Von Weihe & Kadir
Honors Project, 2005
Effect of Heat Stress on Photochemical Efficiency and Recovery of
‘Cynthiana’ and ‘Vignoles’ Winegrapes under Controlled
Environmental Conditions
Michael Von Weihe1 and Sorkel Kadir
1
Department of Horticulture, Forestry and Recreation Resources
Kansas State University, Manhattan, Kansas, 66506
ABSTRACT: Two wine grapes adapted to Kansas, ‘Cynthiana’ an American type
(Vitis aestivalis) and ‘Vignoles’ a French-American hybrid, were exposed to sudden and
gradual temperature increase to 40oC (104oF) under controlled environmental conditions.
Plants were exposed for 0, 3, 6, and 12 days before Photosystem II (PSII) efficiency
(Fv/Fm) and recovery period were determined in the greenhouse after 0, 3, 7, 14, and 21
days. In addition, leaf area and leaf, shoot, and root biomass were determined 21 days
after the recovery periods.
PSII efficiency, measured by Fv/Fm, of ‘Vignoles’ exposed to 40 oC was slightly
higher than that of ‘Cynthiana’. However, significant difference was observed under
gradual exposure to 40oC. Regardless of the treatment, the photosynthetic apparatus of
‘Cynthiana’ was significantly damaged under high temperature, as plants did not recover
under normal environmental conditions. ‘Vignoles’ PSII efficiency recovered to over
100% of the control in both the gradual and sudden exposure to 40ºC after 7 days of
recovery for 3, 6, and 12-days of exposure to 40ºC, except for the 12-day exposure in the
sudden experiment where ‘Vignoles’ recovered after 14 days. Under the two exposure
regimes, leaf area was greater and leaf, shoot, and root biomass were higher in ‘Vignoles’
than that of ‘Cynthiana’. Results of this study indicated that photochemical reaction of
‘Vignoles’ was more efficient and can recover from gradual or sudden exposure to high
temperature than ‘Cynthiana’. Therefore, ‘Vignoles’ is more heat resistant to high
temperature than ‘Cynthiana’.
INTRODUCTION
In the Midwest, the viticulture industry is rapidly growing. Farmers are looking
for alternative crops to diversify their operations and produce value-added farm products.
As a result, grape growing/wine making is fast becoming one of the best and most
appealing options. As the number of Midwest grape growers and wine makers increases,
improved knowledge of the best grape cultivars that grow well in the Midwest is also
increasing. In the Midwestern states of Kansas and Missouri, high summer temperature
is one of the factors that limits grape production and reduces wine quality. Temperatures
above 37ºC (99ºF) are not unusual in a typical Kansas summer.
High temperatures can disrupt many biological plant processes, in particular
photosynthesis. Many studies have shown that exposure to high temperatures can disrupt
the photosynthetic apparatus, thus decreasing the amount of chemical energy a plant can
generate. In one study that dealt with heat stress in corn (Zea mays L.), the
photosynthesis of leaves grown at optimal temperature (25°C) decreased considerably
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Von Weihe & Kadir
Honors Project, 2005
when exposed to temperatures of 35°C or higher (Sinsawat et al., 2004). Also in a study
of photosynthesis in wheat plants, high temperature greatly reduced maximum electron
transport activity of thylakoids in seedlings and as a result decreased overall
photosynthetic output (Al-Khatib and Paulsen, 1989). To be more specific about how
heat stress affects photosynthesis, photosystem II (PSII) is the process that is most
susceptible to high temperature (Al-Khatib and Paulsen, 1989). It appears that the
process by which heat stress affects PSII involves denaturation of certain functional
proteins (Thompson et al., 1989) and dissociation of the peripheral light-harvesting
pigments as well as the PSII complex (Schreiber and Berry, 1977).
