Number of male and female flowers
The results showed that the total number of male and female flowers in cucumber was significantly higher under high nutrient conditions than that in low nutrient conditions, but pollination intensity did not significantly increase the total number of female and male flowers (Table 1, Fig. 1a, b). Furtherly, we found that the effect of nutrient on the number of male flowers was small, except for a peak at P2 level (Fig. 1a), but on 11th day after the end of pollination experiment, nutrient treatment had a significant effect on the number of male flowers (Table 1). This suggests that after the pollination experiment, the plant significantly increased the number of male flowers. Unlike male flower, the response of female flower to nutrient was more obvious, the number of female flowers under high nutrient condition was higher than that under low nutrient condition, and it was statistically significant under the treatment of pollination levels P1, P2 and P4 (Fig. 1b), indicating that the highest number of female flowers did not correspond with the highest pollination level (Fig. 1b).
In addition, we also analyzed the differences of male:female ratio of flower number, found that male:female flower ratio was slightly higher in low nutrient level than in high nutrient level (the mean ratio is 2.25±0.0059 in low nutrient, and in high nutrient the mean ratio is 2.23±0.0058), but the pollination treatment can significantly affect the ratio of male:female flowers per day (Table 1), indicating that cucumber will gradually increase the proportion of male flowers in the flowering stage.
Table 1 ANOVA showing the differences in numbers of male flower female flower, ratio of male:female flowers among different nutrient and pollination treatments. The data of flowers or ratio on the 36th day and the 43th day mean from the start of pollination experiment, that is the day after the pollination experiment (32 days).
Factors
|
Dependent v.
|
df
|
F
|
P value
|
Nutrient
|
Total no. of male flowers
|
1
|
4.80
|
0.031*
|
Total no. of female flowers
|
1
|
21.47
|
0.000**
|
No. of male flowers per day
|
32
|
1.48
|
0.084
|
No. of female flowers per day
|
32
|
1.68
|
0.033*
|
No. male flowers on the 36th day
|
1
|
2.50
|
0.117
|
No. male flowers on the 43th day
|
1
|
10.60
|
0.002**
|
No. female flowers on the 36th day
|
1
|
10.52
|
0.002**
|
No. female flowers on the 43th day
|
1
|
5.05
|
0.027*
|
Ratio of male:female flowers per day
|
32
|
0.943
|
0.562
|
Ratio of male:female flowers on the 36th day
|
1
|
1.01
|
0.317
|
Ratio of male:female flowers on the 43th day
|
1
|
0.35
|
0.554
|
Pollination
|
Total no. of male flowers
|
5
|
0.57
|
0.72
|
Total no. of female flowers
|
5
|
0.15
|
0.98
|
No. of male flowers per day
|
32
|
2.30
|
0.001**
|
No. of female flowers per day
|
32
|
1.61
|
0.045*
|
No. male flowers on the 36th day
|
32
|
0.97
|
0.437
|
No. male flowers on the 43th day
|
5
|
1.82
|
0.115
|
No. female flowers on the 36th day
|
5
|
0.88
|
0.501
|
No. female flowers on the 43th day
|
5
|
2.25
|
0.037*
|
Ratio of male:female flowers per day
|
32
|
1.60
|
0.047*
|
Ratio of male:female flowers on the 36th day
|
5
|
0.59
|
0.708
|
Ratio of male:female flowers on the 43th day
|
5
|
0.29
|
0.919
|
Nutrients and Pollination
|
Total no. of male flowers
|
5
|
0.80
|
0.554
|
Total no. of female flowers
|
5
|
0.15
|
0.98
|
No. of male flowers per day
|
32
|
1.42
|
0.104
|
No. of female flowers per day
|
32
|
2.15
|
0.003*
|
No. male flowers on the 36th day
|
5
|
0.20
|
0.962
|
No. male flowers on the 43th day
|
5
|
1.27
|
0.284
|
No. female flowers on the 36th day
|
5
|
0.62
|
0.688
|
No. female flowers on the 43th day
|
5
|
1.07
|
0.382
|
Ratio of male:female flowers per day
|
32
|
2.38
|
0.001**
|
Ratio of male:female flowers on the 36th day
|
5
|
0.44
|
0.819
|
Ratio of male:female flowers on the 43th day
|
5
|
1.06
|
0.384
|
Although pollination treatment had no effect on the total number of male and female flowers produced during the pollination experiment, the number of male flowers per day was positively affected by pollination, and the number of new female flowers increased significantly every day affected by nutrient or pollination, or their interactions (Table 1). Obviously, with the growth of cucumber plants, the flower number increased gradually during pollination days (Fig. 2) (Repeated Measures ANOVA, df =31, F =15.008, P =0.000<0.05). Moreover, the number of female flowers reached the highest on the 4th day after the pollination experiment (i.e., the 36th day of pollination experiment), while the number of female flowers decreased on the 11th day after the pollination experiment (i.e., the 43th day). Therefore, from the changes of the number of female flowers at the end of the pollination experiment, we can see the positive effect of pollination on the number of female flowers (Fig. 2). Furthermore, this indicated that the pollination may have a persistent effect on the increase of female flowers in cucumber.
