Introduction
Tea-oil
tree (Camellia oleifera Abel; family Theaceae) is an
evergreen shrub and oil plant that has
been cultivated for more
than 2,000
years in China (Tan et al. 2011b). C. oleifera is
one of the four major oil plants in
the world (Li et al. 2016); the others are the olive tree (Olea europaea), oil palm (Elaeis guineensis), and coconut palm (Cocos nucifera). The tea oil extracted from C. oleifera seeds is
edible oil called “eastern olive oil,” because of its high
nutritional value and health care functions (Qu et al. 2019). This oil has a similar
chemical composition
as that of olive oil, as both contain high amounts of unsaturated
fatty acids (Gao et al.
2015; Yang et al. 2016).
Tea oil is not only edible but also a traditional Chinese medicine and superior
nutritional dietary supplement that benefits human digestive system, reduces
blood cholesterol and prevents hypertension and hardening of the arteries (Feás et
al. 2013; Zeng et al. 2015). It
is also an important raw material for the pharmaceutical and chemical
industries. For example, tea meal can be used to extract saponin and produce
feed, and the tea shell can be used to produce potassium carbonate or cultivate
edible and medicinal fungi (Zhang and Liu 2007; Hu et al. 2012; Zhu et al.
2018). In recent years C. oleifera has been widely planted in the
red soil at hilly regions of southern China due to the rapid development of the
C. oleifera industry.
C. oleifera is
a self-incompatible plant at the beginning of flowering in early November and
usually blooms in large numbers in mid-late November (Fig. 1c). In addition,
most C. oleifera is polyploid with many cultivars that display significant
differences in cold resistance (Deng et
al. 2018; Shi et al. 2019).
Cultivars of C. oleifera
with big fruits, high yields, good stability, and strong resistance have been
planted on a large scale by farmers. Hua Shuo and Hua Xin are new high-yielding
C.
oleifera cultivars bred from common C. oleifera in 2009. Hua Shuo (Fig. 1a)
has large fruits, high yields, strong resistance, and late maturity (Tan et al. 2011a). Hua Xin (Fig. 1b) has
high and stable yields, strong resistance, and precocity (Tan et al. 2012). The cold resistance of the
two cultivars is unclear, particularly when the plants are flowering in large
numbers.
The
primary problem during C. oleifera production is low fruit setting
rate caused by low temperature and rainy weather in southern China, resulting
in a lower yield and reduction in the distribution area (Peng and Chen 2008; Wang et al. 2017). Chen (2018) showed that sufficient sunshine and
suitable temperatures improve the seed setting rate of C. oleifera. In addition, C. oleifera plants
flowering in the winter will encounter freeze injury, and low insects
activities, resulting in abnormal pollination and fertilization of C. oleifera,
which seriously affect the development of C.
oleifera industry in China (Fig. 1).
The lowest temperature for suitable growth of the two C. oleifera cultivars is unknown, particularly in
southern China, where rainy and low temperature days are frequent. Little
information is available about the effects of prolonged low temperature on the
growth of the two C. oleifera cultivars during the flowering
phase.
We
investigated the differences in physiological indices between the two cultivars
during the flowering period. We compared cold resistance by exploring the
long-term low-temperature stress on physiological and biochemical processes of
the two C. oleifera cultivars. For this purpose, we measured chlorophyll content,
photosynthesis, chlorophyll fluorescence, and observed the leaf anatomical
structure and chloroplast ultrastructure of the two C. oleifera cultivars.
Materials and Methods
Plant materials and treatments
The
experimental materials for this study were obtained from 4-year-old C.
oleifera potted plants cultured by grafting a shoot each of C. oleifera
Hua Shuo and Hua Xin onto germinated hypocotyls of seeds on the same tree as
rootstock. On February 18, 2017, 120 two year-old C. oleifera young plants were selected and transplanted into plastic
containers (22 × 22 × 20 cm) filled with a 2:1:1 mixture of peat soil, loess,
and perlite. The plants were grown under natural conditions with the same water
and fertilizer management at the Life Science Building of Central South
University of Forestry and Technology, Changsha, China (28°10’ N; 113°23’ E).
