3.1. Prevailing solar radiations
The prevalence of PPFD was regularly monitored on sunny and cloudy days in the morning (9.00-9.30 AM), noon (12.00-12.30 PM) and afternoon (3.00-3.30 PM) using LI-6400XT portable photosynthesis system. Multiple observations were recorded to nullify the temporal effect. Analysis of variance revealed significant differences (P<0.001) in PPFD amongst sunny and cloudy days. The interactive effects of sunlight conditions and timings were found highly significant (P<0.001) (Table 2).
Table 2. Mean square and significance level for sunlight condition, timing and their interactive effect.
Sources of variation
|
DF
|
Sum of Squares
|
Mean sum of squares
|
F value
|
Pr(>F)
|
Sunlight condition (S)
|
1
|
17817392.4
|
17817392.43
|
433.305
|
P<0.001
|
Timings (T)
|
2
|
3867567.5
|
1933783.74
|
47.028
|
P<0.001
|
S × T
|
2
|
912604.6
|
456302.28
|
11.097
|
P<0.001
|
Error
|
132
|
5427809.2
|
41119.77
|
-
|
-
|
Concerning the sunlight conditions, the highest mean PPFD was observed during sunny days (1544.18±290.68) in comparison to cloudy days (825.54±256.16), whereas in case of timings, the PPFD was found during noon (1420.14±539.70) in comparison to morning (1044.36±329.00) and afternoon (1090.08±370.56) (Fig. 1, 2). Sunny × Noon condition resulted in realizing the highest PPFD (1893.62±227.28). It was worth noting that across sunlight conditions and timings, the solar radiations were as low as 220.50. Overall, the cloudy condition caused up to ~46% reduction in solar radiations in comparison to sunny conditions (Table 3 and Fig. 3).
Fig. 1. Box and whisker plot showing the median, median and quartile range of the incident sunlight radiations under cloudy and sunny day conditions. Tukey’s post hoc test was adopted to compare the means at P<0.05 level of significance. The treatment effects were highly significant at P<0.001.
Fig. 2. Box and whisker plot showing the median, median and quartile range of the incident sunlight radiations at morning (9.00-9.30 AM), noon (12.00-12.30 PM) and afternoon Afternoon (3.00-3.30 PM). Tukey’s post hoc test was adopted to compare the means at P<0.05 level of significance. The treatment effects were highly significant at P<0.001.
Fig. 3. Box and whisker plot showing main effects, median, median and quartile range of the incident sunlight radiations at morning (9.00-9.30 AM), noon (12.00-12.30 PM) and afternoon Afternoon (3.00-3.30 PM). Tukey’s post hoc test was adopted to compare the means at P<0.05 level of significance. The treatment effects were highly significant at P<0.001.
Table 3. Summary of the statistics of sunlight condition, timing and their interactive effects.
|
Mean±std
|
Minimum
|
Maximum
|
Sunlight condition (S)
|
|
Cloudy
|
825.54±256.16b
|
220.50
|
1411.22
|
Sunny
|
1544.18±290.68a
|
1126.0
|
2205.07
|
Timings (T)
|
Noon (12.00-12.30 PM)
|
1420.14±539.70a
|
220.56
|
2205.25
|
Afternoon (3.00-3.30 PM)
|
1090.08±370.56b
|
231.06
|
1500.47
|
Morning (9.00-9.30 AM)
|
1044.36±329.00b
|
294.08
|
1569.36
|
Interactive effects (S × T)
|
Cloudy × Noon
|
946.65±274.55c
|
220.57
|
1411.22
|
Cloudy × Afternoon
|
775.84±262.55d
|
231.09
|
1354.58
|
Cloudy × Morning
|
754.13±187.77d
|
294.03
|
1029.01
|
Sunny × Noon
|
1893.62±227.28a
|
1472.16
|
2205.00
|
Sunny × Afternoon
|
1404.32±73.90b
|
1269.04
|
1500.07
|
Sunny × Morning
|
1334.59±100.11b
|
1126.03
|
1569.08
|
3.2. Analysis of variance
A two-way analysis of variance was performed to test the effect of four CO2 concentrations, nine PPFD and their interaction effect on the photosynthetic efficiency of yam bean. All the parameters under consideration were significantly impacted due to CO2 (P<0.0001), PPDF (P<0.0001) and their interaction (P<0.0001) (Table 4). Tukey’s post hoc test for mean comparisons at P<0.05 found that the mean Pn, gs, Ci, Ci/Ca and E differed significantly between CO2 (P<0.0001), PPFD (P<0.0001) and their interactive effect (CO2 × PPFD; P<0.0001) (Table 4). Similar results were reported by Ravi et al. (2022) in yams when exposed to ECO2.
Table….4. Mean square and significance level for net photosynthetic rate (Pn), stomatal conductance (gs), Ci (intercellular CO2 concentration), Ci/Ca (ratio of intercellular to ambient CO2 concentration) and transpiration (E) in yam bean leaves exposed to varying PPFD and carbon dioxide concentration.