It should also be mentioned that plants have the ability to adapt themselves to a
high temperature (e.g. 40°C) if exposed to a slight increase in temperature first (e.g.
between 30°C and 35°C). In a study performed on potato (Solanum tuberosum), it was
shown that in vivo measurements of chlorophyll fluorescence for plants that were
exposed to a temperature of 35°C before being exposed to 40°C could conduct
photosynthesis much more efficiently than control plants that were grown at 25°C and
suddenly placed in 40°C temperatures (Havaux, 1993). This acclimation of plants to
higher temperatures can in part be attributed to the accumulation of xanthophyll
zeaxanthin in the leaves (Havaux and Tardy, 1996). These carotenoids help to stabilize
the lipid phase of the thylakoid membrane (Havaux, 1998).
A measurement of chlorophyll fluorescence was the means by which
photosynthetic capacity of grape cultivars was evaluated in this experiment. When a
photon is absorbed by a chlorophyll molecule and excites an electron, the molecule has
three possible ways to spend this energy. The energy can be converted to heat, used in
photochemistry, or can be re-emitted as a photon of lower wavelength in a process called
fluorescence (Maxwell and Johnson, 2000). When chlorophyll fluorescence increases,
the efficiency of photochemistry decreases. However, a decrease in chlorophyll
fluorescence can result in either an increase in photochemistry, termed photochemical
quenching, or an increase in energy converted to heat, termed non-photochemical
quenching (Maxwell and Johnson, 2000). It is because of this relationship that
fluorescence was used as a means by which to measure the efficiency of photosynthesis,
particularly PSII in this experiment.
Four different values were calculated when measuring chlorophyll fluorescence
for this experiment; Fo, Fm, Fv, and Fv/Fm. Fo is the initial chlorophyll fluorescence level.
Fm represents the maximum fluorescence yield. Fv, or variable fluorescence level, is
calculated by simply subtracting Fo from Fm. From the previous values we can calculate
Fv/Fm, the PSII efficiency. It is this value that was used to evaluate the performance of
each grape cultivars exposed to high temperature.
Identifying cultivars that withstand extreme summer temperatures is critical to
expanding the viticulture industry in the Kansas. Two grape cultivars were selected for
this experiment that are already grown in the Midwest because of their winter hardiness
and ability to produce quality wine. However, little information was available about their
responses to high temperatures. It was the aim of this experiment to 1) study the
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Von Weihe & Kadir
Honors Project, 2005
influence of sudden and gradual heat shock on the efficiency of photosynthesis as well as
growth and development of the vines, 2) determine time required for the vines to recover
from heat treatments, and 3) identify the heat resistant cultivar.
MATERIALS AND METHODS
Dormant grape cuttings of ‘Cynthiana’ (Vitis aestivalis) and ‘Vignoles’ were
planted in May 2004 using black, one gallon polyethylene pots and a media comprised of
sand, peat, and soil in a 1:1:1 ratio. The grapes were grown in a greenhouse at a
temperature of 23-25° C and watered with plain water as needed. Plants were also
fertilized at least once a week with Scotts Peters Professional Peat-Lite Special 20-10-20
at a rate of 200 ppm N. The grapes were grown until roots, shoots, and leaves were
developed in September. The plants were then divided into two groups of 18 (9
Cynthiana and 9 Vignoles) for two different experiments; a sudden exposure to high
temperature (40/35ºC) versus a gradual exposure to high temperature (described
subsequently). These two environments were simulated using two growth chambers
(Control Environments LTD Conviron CMP 3244 growth chambers, Winnipeg,
Manitoba Canada). In addition, three plants of each cultivar were kept in the greenhouse
as the control.
In the sudden exposure experiment, 18 plants (9 ‘Cynthiana’ and 9 ‘Vignoles’)
were immediately placed in a growth chamber with a temperature set to 40/35°C ± 1
day/night. Photoperiod of the chamber was set to a 16 hr/8 hr day/night cycle with the
light intensity of the day being 450 µmol m-²·s-¹. In the gradual exposure experiment, the
18 plants were placed in a growth chamber with a day temperature set to 32° C and
scheduled to increase to 34° C, 36° C and finally 40° C after three days at each
temperature. Once at 40° C, both experiments were carried out in the same manner.