On November 3, 2018, 54 plants of each cultivar with similar growth rates were divided randomly
into three groups. Each group consisted of 18 plants. Four year-old C.
oleifera potted plants were placed in three different temperature for the experiments:
(1) The C. oleifera potted plants were placed in field conditions (CK); (2)
low temperature of 6°C in an artificial climate chamber (6°C); (3) normal
temperature of 25°C in an artificial climate chamber (25°C). Other parameters
in each room of the artificial climate chamber were the same with 70% relative
humidity, a 12 h photoperiod at a photosynthetic photon flux density of 200
μmol·m-2·s-1, and an average CO2 concentration
of 450 μmol∙mol-1.
After 25 days, florescence, chlorophyll
content, photosynthesis, physiological response and anatomical structure were
measured in different treatments. Immediately after measuring gas exchange, the
leaves were cut, weighed, wrapped in tin foil, frozen in liquid nitrogen, and
stored at –80°C until the physiological response measurements were taken.
Plants in all treatments were watered (500 mL/plant) twice and fertilized once
per week with 500 mL Hoagland solution during the experimental period (Li et al. 2017).
Chlorophyll content analyses
Six plants of each of the two cultivars in each treatment
were used for the test. Chlorophyll content was measured with 10 mL an
acetone-ethanol solution (1:1, v/v) (Zhang 1986). The samples were soaked in
the solution for 24 h at 4°C in the dark. The absorbance values at 663 nm (OD663)
and 645 nm (OD645) of the solution were measured with a
spectrophotometer (UV-1100 MAPADA, Shanghai, China). Chlorophyll content was
calculated with the following equations:
Chl a (mg/dm2) = 12.72 × OD663 –
2.59 × OD645
Chl b (mg/dm2) = 22.88 × OD645 –
4.68 × OD663
Chl
(a + b) (mg/dm2) = Chl a + Chl b
Photosynthetic characteristic
measurements
The photosynthetic
characteristics were
measured using an Li-6400xt instrument (LI-COR
Biosciences, Lincoln, NE, USA). Six plants of each of the two cultivars from
the treatments and control were used for the measurements. The
photosynthetic parameters were measured between 9:00 am and 11:00 am with
red–blue light of 1,000 μmol·m-2·s-1 and a CO2
concentration of 400 μmol·mol-1.
Chlorophyll
fluorescence analyses
The chlorophyll fluorescence parameters were
measured using the Li-6400xt device. Six plants of each
of the two cultivars from the treatments and control were
used for the measurements. After a 2 h dark adaptation from
20:00–22:00, the plants were given a saturation pulse for 0.8 s at a light
intensity of 7,200 μmol·m-2·s-1 in the dark. Then
the Li-6400xt collected the initial fluorescence (Fo) and maximal photochemical efficiency (Fv/Fm) data. The
actual photochemical quantum efficiency (ΦPSII)
and electron transport rate (ETR) were determined after activation with light.
Anatomical leaf feature
analyses
Six
plants of each of the two cultivars from the treatment and control were used
for the measurements. The leaf anatomical structure was studied in paraffin
sections using an optical microscope. Mature leaf samples
from the plants
were cut into 5
× 4 mm pieces which were then soaked in FAA fixative solution containing
70% ethanol, glacial acetic acid, and formaldehyde (95:5:5, v/v/v) for 24 h.
The samples were dehydrated in a graded ethanol series (70, 80, 90, 95, and
100%), embedded in paraffin, microtome sliced (Leica RM2235, Germany), and
stained using a Safranin-O and acid fast green staining procedure (Zeng et al. 2008). Using the Leica DMi8
inverted microscope (Leica Inc. Jena, Germany) to observe the images, the
structure of the palisade and spongy tissues was analyzed with application
software (version 4.12.0).