Source of variation
|
df
|
Mean Square
|
F
|
Significance level
|
Pn
|
CO2
|
3
|
935.67
|
555.839
|
P<0.0001
|
PPFD
|
8
|
23260.37
|
13817.870
|
P<0.0001
|
CO2 × PPFD
|
24
|
14.132
|
8.395
|
P<0.0001
|
Error
|
1189
|
1.683
|
-
|
-
|
gs
|
CO2
|
3
|
1.370
|
183.897
|
P<0.0001
|
PPFD
|
8
|
0.096
|
12.829
|
P<0.0001
|
CO2 × PPFD
|
24
|
0.042
|
5.691
|
P<0.0001
|
Error
|
1189
|
0.007
|
-
|
-
|
Ci
|
|
|
|
|
CO2
|
3
|
19873168.28
|
78857.555
|
P<0.0001
|
PPFD
|
8
|
515607.06
|
2045.950
|
P<0.0001
|
CO2 × PPFD
|
24
|
3120.75
|
12.383
|
P<0.0001
|
Error
|
1189
|
252.01
|
-
|
-
|
Ci/Ca
|
CO2
|
3
|
1.051
|
1220.663
|
P<0.0001
|
PPFD
|
8
|
1.046
|
1214.515
|
P<0.0001
|
CO2 × PPFD
|
24
|
0.050
|
58.223
|
P<0.0001
|
Error
|
1189
|
0.001
|
-
|
-
|
E
|
CO2
|
3
|
25.278
|
129.481
|
P<0.0001
|
PPFD
|
8
|
175.821
|
900.602
|
P<0.0001
|
CO2 × PPFD
|
24
|
4.013
|
20.556
|
P<0.0001
|
Error
|
1189
|
0.195
|
-
|
-
|
3.3. Effect of ECO2 and PPFDs on Photosynthetic efficiency
Exposure to various CO2 concentrations and PPFD had a significant effect on the net photosynthetic rate (Table 5a and Fig. 4). ECO2 (600, 800 and 1000 ppm) positively enhanced the net photosynthetic rate of yam bean in comparison to ambient concentration (400 ppm). Whereas, the Pn rate increased linearly across increasing PPFD. The highest net photosynthesis rate was achieved at the interactive level of 600 × 1500 (37.05±02.85) and 800 × 1500 (36.65±0.95) with similar significance levels. Contrastingly, the lowest Pn rate was recorded at the lowest PPFD level (50 µmol m-2 s-1). Pn rate declined severely as PPFD decreased from 600 to 50 μmol m-2 s-1 in relation to 1500 μmol m-2 s-1. Nevertheless, a decrease in Pn rate at PPFD 600 to 50 μmol m-2 s-1, relative to 1500 μmol m-2 s-1, was less at elevated CO2 of 600 and 800 ppm relative to 400 ppm.
Unlike the Pn rate, the trend of gs of yam bean leaves across ECO2 as well as nine PPFD levels PPFD was inconsistent (Table 5a and Fig. 4). The gs in leaves of yam bean varied between 0.26 and 0.59 at 400-1000 ppm. The gs at 1000 ppm were greater by 6.25 to 83.33% relative to ambient CO2. The gs was not affected at lower PPFD 800-50 μmol m-2 s-1 relative to saturation PPFD (1500 μmol m-2 s-1). Stomatal conductance (gs) is diminished under elevated CO2. Partial stomatal closure, and associated reduction in stomatal conductance, are general plant responses under elevated CO2 (Medlyn et al. 2001; Ainsworth and Rogers 2007; Gao et al. 2015).
Table 5a. Main effects and significance level for net photosynthetic rate (Pn) and stomatal conductance (gs) in yam bean leaves exposed to varying PPFD and carbon dioxide concentration.
