Planta.
148:28-34.
Tachibana, S.
1991.
Import of calcium by tomato fruit
in relation to the day-night periodicity. Sci.
Hort. 45:235-243.
Theiler, R. and B.G. Coombe.
1985.
Influence of berry
growth and growth regulators on the development of
grape peduncles in Vitis vinifera L. Vitis.
24:111.
Tromp, J. and J. Oele.
1972. Shoot growth and mineral
composition of leaves and fruit of Apple as affected
by relative air humidity. Physiol. Plant.
27:253258.
88
Tukey, L.D.
1964. A linear electronic device for
continuous measurement and recording of fruit
enlargement and contraction. Proc. Amer. Soc. Hort.
Sci.
84:653-660.
Ureta, F., J.N. Boidron, and J. Bouard.
1981.
Influence
of dessechement de la rafle on grape quality. Amer.
J. Enol. Vitic.
32(2):90-92.
Van de Geijn, S.C. and F. Smeulders.
1981. Diurnal
changes in the flux of calcium toward meristems and
transpiring leaves in tomato and maize plants.
Planta.
151:265-271.
van Die, J. and N.C.M. Willemse.
1980. The supply of
water and solutes by phloem and xylem to growing
fruits of Yucca flaccida Haw. Ber. Dtsch. Bot. Ges.
93:327-337.
van Zyl, J.L.
1987.
Diurnal variation in grapevine
water stress as a function of changing soil water
status and meteorological conditions. S. Afr. J.
Enol. Vitic.
8:45-52.
Wiersum, L.K.
1966. Calcium content of fruits and
storage tissues in relation to the mode of water
supply. Acta Bot. Neerl.
15:406-418.
Westwood, M.N.
1988. Temperate-Zone Pomology.
Press, Portland, Oregon.
Timber
Wilkinson, B.G.
1968. Mineral composition of apples
IX.- Uptake of calcium by the fruit. J. Sci. Fd.
Agric.
19:646-647.
Winkler, A.J. and W.O. Williams.
1935. Effect of seed
development on the growth of grapes. Proc. Amer.
Soc. Hort. Sci.
33:430-434.
Winkler, A.J., J.A. Cook, W.M. Kliewer, and L.A. Lider.
1974. General Viticulture. University of
California Press, Berkeley, California.
Zee, S.Y. and T.P. O'Brien.
1970. A special type of
tracheary element associated with xylem
discontinuity in the floral axis of wheat. Aust. J.
Biol. Sci.
23:783-791.
Ziegler, H.
1963. Verwendung von 45Calcium zur Analyse
der Stoffversorgung wachsender Fruchte.
Planta.
60:41-45.
89
Zimmermann, M.H. 1983. Xylem Structure and the Ascent
of Sap. Springer-Verlag, New York.