Determination of malondialdehyde (MDA) and soluble
sugar contents
Fresh
leaves (0.3 g) were collected at a similar position to determine MDA and
soluble sugar contents. Six plants of each of the two cultivars from the
treatments and control were used to provide the leaf tissues. All samples were
wrapped in sterilized tin foil (Solarbio) and stored at –80°C for later
analyses. MDA content was measured as described by He et al. (2015), and
soluble sugar content was determined as described by
Wang et al. (2015) and Irigoyen et al. (1992). Each determination
included three biological and technical replicates.
Chloroplast ultrastructural observations
The
middle portion of each leaf was cut into 1 mm2 strips and fixed in
2.5% glutaraldehyde solution (prepared with 0.1 mol L-1 sodium
phosphate buffer, pH 7.3) for 24 h at 4°C. After washing three times (30 min
each), the leaf samples were dehydrated in a series of graded ethanol
solutions. After fixing in 1% osmium tetroxide for 2 h at room temperature, the
strips were embedded in epoxy resin and placed in an ion sputtering coating
machine for 20 min. The blade samples were sliced (0.5 µm) using an
ultramicrotome (Leica EM UC7; Heidelberg, Germany) and mounted on copper grids.
Transmission electron microscopy (HT7700; Hitachi, Tokyo, Japan) was used for
the observations.
Statistical analyses
Microsoft Office Excel 2013
was used to process the data. Experiments were conducted as a completely randomized
design (CRD) with eighteen replications each treatment. SPSS 19.0 software was used to analyse the variance
to test for differences. Treatment means were compared using one-way analysis
of variance (ANOVA) and Duncan’s multiple range test with a probability of P≤ 0.05.
Results
Investigation of flowering and fruiting
Compared
to the natural temperature (CK), both Hua Shuo and Hua Xin at 25°C flowered
ahead of schedule, and the first flowering dates were advanced by 4 and 2 days,
respectively. The flowering phases of Hua Shuo and Hua Xin were shortened by 20
and 11 days, respectively compared to CK at 25°C. However, the first flowering
dates of Hua Shuo and Hua Xin were delayed by 4 and 5 days at 6°C, respectively
compared to CK (Table 1).
Under
natural conditions, the petals fell off after flowering of Hua Shuo and Hua
Xin, and the young fruits were very small (Fig. 2aii, bii). The young fruits of
Hua Xin were bigger than those of Hua Shuo at 25°C (Fig. 2ai, bi). Interestingly,
a large number of Hua Xin buds did not flower at 6°C, and remained in their
original state (Fig. 2biii), while Hua Shuo flowered for 49 days. The stamens
and pistils withered but did not fall off, and a significant amount of mucus
appeared at the base of the flower (Fig. 2aiii).
Chlorophyll content
The Chl a, Chl b, and total chlorophyll contents of Hua
Shuo and Hua Xin were highest at 25°C, but significantly decreased in both
cultivars at 6°C compared to the normal temperature of 25°C, except for Chl b
of Hua Shuo (Table 2). In Hua Xin, they significantly decreased by 30.33, 36.88
and 31.75%, respectively, compared to controls (CK) (P < 0.05), at 6°C, but total chlorophyll content of Hua Shuo was
not significantly different (P >
0.05) (Table 2). This indicates that low temperature decreases the chlorophyll
content of C. oleifera.
Photosynthetic characteristics
Net photosynthetic rate (Pn), transpiration rate (Tr), and stomatal conductance (Gs) were significantly affected by temperature, and were
shown to be the highest values at 25°C in both cultivars (Fig. 3a–c). The
values for Hua Shuo decreased by 40.25, 79.43 and 75.00% (P < 0.05), respectively, at 6°C compared to controls, and those
of Hua Xin decreased by 54.45, 74.23 and 80.95% (P < 0.05), respectively. However, Ci was not significantly different between the two
cultivars, except under control conditions (Fig. 3d). In addition, the Pn
of Hua Shuo was lower than that of Hua Xin at 25°C, but was higher than that of
Hua Xin at 6°C.