For both the sudden and gradual experiments, three grape plants of each cultivar
were removed from the growth chambers and placed in the greenhouse for a recovery
period after 3, 6, and 12 days in the growth chambers. Chlorophyll fluorescence was
measured during the recovery period at 0, 3, 7, 14, and 21 days in the greenhouse using a
Hansatech Instruments LTD Fluorescence Monitoring System, Kings Lynn, England.
The values measured by the machine were Fo, Fm, Fv, and Fv/Fm. Leaves tagged and
selected for measurement were of consistent size and placement on each grape plant at
the beginning of the experiment.
After 21 days of recovery in the greenhouse, leaves were harvested from the
plants and total leaf area (LA) was measured using a LI-1905 sensor (Li-Cor, Inc,
Lincoln, Nebraska, USA). In addition, leaves and shoot growth from the current year
were put into brown paper bags and placed in a Thelco oven set at 70ºC for 3 days to dry.
Roots were washed the day of harvest, allowed to dry in the greenhouse for one day then
put into a brown paper bag and placed in the oven for 3 days to dry. Leaf, shoot, and root
dry weight (LDW, SDW, and RDW, respectively) were taken at the end of the drying
period using a Mettler P1200 scale.
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Von Weihe & Kadir
Honors Project, 2005
RESULTS AND DISCUSSION
Objective 1: Influence of Sudden and Gradual Heat Shock on Efficiency of
Photosynthesis as well as Growth and Development of Vines
When analyzing ‘Cynthiana’ and comparing photosystem II (PSII) efficiency
(Fv/Fm) between the sudden and gradual experiments, the sudden exposure to 40ºC
showed higher Fv/Fm after 21 days of recovery than the gradual exposure (Fig. 1 and 2).
When analyzing ‘Vignoles’ and comparing PSII efficiency between the sudden and
gradual experiments, the gradual exposure showed higher Fv/Fm after 21 days than the
sudden experiment showed (Fig. 1 and 2).
Figure 1. Efficiency of photosystem II (PSII) (Fv/Fm) values in percent of control for
‘Cynthiana’ and ‘Vignoles’ exposed to a sudden increase in temperature to 40ºC for 3, 6,
and 12 day exposure periods. Efficiency of PSII was measured after 3, 7, 14, and 21 days
of recovery in the greenhouse.
Fv/Fm (% control)
140
120
100
80
60
40
Cynthiana
Vignoles
LSD0.05=21
20
0
3
7
14
Recovery Period (days)
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Honors Project, 2005
Fv/Fm (% control)
140
120
100
80
60
40
20
0
3
7
14
21
Recovery Period (days)
140
Fv/Fm (% control)
120
100
80
60
40
20
0
3
7
14
Recovery Period (days)
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Honors Project, 2005
Figure 2. Efficiency of PSII (Fv/Fm) in percent of control for ‘Cynthiana’ and ‘Vignoles’
exposed to a gradual increase in temperature to 40ºC for 3, 6, and 12 day exposure
periods. Efficiency of PSII was measured after 3, 7, 14, and 21 days of recovery in the
greenhouse.
