Morpho-Anatomical, Physiological and Biochemical Adjustments in Response to Heat and Drought Co-Stress in Winter Barley

Abstract

This study aimed to investigate the combined effect of high temperatures 10 °C above the optimum and water withholding during microgametogenesis on vegetative processes and determine the response of winter barley genotypes with contrasting tolerance. For this purpose, two barley varieties were analyzed to compare the effect of heat and drought co-stress on their phenology, morpho-anatomy, physiological and biochemical responses and yield constituents. Genotypic variation was observed in response to heat and drought co-stress, which was attributed to differences in anatomy, ultrastructure and physiological and metabolic processes. The co-stress-induced reduction in relative water content, total soluble protein and carbohydrate contents, photosynthetic pigment contents and photosynthetic efficiency of the sensitive Spinner variety was significantly greater than the tolerant Lambada genotype. Based on these observations, it has been concluded that the heat-and-drought stress-tolerance of the Lambada variety is related to the lower initial chlorophyll content of the leaves, the relative resistance of photosynthetic pigments towards stress-triggered degradation, retained photosynthetic parameters and better-preserved leaf ultrastructure. Understanding the key factors underlying heat and drought co-stress tolerance in barley may enable breeders to create barley varieties with improved yield stability under a changing climate.

Keywords: microgametogenesis; phenology; photosynthesis; photosynthetic pigments; ultrastructure; yield.

Figure 1

Control and heat and drought co-stressed (HD) barley plants at anthesis. (a) Control Lambada; (b) HD Lambada; (c) control Spinner; (d) HD Spinner (representative images). Arrow—yellowing, senescent leaf. Scale bar = 10 cm.

Figure 2

Yield components (grain number to productive tiller ratio (a); thousand-grain weight (b); grain production (c); harvest index (d) of control and HD Lambada and Spinner plants. TGW—thousand-grain weight. Mean values, along with standard deviations, are presented. In each histogram, different letters above columns indicate significant differences between means at the p ≤ 0.05 level of probability.

Figure 3

Relative water content (a) and osmotic adjustment (b) of control and HD barley flag leaves. OA—osmotic adjustment, RWC—relative water content. Mean values, along with standard deviations, are presented. In each histogram, different letters within columns indicate significant differences between means minimum at the p ≤ 0.05 level of probability.

Figure 4

Total soluble protein content (a), photosynthetic pigment contents (b–e), chlorophyll a to chlorophyll b ratio (f), and carotenoids to chlorophyll a + b ratio (g) in the flag leaves of the control and HD Lambada and Spinner plants. Chl—chlorophyll, DW—dry weight. Mean values, along with standard deviations, are presented. In each histogram, different letters within columns indicate significant differences between means at the p ≤ 0.05 level of probability.

Figure 5

Maximum quantum yield of the photosystem II (a), relative electron transport rate at PSII (b), actual quantum yield of PS II (c), photochemical quenching coefficient (d) and non-photochemical quenching (e) measured 1 day after the mid-uninucleate stage of microspore development, at anthesis and 5 days after anthesis in barley. C—control, L—Lambada, R5—5th day of regeneration, S—Spinner, S1—1st day of HD co-stress treatment, S5—5th day of HD co-stress treatment. Mean values, along with standard deviations, are presented. In each histogram, different letters above columns indicate significant differences between means at the p ≤ 0.05 level of probability.

Figure 6

Measured time series of net photosynthesis (a), stomatal conductance (b), intracellular CO₂ concentration (c), transpiration rate (d), water use efficiency (e) and instantaneous carboxylation efficiency (f) during the 5-day-long HD co-stress (S1–S5) and regeneration (R1–R5) periods in barley genotypes with different stress-susceptibility. C_i—internal CO₂ concentration in the sub-stomatal chamber, E—transpiration rate, WUE—water use efficiency, F_v/F_m—maximum quantum efficiency, g_s—stomatal conductance, P_n—CO₂ assimilation rate. Mean values, along with standard deviations, are presented. In each histogram, different letters above columns indicate significant differences between means at the p ≤ 0.05 level of probability.

Figure 7

Accumulation of proline (a), glycine betaine (b), total soluble carbohydrates (c) and starch (d) in the flag leaves of the control and HD Lambada and Spinner plants. GB—glycine betaine, TSC—total soluble carbohydrates. Mean values, along with standard deviations, are presented. In each histogram, different letters above columns indicate significant difference of means minimum at the p ≤ 0.05 level of probability.

Figure 8

Structure of the cross-sectioned flag leaves (a–d) and starch distribution in the chloroplasts of the mesophyll cells (e–h) of the control (a,b,e,f) and heat and drought co-stressed (c,d,g,h) Lambada (a,c,e,g) and Spinner (b,d,f,h) plants (representative images). abe—abaxial epidermis, ade—adaxial epidermis, bc—bulliform cell, mc—mesophyll cell, sc—substomatal cavity, st—stoma, vb—vascular bundle, arrowhead—chloroplasts with primary starch granules, blue coloration—proteins, red coloration—carbohydrates. Scale bars: (a–d) = 100 µm; (e–h) = 30 µm.

Figure 9

Ultrastructure of control (a,b) and heat and drought co-stressed (c,d) mesophyll cells of Lambada (a,c) and Spinner (b,d) flag leaves. c—chloroplast, m—mitochondrion, p—peroxisome, s—starch, v—vacuole. Scale bar = 1 µm.