Chlorophyll fluorescence
Different temperature treatments had variable effects on
the chloroplast fluorescence parameters of the two cultivars. Initial
fluorescence (Fo)
significantly improved and maximum phototchemical efficiency (Fv/Fm) decreased
for both Hua Shuo and Hua Xin at 6°C compared to the controls (Fig. 4a, b). No
significant differences were observed between 25°C and CK for either strain.
The actual ΦPSII
and ETR values were higher at 25°C than those at 6°C (P > 0.05) (Fig. 4c, d). The ETR values of Hua Shuo and Hua Xin
decreased by 52.15 and 36.37% (P <
0.05), respectively, at 6°C compared to the controls (Fig. 4d).
Soluble sugar and malondialdehyde contents
Table 1: Florescence of two C. oleifera
cultivars under different
temperatures
|
Species |
Treatment |
Phenological phase |
Florescence
(days) |
||
|
First
flowering stage |
Flowering
stage |
Late
flowering stage |
|||
|
Hua
Shuo |
CK |
11/8-11/14 |
11/15-12/5 |
12/6-12/16 |
38 d |
|
25ºC |
11/4-11/8 |
11/9-11/19 |
11/20-11/22 |
18 d |
|
|
6ºC |
11/12-11/15 |
11/16-12/20 |
12/21-12/31 |
49 d |
|
|
Hua
Xin |
CK |
11/5-11/7 |
11/8-11/21 |
11/22-12/4 |
31 d |
|
25ºC |
11/3-11/4 |
11/5-11/11 |
11/12-11/23 |
20 d |
|
|
6ºC |
11/10-11/28 |
/ |
/ |
/ |
|
%20777-785_files/image002.jpg)
Fig. 1: The
growth habit of the two C. oleifera
cultivars
a) Camellia Hua
Shuo with fruit; b) Camellia
Hua Xin with fruit; c) Camellia Hua Xin with fruit and flowers
(IF: initial flowering; FB: flower bud); d) Frozen Camellia Hua Shuo; e)
Frozen Camellia Hua Xin; f: Frozen flower (FF: frozen flowers); g: Bees
stop moving at low temperatures
%20777-785_files/image004.jpg)
Fig. 2: Growth of two C. oleifera cultivars
under different temperature at 25 DAT
a1: 25°C of Hua Shuo (bar = 0.5 cm); a2: CK of Hua Shuo (bar = 0.5 cm); a3: 6°C of Hua Shuo (bar = 0.3 cm);
b1: 25°C of Hua Xin (bar = 0.5 cm); b2: CK of Hua Xin (bar = 1 cm); b3: 6°C of Hua Xin (bar = 0.5 cm)
Low-temperature
treatment increased soluble sugar and MDA contents. The soluble sugar content
at 6°C increased by 29.03 and 7.86% (P < 0.05) in Hua Shuo and Hua
Xin, respectively, compared to the controls. At 25°C it decreased by 24.72 and
24.07% (P < 0.05), respectively (Fig. 5a). MDA contents were lowest
in both cultivars at 25°C and that of Hua Xin increased by 24.29% (P <
0.05) at 6°C compared to the controls (Fig. 5b).