140
100
80
60
40
Cynthiana
Vignoles
LSD0.05=20
20
0
3
7
14
21
Recovery Period (days)
140
Fv/Fm (% control)
Fv/Fm (% control)
120
120
100
80
60
40
20
0
3
7
14
Recovery Period (days)
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Von Weihe & Kadir
Honors Project, 2005
Fv/Fm (% control)
140
120
100
80
60
40
20
0
3
7
14
21
Recovery Period (days)
‘Cynthiana’ had less leaf area (LA) at 3, 6, and 12 days of gradual exposure to
40ºC in contrast to higher LA in the sudden exposure to 40ºC (Table 1). This reinforces
the fact that ‘Cynthiana’ responded better to a sudden exposure to heat stress than a
gradual exposure to 40ºC. Another conclusion that can be drawn is that ‘Cynthiana’ had
significantly more LA in the 3 and 6-day sudden exposures to heat stress when compared
to a gradual exposure to heat stress. Leaf dry weight (LDW) is similar to the LA values
in that the sudden exposure had significantly more LDW in the 3 and 6-day sudden
exposures to heat stress than the gradual experiment (Table 2). Shoot dry weight (SDW)
was inconclusive to show a significant difference between the sudden and gradual
exposure to 40ºC (Table 3). However, root dry weight (RDW) showed the opposite
effect, the gradual exposure to 40ºC had much higher RDW than the sudden exposure to
40ºC (Table 4). This demonstrates that when ‘Cynthiana’ is exposed to heat stress it will
divert growth capacity to the roots instead of to leaves and shoots, possibly in an attempt
to find more water.
Table 1. Total Leaf area (LA) in percent of control for ‘Cynthiana’ and ‘Vignoles’
exposed 3, 6, and 12 days to sudden and gradual temperature increases up to 40/35ºC.
LA was measured after a 21-day recovery period.
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Honors Project, 2005
Cultivar
Cynthiana
Vignoles
3
108.2545
121.2881
Cynthiana
Vignoles
LSD0.05
3
34.22367
129.6449
22
Sudden exposure (days)
6
126.6952
106.7127
Gradual exposure (days)
6
10.70316
146.6329
22
12
60.11813
106.881
12
56.3843
74.71667
22
Table 2. Leaf dry weight (LDW) in percent of control for ‘Cynthiana’ and ‘Vignoles’
exposed 3, 6, and 12 days to sudden and gradual temperature increases up to 40/35ºC.
LDW was measured after a 21-day recovery period.
Cultivar
Cynthiana
Vignoles
3
76.56523
104.438
Cynthiana
Vignoles
LSD0.05
3
38.13782
102.6424
21
Sudden exposure (days)
6
91.69408
84.14079
Gradual exposure (days)
6
17.25031
127.7933
21
12
53.90476
78.59649
12
38.31807
55.61878
21
Table 3. Shoot dry weight (SDW) in percent of control for ‘Cynthiana’ and ‘Vignoles’
exposed 3, 6, and 12 days to sudden and gradual temperature increases up to 40/35ºC.
SDW was measured after a 21-day recovery period.
Cultivar
Cynthiana
Vignoles
3
81.01793
92.20611
Cynthiana
Vignoles
LSD0.05
3
135.879
68.01558
17
Sudden exposure (days)
6
103.0135
58.33182
Gradual exposure (days)
6
101.7102
117.8521
17
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12
113.142
47.62184
12
100.1803
59.2157
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Von Weihe & Kadir
Honors Project, 2005
Table 4. Root dry weight (RDW) in percent of control for ‘Cynthiana’ and ‘Vignoles’
exposed 3, 6, and 12 days to sudden and gradual temperature increases up to 40/35ºC.
RDW was measured after a 21-day recovery period.
Cultivar
Cynthiana
Vignoles
3
73.08603
92.36031
Cynthiana
Vignoles
LSD0.05
3
205.7659
108.04
18
Sudden exposure (days)
6
106.4427
61.48434
Gradual exposure (days)
6
148.2145
93.09715
18
12
83.77405
67.17622
12
165.2623
61.62148
18
‘Vignoles’ had considerably more LA than ‘Cynthiana’ (Table 1) for the 12-day
sudden exposure to 40ºC and for the 3 and 6-day gradual exposure. Additionally,
‘Vignoles’ had higher LDW (Table 2) than ‘Cynthiana’ for the 3 and 12-day sudden
exposure to 40ºC and for the 3 and 6-day gradual exposure. Interestingly, ‘Vignoles’ had
less SDW (Table 3) and RDW (Table 4) than ‘Cynthiana’ in SDW and RDW. This
indicates that internodes were shorter and growth was diverted to leaves in ‘Vignoles’
than in ‘Cynthiana’. It has been shown before that heat stress affects meristem tip growth
in ‘Cynthiana’, but meristem tips on ‘Vignoles’ are more heat resistant and thus can
continue to develop more leaves (Personal communication with Kadir, S.)