If we consider the effect of daily pollination on the number of female flowers, we found that there were significant effects on some days in low or high nutrient level, which mainly appeared during the later stages of the pollination experiment (Fig. 2). In addition, different pollination levels lead to differences in the peak number of female flowers in low or high nutrient level during the pollination experiment, i.e., with the increase in the pollination degree, the number of female flowers gradually increased in the late stage of the pollination experiment, while under low pollination (P1), there was an early decrease in female flowers, esp. when the nutrient level is low (Fig. 2). Further, after the pollination experiment, the number of female flowers showed a significant decrease on the 11th day compared with the 4th day in low and high nutrient level (T-test, df=19, t=2.269, p=0.035<0.05) (Fig. 2), while the number of flowers in the control group without pollination did not decrease significantly.
Reproduction set
Nutrient and pollination treatments significantly affected the mass of fruits per plant produced in the pollination experimental stage but did not affect the fruits that were produced after the pollination experiment (Table 2, Fig. 3). Moreover, results showed that the maximum mass of fruits occurred at the low pollination level (P0) under high nutrient treatment, in other words, as the pollination strength increased, the mass of fruits did not increase accordingly. This trend was the same as that of the number of female flowers. At low nutrient levels, the pollination treatment did not significantly change the mass of fruits per plant compared with the blank control (Fig. 3). Nonetheless, the effect of pollination on a single fruit weight was different from themass of fruits per plant. The results showed that in high nutrient level, pollination increased the weight of a single fruit, esp., in P3 level, the weight of fruit was maximal and significantly higher than that in the low nutrient treatment (Fig. 4a). The results can be explained by the number of fruits and fruit set. In addition to the weight of fruit, pollination also significantly affected the number of fruit and the percentage fruit set (Table 2, Fig. 4b, c). In the results, the percentage of fruit set was very low, only around 10%.This also indicated that a large number of female flowers fail to produce fruit, which is consistent with the results of the number of female flowers increased by pollination. Pollination leads to an increase in female cucumber flowers, but the plant does not guarantee that all pollinated female flowers develop into fruit, thus increasing flower abortion. The patterns of fruit number and relative fruit set were similar to those of the mass of fruits at the two nutrient levels; the values were greatest at the P1 level, and there was also a small downward fluctuation with the increase in pollination levels (Fig. 4b, c). This means that a continued increase in pollination will reduce the number of fruit, thus decreasing the mass of fruits per plant; however, because the fruit number decreases, the average weight of a single fruit will increase.
The mean days of growing required to grow mature fruit and the fruit diameter and length were also recorded. The results indicated that the effects of the nutrient and pollination treatments were not significant on the average days of fruit growing from pollinated to harvest (Table 2, Fig. 5a), but at high nutrient levle, the days for growing to ripen appeared to be more stable than that at low nutrient level (Fig. 5). Though from the whole effects of nutrient and pollination, there were no statistical differences on fruit diameter and length (Table 2), we found that under high nutrient condition, the data of diameter and length of fruit have a peak, both in P3 pollination level reached maximum by nonlinear curve fitting, and the minimum in the control of pollination (P0), also, in the levels of pollination P4 and P5, fruit diameter and length were gradually decline (Fig. 5). So, this suggests that high intensity pollination didn’t increase the size of single fruit. In addition, interestingly, at the low nutrient levels, the effect of pollination on fruit diameter and fruit length was slightly opposite to that at high nutrient levels and high nutrient (Fig. 5). There is a trough at the level of pollination P4 in fruit diameter and P2 in fruit length. Thus, this suggests that pollination (P3) makes the fruit thinner and longer in nutrient abundance, while in nutrient deficiency, pollination makes the fruit thicker (in P4) and shorter (in P2) (Fig. 5).
