Physiological and biochemical responses of hybrid maize (Zea mays L.) varieties grown under heat stress conditions

Experimental site and climate characteristics

The field experiments were conducted at the Talat Demiroren research station of the South Eastern Anatolian Project Agricultural Research Institute, Sanliurfa Province, Turkey (36° 54′ 13″ N, 38° 55′ 03″E, altitude 378 m). The field studies were carried out in the 2020 and 2021 corn growing periods. The soil of the experimental field had a heavy clay textured, granular structure and the infiltration rate was within the optimum range. The organic matter content and pH levels were between 0.81% and 1.11% and 7.55, and 8.11, respectively.

The study area is located in a semi-arid climate with high temperatures and low relative humidity during the summer season. The corn growing periods in both 2020 and 2021 were drier and hotter compared to the historical temperature and precipitation averages. The amount of precipitation and duration of sunshine during the experiments verified that the region has a semi-arid climate close to the aridity line. The climate data of the study area recorded during the corn growing seasons of 2020 and 2021 are provided in Table 1.

‘HS’ conditions were present between May 1 and September 1, while August 1 to December 1 had ‘C’ conditions, which are the optimum conditions. The ‘HS’ and ‘C’ conditions in the study area were determined using historical climate data. The temperature difference between the ‘HS’ and ‘C’ conditions was 4.77 °C in the first year of the study (2020), and increased to 6.65 °C in the second year (2021). Temperature differences were recorded both between years and corn growing conditions (GC). More extreme climate conditions occurred in the first year of the experiment.

The average relative humidity also differed between the experimental years and the ‘HS’ and ‘C’ conditions. In the first year of the study, the relative humidity during ‘HS’ conditions was 13.7% lower than under ‘C’ conditions, but 12.7% lower in the second year. The mean relative humidity values during ‘HS’ conditions in both experimental years were below 40%. Similar to the temperature and relative humidity values, the daily average maximum temperature values between the tasseling and silking periods during ‘HS’ conditions were higher than those recorded in ‘C’ conditions of both experimental years. The temperature exceeded 40 °C on some days during the 2021 growing season, especially under ‘HS’ conditions (Fig. 1).

Plant material, experimental design, and cultural practices

Eleven candidate hybrid corn varieties from the national corn breeding program and two commercial hybrid corn varieties, used as controls, were used in this study. All included varieties were classified as having moderate maturity (FAO 650) and are widely grown in ‘HS’ areas and regions of the world. Some traits of the corn varieties used in the study are given in Table 2.

This experiment used split plots in randomized blocks with three replications, with ‘HS’ and ‘C’ growing conditions applied to the main plots and the different corn varieties planted in the sub plots. The inter row and intra row spacings were 70 and 17 cm, respectively. Each plot had four rows, 5 m in length, and the area of each plot was 14 m². Each plot had 115 to 120 corn plants. Fertilizer was applied in the seedbeds at a rate of 350 kg ha⁻¹ in the form of diammonium phosphate (18%N-46%P₂O₅-0%K₂O) during sowing (Kucuk, Cevheri & Mutlu, 2019). Nitrogen (350 kg N ha⁻¹) was applied as urea (46% N) at the second and fourth weeks of sowing. Considering climate conditions and the water requirements of corn plants, a total of 660 and 600 mm irrigation water was applied in the ‘HS’ and ‘C’ conditions of the first year, respectively, and 710 and 630 mm irrigation water was applied in ‘HS’ and ‘C’ conditions of the second year, respectively, when the ‘HS’ was more serious. Corn cobs were harvested between August 25–29 and December 1–3 in the ‘HS’ and ‘C’ conditions of the first year, respectively, and between August 24–25 and November 28–30 in the second year. The harvest dates differed between the years due to changes in climate, and the cobs were harvested when the corn kernels had 24–25% moisture.

Data collection

Data analysis

A statistical analysis was carried out using JUMP (Version 13.2.0) statistical software. The differences in the traits between the growing conditions were tested using an analysis of variance (ANOVA). A statistical analysis was carried out for each experimental year, separately, as well as the 2-year combined data. The degrees of freedom and F ratio for all parameters were calculated in the combined-year data. The least significant difference (LSD) multiple comparison test, set at a 95% probability, was used where ANOVA indicated significant differences between the mean values for year, the growing conditions, and the plant varieties. The relationships between all traits examined in the study were determined using the Spearman’s correlation test.

Correlations

The lowest and highest correlation coefficients and significance levels of the traits are shown in Table 7. Significant (P ≤ 0.01) positive correlations were recorded between GY and TPC (r = 0.83), LWP (r = 0.78), Chl-a (r = 0.71), DT (r = 0.54), LWP and TPC (r = 0.77), Chl-a (r = 0.77), DT (r = 0.65), DT and Chl-a (r = 0.57), TPC (r = 0.53), and Chl-a and TPC (r = 0.67). Significant (P ≤ 0.01) negative correlations were observed between CMD and GY (r = −0.72), LWP (r = −0.71), TPC (r = −0.71), Chl-a (r = −0.64), and DT (r = −0.43). Damage to cell membranes caused a decrease in GY, TPC, and other physiological parameters. The lowest correlation coefficient was obtained between CMD and DT parameters, while the highest correlation coefficients were recorded between GY and TPC and LWP.

Table 7:

Correlation analysis between the traits investigated in the study.

High correlation coefficients and significant relationships were observed between grain yield and several physiological and biochemical parameters.