CO2
|
PPFD
|
Mean±std
|
No. of observations
|
CO2
|
PPFD
|
Mean±std
|
No. of observations
|
Pn
|
400
|
50
|
00.37±00.32r
|
n=47
|
800
|
50
|
01.68±0.83q
|
n=36
|
100
|
03.35±00.58p
|
n=37
|
100
|
05.32±0.73o
|
n=48
|
200
|
09.03±00.33m
|
n=28
|
200
|
11.86±0.62l
|
n=33
|
400
|
18.48±00.63k
|
n=26
|
400
|
21.94±0.51ij
|
n=31
|
600
|
25.03±00.74h
|
n=31
|
600
|
26.90±0.58fg
|
n=50
|
800
|
27.88±01.35f
|
n=33
|
800
|
29.76±0.37e
|
n=38
|
1000
|
31.15±00.67cd
|
n=29
|
1000
|
32.08±0.39c
|
n=32
|
1200
|
32.07±00.86c
|
n=30
|
1200
|
34.33±0.59b
|
n=29
|
1500
|
34.09±00.51b
|
n=26
|
1500
|
36.65±0.95a
|
n=38
|
600
|
50
|
1.92±01.96q
|
n=45
|
1000
|
50
|
-2.63±0.52s
|
n=29
|
100
|
5.96±02.09o
|
n=25
|
100
|
00.89±0.23qr
|
n=35
|
200
|
11.63±02.15l
|
n=34
|
200
|
07.26±0.27n
|
n=39
|
400
|
21.17±02.11j
|
n=32
|
400
|
18.02±0.40k
|
n=35
|
600
|
26.62±02.52g
|
n=32
|
600
|
23.01±0.40i
|
n=47
|
800
|
30.50±02.66de
|
n=32
|
800
|
26.03±0.38gh
|
n=34
|
1000
|
31.38±02.79cd
|
n=29
|
1000
|
29.49±0.86e
|
n=35
|
1200
|
34.75±02.64b
|
n=28
|
1200
|
31.69±1.13cd
|
n=30
|
1500
|
37.05±02.85a
|
n=21
|
1500
|
34.81±1.26b
|
n=41
|
gs
|
400
|
50
|
0.41±0.06hijk
|
n=47
|
800
|
50
|
0.29±0.02m
|
n=36
|
100
|
0.37±0.09ijkl
|
n=37
|
100
|
0.31±0.06lm
|
n=48
|
200
|
0.44±0.17defghij
|
n=28
|
200
|
0.34±0.04klm
|
n=33
|
400
|
0.42±0.15efghijk
|
n=26
|
400
|
0.33±0.03klm
|
n=31
|
600
|
0.49±0.14abcdef
|
n=31
|
600
|
0.35±0.03klm
|
n=50
|
800
|
0.52±0.06abcd
|
n=33
|
800
|
0.35±0.03klm
|
n=38
|
1000
|
0.49±0.02abcdefg
|
n=29
|
1000
|
0.37±0.03ijklm
|
n=32
|
1200
|
0.44±0.02defghij
|
n=30
|
1200
|
0.37±0.03ijklm
|
n=29
|
1500
|
0.36±0.02jklm
|
n=26
|
1500
|
0.37±0.05ijkl
|
n=38
|
600
|
50
|
0.38±0.11ijk
|
n=45
|
1000
|
50
|
0.47±0.04bcdefgh
|
n=29
|
100
|
0.45±0.13cdefghi
|
n=25
|
100
|
0.48±0.03bcdefgh
|
n=35
|
200
|
0.44±0.16defghij
|
n=34
|
200
|
0.49±0.05abcdef
|
n=39
|
400
|
0.41±0.13fghijk
|
n=32
|
400
|
0.51±0.05abcd
|
n=35
|
600
|
0.40±0.14ghijk
|
n=32
|
600
|
0.54±0.06ab
|
n=47
|
800
|
0.45±0.13cdefghi
|
n=32
|
800
|
0.49±0.04abcdef
|
n=34
|
1000
|
0.45±0.16cdefghi
|
n=29
|
1000
|
0.53±0.02abc
|
n=35
|
1200
|
0.56±0.15a
|
n=28
|
1200
|
0.50±0.02abcde
|
n=30
|
1500
|
0.48±0.14abcdefgh
|
n=21
|
1500
|
0.49±0.05abcdef
|
n=41
|
Fig. 4. Response of leaf net photosynthetic rate (Pn) and stomatal conductance (gs) in yam bean to interactive effects of varying irradiance and elevated carbon dioxide. Values are mean±std. Tukey’s post hoc test was adopted to compare the means at P<0.05 level of significance. The treatment effects were highly significant at P<0.0001. The length of bars shows the variability with standard error. Note: PPFD: photosynthetic photon flux density.
It was found that Ci increased linearly as an effect of ECO2 (Table 5b and Fig. 5). Strikingly, the mean Ci decreased linearly across increasing PPFD levels irrespective of the ECO2 concentration. The rise in Ci at 50 μmol m-2 s-1 relative to saturation PPFD (1500 μmol m-2 s-1) amounted to 17.99% at 1000 ppm to 92.38% at 400 ppm. The drastic reduction in Pn rates at lower PPFD (800-50 μmol m-2 s-1) accounted for 4.62-92.38% greater intercellular CO2 concentrations at the lower PPFD relative to saturation PPFD. This indicates that at lower PPFD Ci was not a rate-limiting factor for photosynthesis.
Analysis of the data revealed that the mean Ci/Ca increased significantly and linearly across CO2 concentration (Table 5b and Fig. 5). Whereas, the mean Ci/Ca reduced significantly at increasing PPFD. Interestingly, the extent of reduction of Ci/Ca as an effect of PPFD was lesser at ECO2 (600-1000 ppm) in comparison to ambient CO2 concentration (400 ppm). Amongst ECO2, the reduction in Ci/Ca as an effect of PPFD was relatively small at 1000 ppm in comparison to 600 and 800 ppm, indicating that ECO2 restricted the Ci/Ca reduction under limited PPFD. The mean Ci/Ca increased in the range of 8-19% as an effect of ECO2. The highest mean Ci/Ca was observed at 50 and 100 PPFD across all CO2 concentrations. However, statistically, the effects were the same.