Chloroplast
ultrastructure
Table 2: Chlorophyll
content of
the two C. oleifera cultivars under
different temperatures
|
Species |
Treatment |
Chl
a content (mg/dm-2) |
Chl
b content (mg/dm-2) |
Total
Chl content (mg/dm-2) |
|
|
Hua Shuo |
CK |
4.56±0.06b |
1.13±0.10b |
5.69±0.16b |
|
|
25ºC |
5.92±0.08a |
1.56±0.05a |
7.48±0.13a |
||
|
6ºC |
4.71±0.06b |
1.61±0.08a |
6.32±0.14b |
||
|
Hua Xin |
CK |
5.11±0.07b |
1.41±0.07a |
6.52±0.14ab |
|
|
25ºC |
5.36±0.07a |
1.34±0.11a |
6.70±0.18a |
||
|
6ºC |
3.56±0.10c |
0.89±0.12b |
4.45±0.22c |
||
Different lowercase letters within a column indicate a
significant difference at P < 0.05 a (n = 6)
%20777-785_files/image006.png)
Fig. 3: Effects
of different temperatures on net photosynthetic rate (Pn), stomatal conductance (Gs), intercellular CO2 concentration (Ci), and transpiration rate (Tr) in the two C. oleifera cultivars. Different
lowercase letters within a column indicate a significant difference at P <
0.05 (n = 6)
%20777-785_files/image008.png)
Fig. 4: Effects
of different temperatures on initial fluorescence (Fo), maximum
phototchemical efficiency (Fv/Fm), actual
photochemical quantum efficiency (ΦPSII),
and electron transport rate (ETR) in the two C. oleifera cultivars. Different lowercase letters in the columns
indicate a significant difference at P < 0.05 (n = 6)
The
chloroplast ultrastructure of both cultivars changed at different temperatures (Fig. 6). At 25°C, both cultivars had intact cell
morphology, clear cell walls, and normal organelle structures (e.g., chloroplasts).
Clearly, all chloroplasts were in the shape of a convex lens and distributed
close to the cell edge (Fig. 6ai, bi). In addition, the grana and stroma thylakoid
structures were clear, and the cytoplasm contained a small number of randomly
distributed starch granules (Fig. 6ai, bi). However, at 6°C, the chloroplast
reticulate structures of the photosynthetic lamellae were damaged in both
cultivars, although that of Hua Xin had completely disintegrated and the
thylakoid membrane was loose (Fig. 6bii), while the grana lamellae of Hua Shuo
were loosely arranged and slightly swollen and dilated (Fig. 6bi). The
chloroplasts of Hua Shuo was intact in the field environment, while the
chloroplasts of Hua Xin were elongated into strips, deviated from the cell
membranes, and loosely arranged (Fig. 6aiii, biii).
%20777-785_files/image010.png)
Fig. 5: Effects of different temperatures on soluble
sugar and MDA contents of the two C.
oleifera cultivars. Different lowercase letters within a column indicate a
significant difference at P < 0.05 (n = 6)
%20777-785_files/image012.jpg)
Fig. 6: Effects of different temperatures on
mesophyll cell ultrastructure of the two C.
oleifera cultivars
a:
Hua Shuo; b: Hua Xin; 1: 25°C; 2: 6°C; 3: control
(CK)
Leaf
anatomical features
C.
oleifera leaves
are composed of the upper epidermis, lower epidermis, and mesophyll. The
epidermis consisted of irregular oblong monolayers of varying sized cells, and
the mesophyll consisted of a layer of palisade tissue cells and multiple layers
of spongy tissue cells. The mesophyll cells were closely arranged in palisade
tissue. There were more chloroplasts in the cells. The thicknesses of palisade
tissue were significantly increased at 6°C in both cultivars than at 25°C. The sponge thickness of Hua Xin was the greatest among all
samples. Leaf thickness significantly increased in both cultivars at 6°C
compared to the controls and the leaf thicknesses of Hua Shuo and Hua Xin
significantly increased by 19.49 and 13.72%, respectively (Table 3). At 6°C,
the palisade tissue cells of both cultivars were elongated and the leaves were
significantly thicker. The outer cuticle of the upper epidermis cells of Hua
Shuo leaves was obviously thicker, while the spongy tissue cells was loosely
arranged and the intercellular space was enlarged (Fig. 7). The results
indicated that low temperature (6°C)
increased leaf thickness of C.
oleifera, which would help protect the plants from the impact of chilling
stress.
Discussion
Temperature plays an important role in the flowering and
fruiting of plants. Some environmental signals, particularly warm temperatures,
promote flowering by activating FT transcription; however, exposure to high
temperatures reduces the activity of floral repressors (Fernández et al. 2016). In this study, temperature
had a significant effect on the flowering stage of two C. oleifera cultivars.