Table 2 ANOVA showing the differences in Mass of fruits per plant, number of fruit, fruit set, fruit size, fruit weight and number of seeds among different nutrient and pollination treatments. The data of mass of fruits per plant (non-pollinated fruits) are statistics of the fruits produced after the pollination experiment was stopped, and the data of mass of fruits per plant (pollinated fruits) are statistics of the fruits produced during pollination experiment.
Factors
|
Dependent v.
|
df
|
F
|
P value
|
Nutrient
|
Mass of fruits per plant (non-pollinated fruits)
|
1
|
1.20
|
0.278
|
Mass of fruits per plant (pollinated fruits)
|
1
|
31.27
|
0.000***
|
Weight of fruit
|
1
|
0.06
|
0.810
|
No. fruit
|
1
|
18.11
|
0.000***
|
Percentage fruit set
|
1
|
7.03
|
0.009*
|
Mean days of fruit growing
|
1
|
0.15
|
0.702
|
Mean diameter of fruit
|
1
|
0.23
|
0.635
|
Mean length of fruit
|
1
|
0.04
|
0.839
|
Total no. seeds per plant
|
1
|
15.06
|
0.000***
|
No. of seeds per fruit
|
1
|
0.09
|
0.766
|
Pollination
|
Mass of fruits per plant (Non-pollinated fruits)
|
5
|
0.63
|
0.681
|
Mass of fruits per plant (Pollinated fruits)
|
5
|
2.39
|
0.043*
|
Weight of fruit
|
5
|
1.64
|
0.157
|
No. fruit
|
5
|
2.49
|
0.036*
|
Percentage fruit set
|
5
|
2.62
|
0.029*
|
Mean days of fruit growing
|
5
|
1.59
|
0.173
|
Mean diameter of fruit
|
5
|
0.76
|
0.580
|
Mean length of fruit
|
5
|
1.15
|
0.339
|
Total no. seeds per plant
|
5
|
6.78
|
0.000***
|
No. of seeds per fruit
|
5
|
6.53
|
0.000***
|
Nutrients and Pollination
|
Mass of fruits per plant (Non-pollinated fruits)
|
5
|
0.30
|
0.915
|
Mass of fruits per plant (Pollinated fruits)
|
5
|
4.09
|
0.002*
|
Weight of fruit
|
5
|
1.40
|
0.232
|
No. fruit
|
5
|
1.32
|
0.262
|
Percentage fruit set
|
5
|
0.91
|
0.476
|
Mean days of fruit growing
|
5
|
1.40
|
0.235
|
Mean diameter of fruit
|
5
|
2.99
|
0.016*
|
Mean length of fruit
|
5
|
2.12
|
0.072
|
Total no. seeds per plant
|
5
|
1.30
|
0.564
|
No. of seeds per fruit
|
5
|
0.78
|
0.270
|
Seed production
Because the fruit set was very low, at some plants, there were few fruits produced, so we harvest all the fruits and counted the seeds number of each fruit. The results showed that the seed production was significantly affected by nutrients and pollination. Obviously, no seeds were produced in the non-pollination experiments, so the plots is empty (Fig. 6). At high nutrient levels, the number of seeds per fruit was higher at the P2 and P3 levels than in the control (Fig. 6a). At the low nutrient level, however, the number of seeds per fruit at the P2 level was almost the lowest. Unlike seed number per fruit, the total number of seeds per plant was significantly affected by the interaction between nutrients and pollination (Table 1), and differed from the number of seeds per fruit among pollination levels under low nutrient treatment (Fig. 6b). By contrast, the lowest value was observed at the P2 level, but an increase was observed at the P4 level (Fig. 6b).