Table 5b. Main effects and significance level for leaf Ci (intercellular CO2 concentration) and Ci/Ca (ratio of intercellular to ambient CO2 concentration) in yam bean leaves exposed to varying PPFD and carbon dioxide concentration.
CO2
|
PPFD
|
Mean±std
|
N=
|
CO2
|
PPFD
|
Mean±std
|
No. of observations
|
Ci
|
400
|
50
|
391.96±2.37v
|
n=47
|
800
|
50
|
776.28±4.66h
|
n=36
|
100
|
375.28±6.39w
|
n=37
|
100
|
754.56±1.81i
|
n=48
|
200
|
347.74±16.40x
|
n=28
|
200
|
718.97±4.88j
|
n=33
|
400
|
298.79±24.02y
|
n=26
|
400
|
659.44±10.97k
|
n=31
|
600
|
283.48±26.06yz
|
n=31
|
600
|
635.42±14.31l
|
n=50
|
800
|
282.25±9.46z
|
n=33
|
800
|
620.20±13.85m
|
n=38
|
1000
|
265.24±5.14A
|
n=29
|
1000
|
613.87±11.05mn
|
n=32
|
1200
|
248.40±3.84B
|
n=30
|
1200
|
601.53±13.87n
|
n=29
|
1500
|
210.47±10.73C
|
n=26
|
1500
|
584.95±32.27o
|
n=38
|
600
|
50
|
583.21±6.57o
|
n=45
|
1000
|
50
|
996.00±2.01a
|
n=29
|
100
|
565.75±2.39p
|
n=25
|
100
|
981.22±1.55a
|
n=35
|
200
|
536.21±9.32q
|
n=34
|
200
|
954.20±2.76b
|
n=39
|
400
|
483.77±21.54r
|
n=32
|
400
|
911.77±6.03c
|
n=35
|
600
|
450.45±33.33st
|
n=32
|
600
|
894.49±9.25d
|
n=47
|
800
|
443.97±36.21st
|
n=32
|
800
|
872.95±7.76e
|
n=34
|
1000
|
435.80±32.68tu
|
n=29
|
1000
|
865.42±2.19e
|
n=35
|
1200
|
452.16±27.99s
|
n=28
|
1200
|
849.58±3.57f
|
n=30
|
1500
|
421.91±32.21u
|
n=21
|
1500
|
832.49±10.62g
|
n=41
|
Ci/Ca
|
400
|
50
|
0.98±0.00ab
|
n=47
|
800
|
50
|
0.97±0.00abcd
|
n=36
|
100
|
0.95±0.00ef
|
n=37
|
100
|
0.95±0.00def
|
n=48
|
200
|
0.88±0.01hi
|
n=28
|
200
|
0.91±0.01gh
|
n=33
|
400
|
0.77±0.01opq
|
n=26
|
400
|
0.84±0.01kl
|
n=31
|
600
|
0.74±0.01rs
|
n=31
|
600
|
0.81±0.00mn
|
n=50
|
800
|
0.74±0.01rs
|
n=33
|
800
|
0.80±0.00no
|
n=38
|
1000
|
0.70±0.01t
|
n=29
|
1000
|
0.79±0.01nop
|
n=32
|
1200
|
0.66±0.01u
|
n=30
|
1200
|
0.78±0.01opq
|
n=29
|
1500
|
0.56±0.01v
|
n=26
|
1500
|
0.76±0.00qrs
|
n=38
|
600
|
50
|
0.98±0.00abc
|
n=45
|
1000
|
50
|
1.00±0.01a
|
n=29
|
100
|
0.95±0.01cdef
|
n=25
|
100
|
0.98±0.00ab
|
n=35
|
200
|
0.91±0.01gh
|
n=34
|
200
|
0.96±0.00bcde
|
n=39
|
400
|
0.83±0.01lm
|
n=32
|
400
|
0.93±0.00fg
|
n=35
|
600
|
0.78±0.01opq
|
n=32
|
600
|
0.91±0.00gh
|
n=47
|
800
|
0.77±0.01pqr
|
n=32
|
800
|
0.89±0.01hi
|
n=34
|
1000
|
0.76±0.01qrs
|
n=29
|
1000
|
0.89±0.00hi
|
n=35
|
1200
|
0.79±0.01op
|
n=28
|
1200
|
0.87±0.01ij
|
n=30
|
1500
|
0.74±0.01s
|
n=21
|
1500
|
0.86±0.00jk
|
n=41
|
Fig. 5. Response of leaf Ci/Ca (ratio of intercellular to ambient CO2 concentration) and Ci (intercellular CO2 concentration) in yam bean to interactive effects of varying irradiance and elevated carbon dioxide. Values are mean±std. Tukey’s post hoc test was adopted to compare the means at P<0.05 level of significance. The treatment effects were highly significant at P<0.0001. The length of bars shows the variability with standard error. Note: PPFD: photosynthetic photon flux density.