The normal temperature of 25°C during the flowering phase promoted early
flowering, whereas 6°C prolonged florescence. Similarly, Daba et al. (2016) reported earlier flowering
under long days and higher temperatures than under short days and lower
temperatures. Table 3: The anatomical features of leaves of the two
C. oleifera cultivars under
different temperatures
|
Species |
Treatment |
Palisade tissue thickness/µm |
Sponge tissue thickness/µm |
Leaf thickness/µm |
|
Hua Shuo |
25ºC |
151.38 ± 7.82 b |
254.48 ± 8.14 a |
471.12 ± 6.72 b |
|
6ºC |
224.67 ± 10.96 a |
267.11 ± 10.97 a |
544.47 ± 10.00 a |
|
|
CK |
148.08 ± 9.51 b |
261.19 ± 7.77 a |
455.66 ± 4.09 c |
|
|
Hua Xin |
25ºC |
155.90 ± 9.83 b |
234.56 ± 7.50 b |
454.97 ± 16.26 b |
|
6ºC |
209.23 ± 18.28 a |
228.99 ± 8.05 b |
500.04 ± 19.45 a |
|
|
CK |
107.75 ± 6.43 c |
276.34 ± 7.58 a |
439.72 ± 5.51 b |
Different
lowercase letters within a column indicate a significant difference at P <
0.05 according to Duncan’s tests (n = 9)
%20777-785_files/image014.png)
Fig. 7: Effects of different temperatures on leaf
anatomical structure of the two C.
oleifera cultivars (40×)
a: Hua Shuo; b: Hua Xin; 1:
25°C; 2: 6°C; 3: control (CK)
A
large number of Hua Xin flower buds did not blossom at 6°C, while Hua Shuo
flowered, the stamens and pistils withered but did not fall off, and a
significant amount of mucus appeared at the base of the flower. The main reason
may be that Hua Shuo secretes mucus to protect the young fruit against freezing
injury, which is a self-protective mechanism in plants to adapt to a new
environment. The phenotypic differences between the two cultivars were due to
flowering response to low temperature, and that the differences were related to
the expression of cold-resistance genes (Catt and Paul 2017).
Chlorophyll captures light energy in green
leaves during photosynthesis, which is a series of enzymatic reactions. Studies
have shown that low or high temperature stress can change the characteristics
of the chloroplast membrane, leading to destruction of the chloroplast and a
decrease in enzyme activities, which hinder chlorophyll synthesis and
accelerate chlorophyll decomposition (Kowitcharoen et al. 2015; Jespersen et al. 2016; Li et
al. 2018). In this experiment, the chlorophyll contents of the two C. oleifera
cultivars showed a significant downward trend under the low-temperature stress,
indicating that formation of chloroplasts and the synthetic rate of chlorophyll
were significantly decreased by the low temperature (Cai et al. 2019). The contents of Chl-a, Chl-b, and total chlorophyll
of Hua Xin were much lower than those of Hua Shuo at 6°C. The leaves of Hua Xin
gradually turned yellow, indicating that Hua Shuo was more cold-resistant than
Hua Xin.
Photosynthesis
requires a balance between the light energy absorbed by the light harvesting system
and the energy consumed by the plant; therefore, it is very sensitive to any
change in the environmental conditions. Low temperature exacerbates the
imbalance between the energy source and the metabolic sink, causing
photosynthesis to significantly change (Ensminger et al. 2006). The main factors causing a decline in the
photosynthetic rate of plant leaves can be classified into stomatal and
non-stomatal limitations caused by the decrease in photosynthetic activity in
mesophyll cells under external environmental stress (Gu et al. 2019). Farquhar and Sharkey (1982) showed that a decrease
in Pn is mainly caused by
stomatal constraints when both gs
and Ci decrease, while a
decrease in gs is
accompanied by an increase in Ci,
and a decrease in Pn is
mainly caused by non-stomatal factors. In this study, the increase in Ci lead us to hypothesize that the drop in Pn was mainly due to non-stomatal factors,
such as damage to chloroplasts or reduced photosynthetic enzyme activities. However, the simultaneous decline of Pn, gs, and Ci under natural conditions in the two C. oleifera cultivars clearly indicated that stomatal closure was
the main factor responsible for the reduced photosynthetic assimilation rate.