Correlation between flowers and fruits and seeds
Correlation analysis showed that the female flowers were positively correlated with the number of male flowers, number of fruit, mass of fruits per plant, total number of seeds per plant, and the shoot mass but were not correlated with the weight of fruit and the number of seeds per fruit (Table 2). Moreover, the number of fruit per plant was positively correlated with the mass of fruits and the total number of seeds per plant but was uncorrelated or negatively correlated with the weight of single fruit, the number of seeds per fruit, and the shoot mass. Additionally, the mass of fruits per plant was also related to the total seed number per plant (Table 2). These results showed that the number of fruits increased with the number of female flowers and that the fruits per plant had a greater mass and a higher number of total seeds; however, there was also a decline in the weight of fruit (Fig. 4a) and the number of seeds per fruit. Accordingly, the total number of seeds per plant was negatively correlated with the number of seeds per fruit, and the number of seeds per fruit was positively correlated with the fruit weight (Table 2). These results showed that a high number of fruit per plant can lead to reductions in the weight of a single fruit and that smaller fruit produces fewer seeds. Additionally, the shoot mass was negatively correlated with the number of fruit and the mass of fruits, and the total number of seeds showed a trade-off between vegetation growth and reproductive allocation (Table 2).
Table 2 Correlation analysis of cucumber flower, fruit, and seed traits using Pearson’s linear simple correlation with two-tailed P values.
|
|
Female flowers
per plant
|
No. fruits per plant
|
Non-pollinated female flowers
|
Male flowers
per plant
|
Non-pollinated male flowers
|
Mass of fruits per plant
|
Total seeds per plant
|
No. fruits per plant
|
r
|
0.337**
|
|
|
|
|
|
|
p
|
0.000
|
|
|
|
|
|
|
n
|
117
|
|
|
|
|
|
|
Non-pollinated female flowers
|
r
|
0.349**
|
0.049
|
|
|
|
|
|
p
|
0.000
|
0.601
|
|
|
|
|
|
n
|
117
|
117
|
|
|
|
|
|
Male flowers per plant
|
r
|
0.451**
|
0.088
|
0.246**
|
|
|
|
|
p
|
0.000
|
0.345
|
0.007
|
|
|
|
|
n
|
117
|
117
|
117
|
|
|
|
|
Non-pollinated male flowers
|
r
|
0.259**
|
-0.173
|
0.147
|
0.444**
|
|
|
|
p
|
0.005
|
0.063
|
0.113
|
0.000
|
|
|
|
n
|
117
|
117
|
117
|
117
|
|
|
|
Mass of fruits per plant
|
r
|
0.337**
|
.892**
|
-0.053
|
0.135
|
-0.152
|
|
|
p
|
0.000
|
0.000
|
0.569
|
0.145
|
0.101
|
|
|
n
|
117
|
117
|
117
|
117
|
117
|
|
|
Total seeds per plant
|
r
|
0.236*
|
0.721**
|
-0.033
|
0.139
|
-0.146
|
0.849**
|
|
p
|
0.01
|
0.000
|
0.725
|
0.136
|
0.116
|
0.000
|
|
n
|
117
|
117
|
117
|
117
|
117
|
117
|
|
Shoot mass
|
r
|
0.387**
|
-0.039
|
0.380**
|
0.293**
|
0.523**
|
-0.065
|
-0.067
|
p
|
0.000
|
0.681
|
0.000
|
0.001
|
0.000
|
0.488
|
0.473
|
n
|
117
|
117
|
117
|
117
|
117
|
117
|
117
|
Seeds per fruit
|
r
|
0.051
|
-0.034
|
0.135
|
0.124
|
-0.007
|
-0.057
|
-0.042
|
p
|
0.593
|
0.714
|
0.146
|
0.182
|
0.942
|
0.539
|
0.656
|
n
|
117
|
117
|
117
|
117
|
117
|
117
|
117
|
Fruit weight
|
r
|
0.019
|
0.077
|
0.093
|
0.101
|
-0.051
|
0.053
|
0.012
|
p
|
0.861
|
0.465
|
0.379
|
0.336
|
0.629
|
0.614
|
0.911
|
n
|
92
|
92
|
92
|
92
|
92
|
92
|
92
|