Analysis of variance showed that E at all CO2 concentrations and PPFD increased significantly and linearly (Table 5c and Fig. 6). Irrespective of the CO2 concentrations, the lowest E was observed at the lowest PPFD. Subjecting yam bean leaves to ECO2 substantially increased E across PPFD. The highest E was recorded at 600, 800 and 1000 ppm between 1000-1500 PPFD concerning the least PPFD and ECO2 concentration. The quantum of rise in E at the highest PPFD was higher at 600 (128%) and 800 ppm (135%) in comparison to 1000 ppm (80%) and 400 ppm (58%) in comparison to the lowest PPFD. This indicated that ECO2 facilitated the sustenance of transpiration at the lowest PPFD.
Table 5c. Main effects and significance level for transpiration (E) in yam bean leaves exposed to varying PPFD and carbon dioxide concentration.
CO2
|
PPFD
|
Mean±std
|
No. of observations
|
CO2
|
PPFD
|
Mean±std
|
No. of observations
|
E
|
400
|
50
|
3.14±0.11mnop
|
n=47
|
800
|
50
|
2.65±0.20q
|
n=36
|
100
|
3.02±0.52nopq
|
n=37
|
100
|
2.94±0.53opq
|
n=48
|
200
|
3.52±1.00klm
|
n=28
|
200
|
3.30±0.34lmno
|
n=33
|
400
|
3.69±1.07kl
|
n=26
|
400
|
3.66±0.28kl
|
n=31
|
600
|
4.76±0.98ghi
|
n=31
|
600
|
4.36±0.18j
|
n=50
|
800
|
5.44±0.31de
|
n=33
|
800
|
4.74±0.16ghi
|
n=38
|
1000
|
5.51±0.20d
|
n=29
|
1000
|
5.33±0.21def
|
n=32
|
1200
|
5.36±0.18def
|
n=30
|
1200
|
5.64±0.30cd
|
n=29
|
1500
|
4.95±0.30fgh
|
n=26
|
1500
|
6.23±0.62ab
|
n=38
|
600
|
50
|
2.89±0.44pq
|
n=45
|
1000
|
50
|
3.57±0.10kl
|
n=29
|
100
|
3.35±0.54lmno
|
n=25
|
100
|
3.67±0.08kl
|
n=35
|
200
|
3.33±0.60lmn
|
n=34
|
200
|
3.91±0.12k
|
n=39
|
400
|
3.56±0.30kl
|
n=32
|
400
|
4.46±0.17ij
|
n=35
|
600
|
3.91±0.42k
|
n=32
|
600
|
5.07±0.11efg
|
n=47
|
800
|
4.63±0.53hij
|
n=32
|
800
|
5.30±0.27def
|
n=34
|
1000
|
5.53±0.74d
|
n=29
|
1000
|
6.01±0.28bc
|
n=35
|
1200
|
6.51±0.55a
|
n=28
|
1200
|
6.04±0.22bc
|
n=30
|
1500
|
6.61±0.51a
|
n=21
|
1500
|
6.46±0.42a
|
n=41
|
Fig. 6. Response of transpiration (E) in yam bean to interactive effects of varying irradiance and elevated carbon dioxide. Values are mean±std. Tukey’s post hoc test was adopted to compare the means at P<0.05 level of significance. The treatment effects were highly significant at P<0.0001. The length of bars shows the variability with standard error. Note: PPFD: photosynthetic photon flux density.
Solar radiation is the pre-requisite of photosynthesis (Taiz and Zigger 2002). It is of tremendous importance to sustain the photosynthetic efficiency of cultivated plants under climatic uncertainties to ensure food supply to ever-growing populations (Greenwald et al. 2006). However, plants often come across fluctuating light intensities resulting in the employment of efficient photosynthetic apparatus under limited PPFD availability (low light condition/global dimming) or the adoption of excessive sunlight dissipation strategy to hinder associated photo-oxidative stress. It would be crucial to understand the effect of varying PPFD on photosynthetic efficiency and its implications for crop productivity (Durand et al. 2021). Crop physiological responses under varying PPFD are further convoluted with the co-occurrence of other abiotic stresses (Demmig-Adams et al. 2015). In addition, elevated carbon dioxide is likely to impact the physiological processes of plants. It is well documented that ECO2 has sizeable effects on crop physiology ranging from physio-molecular levels to the ecosystem level because of its central role in the photosynthetic process (Becklin et al. 2016; Dong et al. 2018; Poorter et al. 2022). The photosynthetic responses under combined effects of varying PPFD and ECO2 could provide vital information on the physiological efficiency of the plant (Urban et al. 2014). Aiming this, the present study was undertaken to elucidate the photosynthetic responses of yam bean to varying PPFD and ECO2.