This result was consistent with the chloroplast ultrastructural observations.
Therefore, low temperature had a serious effect on photosynthetic physiology
and carbon assimilation in C. oleifera. In conclusion, under natural conditions, C. oleifera first reduced the number of CO2 photosynthetic
reaction sites entering mesophyll cells by closing a portion of the stomata or
adjusting the stomatal opening, and then photoinhibition occurred to protect
the photosynthetic organs from low temperature damage (Lu et al. 2015; Xu et al.
2019).
Analysis of chlorophyll fluorescence
parameters is helpful to elucidate the location and extent of photosynthetic
apparatus injured by stress (Kooten and Snel 1990). Chlorophyll fluorescence
parameters play a unique role in the study of light absorption and transmission
(Wei et al. 2011; Geng et al. 2014). The scientific community
has reached a consensus that the Fv/Fm
of most plants is between 0.8 and 0.85 under healthy physiological conditions.
An Fv/Fm value
< 0.75 indicates that the plants are under stress (Perks et al. 2004). PSII electron transfer is carried out after the
photochemical reaction, which leads to splitting (oxidation) of water
molecules. Therefore, ETR is valuable for many types of plant stress
investigations. Our results show
that the low
temperature (6°C) significantly decreased Fv/Fm, ФPSII, and ETR in the two C. oleifera cultivars, indicating that PSII
photochemical efficiency of leaves decreased under low-temperature stress,
which could be the result of photochemical damage in the PSII reaction center
or photoprotection (Demmig and Björkman 1987; Hao et al. 2019). In this study, F0 increased
while total
chlorophyll content
decreased under the low temperature. This is a clear indication that the number
of inactive PSII reactive centers decreased PSII activity due to stress (Li et al. 2017).
Soluble sugars are an osmotic regulator used
for plant cold resistance. Studies have shown that soluble sugar content is
positively correlated with cold resistance in plants (Yoon et al. 2017; Hu et al. 2018). MDA is the final product of membrane lipid peroxidation.
MDA binds and cross-links with proteins and enzymes on the cell membrane to
inactivate the structure and function of the biofilm, thereby destroying the
structure and function of the biofilm (He
et al. 2015). In our experiments, the soluble sugars of the two C.
oleifera cultivars significantly increased under the low-temperature
stress, and the results were similar to previous studies showing the soluble
sugar content is positively correlated with cold resistance in plants.
We observed the morphological structure of
the chloroplasts in the two C. oleifera cultivars. They suffered
serious injury under low-temperature stress, deformed by expansion of the inner
cyst lamella (Fig. 6). This hindered metabolism and decreased photosynthetic
efficiency, thus affecting the normal growth of both cultivars. Paraffin
sections revealed that the low-temperature stress increased leaf thickness in
both cultivars, suggesting that the plants changed their growth and morphology
in response to the stress. This may be an adaptive mechanism for coping with
low temperatures (Hu et al. 2016). The specific reasons need
to be further investigated.
Conclusion
C. oleifera
requires a particular temperature for flowering as low temperatures reduced
chlorophyll content, photosynthetic efficiency, fruit set rate, and yield. A
temperature of 6°C reduced net photosynthesis by 40 and 54% in Hua Shuo and Hua
Xin, respectively, compared to the normal temperature. Hua Shuo was better
adapted to low temperatures than Hua Xin as reflected by flowering phase and
photosynthetic parameters. Thus, low temperatures should be avoided to ensure
proper flowering and yield in C. oleifera.
Acknowledgments
This
study was supported by the Major Projects of Science and Technology Project of
Hunan Province [grant number 2018NK1030], and the Hunan Postgraduate Science
and Technology Innovation Project [grant number CX2018B438] and the Central
South University of Forestry and Technology Postgraduate Science and Technology
Innovation Fund Project [grant number 20181004].
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