Objective 2: Time Required for Vines to Recover from Heat Treatments
‘Cynthiana’, irrespective of gradual or sudden exposure, never fully recovered the
efficiency of PSII compared to control plants at the end of a 21-day recovery period (Fig.
1 and 2). When suddenly exposed to high temperature for 3 days, ‘Cynthiana’ PSII
efficiency was over 100% after 7 and 14 days of recovery but dropped below 100% after
21 days of recovery (Fig. 1). This sudden decrease in PSII efficiency could likely be due
to factors other that were not investigated in this study.
‘Vignoles’ PSII efficiency recovered to over 100% of the control in both the
gradual and sudden exposure to 40ºC after 7 days of recovery for 3, 6, and 12-day
exposure to 40ºC except for the 12-day exposure in the sudden experiment where
‘Vignoles’ recovered after 14 days of recovery. ‘Vignoles’ had a higher Fv/Fm when
exposed to the gradual temperature exposure compared to the sudden temperature
exposure. The ability of ‘Vignoles’ to adapt itself to gradual temperature increases could
be a result of carotenoids stabilizing the lipid phase of the thylakoid membrane, as stated
earlier in the example of potatoes exposed to heat stress (Havaux, 1998).
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Von Weihe & Kadir
Honors Project, 2005
Objective 3: Identify the Heat Resistant Cultivar
Data gathered from this study points to the fact that ‘Vignoles’ is more heat
resistant than ‘Cynthiana’. ‘Vignoles’ is able to conduct more efficient photosynthesis
than ‘Cynthiana’ for 3, 6, and 12-day exposure to sudden increase in temperature to 40ºC
(Fig. 1). In addition, ‘Vignoles’ conducts significantly more efficient photosynthesis in
the gradual experiment (Fig, 2). Also ‘Vignoles’ produced higher LA and LDW but
lower amounts of SDW and RDW compared to ‘Cynthiana’. This information illustrates
that ‘Vignoles’ under heat stress will form more leaves where as ‘Cynthiana’ will
produce more roots.
REFERENCES
Al-Khatib, K., Paulsen, G. M., 1989. Enhancement of thermal injury to photosynthesis
in wheat plants and thylakoids by high light intensity. Plant Physiology. 90: 1041-1048.
Havaux, M., 1993. Rapid photosynthetic adaptation to heat stress triggered in potato
leaves by moderately elevated temperatures. Plant Cell Environment. 16: 461-467.
Havaux, M., 1998. Carotenoids as membrane stabilizers in chloroplasts. Trends in Plant
Science. 3: 147-151.
Havaux, M., Tardy, F., 1996. Temperature-dependent adjustment of the thermal stability
of photosystem II in vivo: possible involvement of xanthophylls-cycle pigments. Planta.
198: 324-333.
Maxwell, K., Johnson, G. N., 2000. Chlorophyll fluorescence – A practical guide.
Journal of Experimental Botany. 51: 659-668
Schreiber U., Berry J. A., 1977. Heat-induced changes of chlorophyll fluorescence in
intact leaves correlated with damage of the photosynthetic apparatus. Planta. 136: 233238.
Sinsawat, V., Leipner, J., Stamp, P., Fracheboud, Y., 2004. Effect of heat stress on the
photosynthetic apparatus in maize (Zea mays L.) grown at control or high temperature.
Environmental and Experimental Botany. 52: 123-129.
Thompson L. K., Blaylock R., Sturtevant J. M., Brudvig G. W., 1989. Molecular basis of
heat denaturation of photosystem II. Biochemistry. 28: 6686-6695.
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