Several factors impact the photosynthetic efficiency of plants namely, Sunlight or cloudy condition (Wang et al. 2020), canopy cover (Urban et al. 2007; Durand et al. 2021), angle of radiation incidence/penetration (Brodersen and Vogelmann 2010), leaf anatomy and chlorophyll distribution (Brodersen and Vogelmann 2010; Ichiro et al. 2016) and species-specific (Berry and Goldsmith 2020). Among this sunlight availability is the crucial determining factor for the photosynthesis. The Pn rate increased steadily at each increasing PPFD. The Pn rate was maximum at 1500 μmol m-2 s-1 in comparison to 50 μmol m-2 s-1 at each CO2 level. Whereas the Pn rate increased steadily up to 600 and 800 ppm in comparison to 400 ppm followed by acclimation at 1000 ppm. Based on these results, PPFD were categorized into low light (400-800 μmol m-2 s-1) and high light conditions (1000-1500 μmol m-2 s-1). Consequences of low light conditions were offset by elevated CO2 concentration as the Pn rate increased by 9.02 and 9.44% at 600 and 800 ppm, respectively, in comparison to 400 ppm. However, effects were less pronounced under high light conditions as there was only a 5.52 and 4.48% increase in net photosynthetic rate under 600 and 800 ppm, respectively, over 400 ppm. This indicates on cloudy days elevated CO2 up to 600-800 ppm can sustain the Pn rate of yam bean. Although the Pn increased with increasing PPFD from 50 to 1500 μmol m-2 s-1, the degree of increase in Pn declined as PPFD and CO2 concentration increased. At 1000 ppm CO2, the Pn rate decreased relative to 800, 600 and 400 ppm CO2 at PPFD between 50 and 1500 μmol m-2 s-1. This indicated that even under higher PPFD of 1000-1500 μmol m-2 s-1, a rise in CO2 above 800 ppm is not beneficial for photosynthesis in yam bean. Previous reports by (Ravi et al. 2017) in sweet potato, (Ravi et al. 2018) in elephant foot yam, (Ravi et al. 2020) in yam bean and (Ravi et al. 2022) in yams have depicted the maximum Pn rate at 1500 μmol m-2 s-1. PPFD changes constantly within a day because of fluctuating sunlight radiation or cloudy conditions causing severe changeability in net photosynthetic rates. Moreover, the persistence of cloudy conditions for a longer period amplifies the ratio of diffuse to direct solar radiation leading to reduced temperature and VPD (Urban et al. 2014; Wimalasekera 2019; Durand et al. 2021). However, the Pn rate under these effects is very much aligned with the light compensation point (LCP) of the respective species (Wimalasekera 2019). The capacity of LCP is a crucial determinant of a plant’s tolerance to varying PPFD (Walters and Reich 2000). Varying sunlight availability could potentially impact leaf level (Brodersen et al. 2008) to canopy level (Kanniah et al. 2013) CO2 assimilation and stomatal aperture individually or in combination with other environmental stressors (Urban et al. 2007, 2012, 2014; Barillot et al. 2010; Durand et al. 2021). According to the results of (Brodersen et al. 2008), leaf photosynthesis could be as high as 15-20% under direct light compared to diffused sunlight under cloudy conditions. Multiple palisade layers adapted under high-light conditions amplified the photosynthetic efficiency in comparison to leaves subjected to low-light conditions (Vogelman et al. 1996). Sun-adapted leaves grown under ECO2 (700 ppm) elicited higher carbon assimilation (~315%) as compared to the leaves grown under cloudy conditions. Moreover, leaf photosynthesis was significantly decelerated in leaves grown under a cloudy sky with ambient CO2 concentration (385 ppm) in comparison to sun-adapted-ECO2 condition (Urban et al. 2014). The fluctuation in photosynthesis under varying PPFD or sunlight conditions (Morales and Kaiser 2020) is attributed to the inherent PSII saturation capacity. Leaves exposed to cloudy conditions became light-saturated more easily (Earles et al. 2017) in comparison to the leaves exposed to sunny conditions. More work is needed to decipher plant’s photosynthetic responses at the canopy level given long-term changes in the diffuse fraction of solar radiation (Durand et al. 2021). Cloudy conditions offer limited substrate for photosynthesis leading to restricted CO2 assimilation activity. These results corresponded with Bernacchi et al. (2003) where the photosynthetic efficiency of leaves under the cloudy conditions was down-regulated because of limited RuBP activity, ATP and NADPH production and reduced electron transport rate. According to Urban et al. (2014), daily sum of fixed CO2 and light use efficiency of Fagus sylvatica was significantly reduced under the cloudy condition in comparison to sunlight condition mediated by xanthophylls dependent thermal dissipation of absorbed light energy. Further, the Pn rate was reduced as a consequence of increased thermal dissipation of solar radiation within PS II antennae associated with increased de-epoxidation state of the xanthophyll cycle pigments (Eskling et al. 1997; Urban et al. 2014). However, Urban et al. (2014) and Kets et al. (2010) inferred that studies assessing the photosynthetic efficiency under interactive effects of ECO2 and varying PPFD have received limited attention.
ECO2 offsets the negative impact of limited PPFD on photosynthetic efficiency. As per the research results of (Vanaja et al. 2011; Reich et al. 2018) C3 and C4 crops exhibited varied responses concerning the photosynthesis under enriched carbon enrichment. ECO2 is known to induce positive responses in plants, especially in the form of enhanced photosynthetic rates. Enhancement in photosynthetic efficiency as a consequence of ECO2 has been reported in other leguminous crops. Increases in photosynthetic rate in soybean Vu et al. (1997) (up to 33%) and Prasad et al. (2005) (up to 66%) at 1000 ppm compared to 400 ppm CO2 have been reported. Similar results were reported in soybean (Bhatt et al. 2010) as Pn rates were elevated up to 48-94% under long-term exposure to 600±50 and 700 ppm CO2 compared to ambient 360 and 350 ppm CO2. The results agree with chickpea (increment up to 70%) at 550 ppm compared to 370 ppm CO2 (Madan Pal and Sangeeta 2009), mung bean (increment up to 11.7%) at 550 ±60 ppm (Hao et al. 2011), black gram (increment up to 78 and 30%, at 700 and 550 ppm CO2, respectively) (Sathish et al. 2014). ECO2 is known to ameliorate the negative effects of several abiotic stresses. The water content of this soil was found to be higher under ECO2 aiding in delaying the onset of drought (De Kauwe et al. 2021). ECO2 mitigated the negative impact of high-temperature stress on wheat photosynthesis and biomass (Chavan et al. 2019). ECO2 stimulated photosynthetic responses by modulating electron transport rate, and Rubisco activity and by maintaining the quantum efficiency of PSII under various abiotic stresses (Shanmugam et al. 2013; Javaid et al. 2022). It can be said that ECO2 regulated PSII quantum efficiency and electron transport rate under varying PPFD to sustain the photosynthetic efficiency as observed in the current study.
In addition, a reduction in photosynthesis under limiting PPFD could be linked with reduced the stomatal conductance. Reduced gs rate is one of the common (but not universal) responses when exposed to elevated carbon dioxide (Xu et al. 2016). Cloudy day conditions resulted in closed stomata even though the plants were exposed to ECO2 (Urban et al. 2014). In yet another research result, variation in blue light incidence significantly impacted the gs and E rate. The reduction of blue light triggered a drastic reduction in gs by 43.2% and by 40.0% in E. It can be inferred that varied PPFD, quality of light and ECO2 exerted negative effects in the form of reduced stomatal conductance. The stomatal aperture consumes a significant amount of energy (Lu et al. 1997) and it seems that irrespective of the CO2 concentration, limited radiation energy prevailing under cloudy conditions couldn’t provide the required light use efficiency to regulate the stomatal aperture (Lu et al. 1997; Urban et al. 2014). Along with varying PPFD, rising carbon dioxide concentration, as a consequence of climate change have markedly affected phenotypic and molecular responses of stomata (Xu et al. 2016). Stomatal closure is one of the primal responses under prolonged ECO2 environments (Bunce 2004, 2021; Engineer et al. 2016), however, Xu et al. (2016) and Jordan (2011) opined that this is a general rather than a universal response. The closure of stomata as observed in this study corresponded with above mentioned results. The closure of stomata was mainly attributed to the reduced number of stomata per unit leaf area. Stomata closure mediated optimized water use efficiency under ECO2 (Engineer et al. 2016; Ravi et al. 2022) could be beneficial to maintain photosynthetic efficiency and productivity in the anticipated climatic variability (Sreeharsha et al. 2015). However, stomatal closure with reduced stomata numbers is anticipated to increase the temperature load in the view of reduced evapo-transpiration cooling ability, especially under locations with frequent drought episodes (Long et al. 2004; Leakey et al. 2009; Long and Ort 2010). Stomata tend to open when the intracellular carbon dioxide concentration is low (Outlaw Jr. 2003) whereas, stomata remain closed to maintain the mesophyll conductance for CO2 and leaf water demand (Lawson et al. 2014). In addition, stomatal closure is associated with enriched CO2 concentration-modified extracellular malate in the guard cell (Hedrich and Marten 1993; Hedrich et al. 1994). Malate functions as a CO2 sensor in the guard cell facilitating mesophyll photosynthesis and guard cell function. Plants lacking ABC transporter AtABCB14 exhibited a greater quantum of stomatal closure. ABC transporter facilitated malate accumulation in guard cells and played an important role in stomatal regulation (Lee et al. 2008). Furthermore, molecular studies using Arabidopsis thaliana have revealed that stomatal closure at high CO2 is mediated by carbonic anhydrases (CAs) localized in the plasma membrane of guard cells (Fabre et al. 2007; Hu et al. 2010; Kim et al. 2010). Diminished stomatal regulation responses under ECO2 were further also ascribed to the pH value of guard cells, reduced K+ ion, cytosolic Ca2+ and Cl- ion content and depolarization of guard cells. As per the research results of Fujita et al. (2013), mesophyll is central to stomatal closure via S-type anion channel activation and abscisic acid (ABA) accumulation. Moreover, stomatal closure. In the current study, irrespective of the PPFD, exposure of yam beal leaves to ECO2 sustained the transpiration rate even though stomatal conductance was reduced. Several researchers have reported steady (Ceulemans et al. 1995; Poole et al. 2000) or increased (Atkinson et al. 1997; Lawson et al. 2014) stomatal density or gs rate under ECO2. Stomatal conductance and transpiration rate increased slightly in Quercus leaves exposed to ECO2 (700 ppm) in comparison to ambient CO2 (350 ppm) (Atkinson et al. 1997). The increment in transpiration rate under ECO2 is attributed to varied patterns of epidermal cell differentiation and stable Rubisco content (Lawson et al. 2002). Increased or steady transpiration rate is contradictory to the findings of previous research reports. Teng et al. (2006) found a significant reduction in the transpiration rate of A. thaliana as the stomata density was decreased by 19 and 14% on the adaxial and abaxial surfaces, respectively, in the leaves exposed to 700 ppm CO2 in comparison to ambient CO2 concentration. The transpiration process was substantially diminished in the wheat leaves under ECO2 (700 ppm) in comparison to 390 ppm (ambient) (Houshmandfar et al. 2015).
Increased Ci content under ECO2 as observed in the current experiment was also observed as reported by Fernández et al. (2002), Gleadow et al. (2009), Rosenthal et al. (2012) and Cruz et al. (2014) in cassava; Ravi et al. (2017) in sweet potato; Ravi et al. (2018) in elephant foot yam; Ravi et al. (2019) in taro and Ravi et al. (2020) in yam bean and Ravi et al. (2022) in yams. The Ci is increased at elevated CO2 concentrations (Hao et al. 2011). A Similar increment in Ci concentration in response to short-term exposure to CO2 between 400 and 1000 ppm at saturation PPFD of 1500 μmol m-2 s-1 occur in leaves of other tuber crops: sweet potato (Ravi et al. 2017), elephant foot yam (Ravi et al. 2018) and taro (Ravi et al. 2019). The Pn rate increased as Ci increased between 100 and 1000 ppm CO2. Under elevated CO2 the increase in Pn was limited by the maximal carboxylation rate of Rubisco at lower Ci (< 400 ppm CO2) and by regeneration of the primary substrate RuBP and triose phosphate utilization at higher Ci (600-1000 ppm CO2) (Long and Bernacchi 2003; Kromdijk and Long 2016). Although the Pn rate increased with short-term exposure to exogenous CO2 between 400 and 1000 ppm, the decline in the increment of the Pn rate resulted in an increase in Ci concentration at CO2 concentration between 400 and 1000 ppm. Sweet potato plants grown under controlled conditions had greater photosynthetic rates due to an increase in Ci from 250 to 560 ppm (Cen and Sage 2005). Increased Ci was responsible for increased Pn rates as CO2 concentration increased between 400 and 1000 ppm which agrees with Hikosaka et al. (2005) and Ravi et al. (2017, 2018, 2019). To our knowledge, this is the first study of yam bean depicting the suitability of yam bean for a high CO2 environment under global dimming, and the results will help in developing crop simulation models and for genetic improvement of this crop to high CO2.
In the current study, photosynthetic efficiency across PPFD was substantially improvised under ECO2 (600-1000 ppm) in comparison to 400 ppm. It implies that ECO2 triggered the required mechanism to sustain the photosynthetic process even under low PPFD. As per the research results of Urban et al. (2014), ECO2 (700 ppm) stimulated the net photosynthetic rate in Fagus sylvatica at irradiances above550 µmol m-2 s-1 in comparison to 385 ppm. ECO2 also regulated the sum of daytime-fixed CO2 and light use efficiency under contrasting sky conditions. However, they assumed that the constant prevalence of cloudy conditions as a manifestation of climate change may offset the positive effect of ECO2 on photosynthetic efficiency. The underlying mechanism for deciphering the positive effect of ECO2 on physiological processes under contrasting sky conditions remains unclear and warrants urgent investigation. Unlike other legume crops, yam bean is a unique crop, the only tuber-bearing legume crop where the sink is flower as well as a tuber. Source and sink grow simultaneously. In this regard results reported here are of utmost importance in connection with future anticipated extreme climatic conditions and the potential role of yam bean in supporting food and nutrition security.