Traits	By traits	Correlation	Count	Lower 95%	Upper 95%	Signif prob (P)	Correlation levels
Chl-a	TPC	0.6697	156	0.5728	0.7481	<0.0001**
DT	TPC	0.5302	156	0.4069	0.6344	<0.0001**
DT	Chl-a	0.5712	156	0.4549	0.6683	<0.0001**
CMD	TPC	−0.7064	156	−0.7773	−0.6178	<0.0001**
CMD	Chl-a	−0.6383	156	−0.7229	−0.5348	<0.0001*
CMD	DT	−0.4254	156	−0.5460	−0.2875	<0.0001**
LWP	TPC	0.7723	156	0.7001	0.8288	<0.0001**
LWP	Chl-a	0.6768	156	0.5815	0.7537	<0.0001**
LWP	DT	0.6498	156	0.5487	0.7322	<0.0001**
LWP	CMD	−0.7132	156	−0.7826	−0.6262	<0.0001**
GY	TPC	0.8263	156	0.7690	0.8704	<0.0001**
GY	Chl-a	0.7120	156	0.6247	0.7816	<0.0001**
GY	DT	0.5441	156	0.4232	0.6460	<0.0001**
GY	CMD	−0.7163	156	−0.7850	−0.6300	<0.0001**
GY	LWP	0.7794	156	0.7092	0.8344	<0.0001**

DOI: 10.7717/peerj.14141/table-7

Notes:

* Significant at 0.05 level of probability.

** Significant at 0.01 level of probability.

LWP, Leaf water potential (%); GY, Grain yield (kg ha⁻¹); Chl-a: Chlorophyll-a (mg g⁻¹); DT, Days to 50% tassel (day); CMD, Cell membrane damage (%); TPC, Total phenol content (μg g⁻¹).

Days to 50% tassel (day) and grain yield (kg ha⁻¹)

The decrease in DT values during ‘HS’ may be attributed to the way the plants responded to the adverse environmental conditions; in order to produce grain under such harsh conditions, the plants may have accelerated their life cycles and tasseling stages (Wang et al., 2022). The disruptions to protein and chlorophyll synthesis, insufficient gas exchange through stomatal openings, and water loss through excessive leaf transpiration likely triggered early flowering (tasseling). Early flowering in the maize plants caused a decrease in other physiological parameters, especially GY. The stressed plants could not sufficiently complete their vegetative and morphological processes and experienced some problems at cellular level. Moderate correlations were seen between DT and other physiological parameters, especially GY. Similarly, Cao et al. (2008) observed that early flowering accelerated the tasseling process in maize when the temperature was above 35 °C. The male and female organs of the plants are damaged under ‘HS’ conditions. In addition, pollen germination, pollen tube growth, and viability of the ovules are inhibited, sterility is significantly increased, and grain yield is decreased with the increased ‘HS’ conditions (Maheswari et al., 2012). Lobell et al. (2011) found that a slight increase in temperature during flowering may cause a significant decrease in GY. In ‘HS’ conditions, the pollen fertility of maize plants likely decreases the tasseling period, which is the most sensitive period, disrupting the synchronization between tasseling and silking, resulting in decreased GY.

Previous studies have shown that both environmental conditions and the genetic characteristics of cultivars impact GY (Gao et al., 2021; Liu et al., 2021; Han et al., 2022). Environmental factors, such as high temperature and low relative humidity, adversely affect plant growth leading to yield losses because of the amount of water lost through evapotranspiration. Pollen viability decreases when daily temperatures are 40 °C and above during the reproductive stage (between tasseling and silking), decreasing GY. A correlation analysis indicated that an increase in physiological parameters such as LWP and Chl-a also increased GY, while an increase in CMD decreased GY. ‘HS’ conditions caused damage at the cellular level, deteriorated protein synthesis, disrupted photosynthesis and respiration, and decreased the carbohydrate reserves of the plants, chlorophyll amounts, and the rate of photosynthesis. Because of these changes, the plants could not grow properly, and GY significantly decreased. Yang et al. (2019) observed similar adverse effects on the protein and starch metabolism in leaves under serious ‘HS’ conditions leading to decreases in yield and quality. These GY decreases due to ‘HS’ conditions are also in line with the findings of Cairns et al. (2013). Similarly, Yang et al. (2020) reported a significant decrease in dry matter accumulation, tasseling day, chlorophyll content, and GY in maize under ‘HS’ conditions. The GY loss (22.9%) reported by Yang et al. (2020) was close to the GY loss recorded in this study (15%). High temperatures cause the denaturation of the chlorophyll sub-pigments involved in photosynthesis (Waraich et al., 2012; Mutlu, 2021) and deteriorate other major physiological processes (Mangani et al., 2019), shortening the grain filling time, and reducing the GY. In parallel with these findings, Siebers et al. (2017) found that temperatures over 35 °C inhibited ovary fertilization and the grain filling process of corn plants.

Leaf water potential (%) and Chlorophyll-a (mg g⁻¹)

The leaves are the plant organelles most affected by stress conditions (Killi et al., 2017), and they provide important information about the level of plant stress (Li et al., 2020). High temperatures cause water loss in the leaves, decrease the fluid of the cytoplasm organelle in plant cells, and decrease the flexibility of the cell membranes (Young, Wilen & Bonham-Smith, 2004; Li & Howell, 2021). The chloroplast organelles are also adversely affected by ‘HS’ conditions (Li & Zhang, 2022). Plant leaves wilt, shrink, and curl inward when the stress level is higher (Butler, Mueller & Huybers, 2018). Despite these protective reflexes, the water carried from the roots cannot compensate for the amount of water lost by the transpiration in the leaves (Zhu et al., 2019). Significant negative correlations were observed between LWP and CMD, while significant positive correlations were recorded between the remaining parameters. Flexibility of the cell membranes decreases under ‘HS’ conditions. An increase in ‘HS’ increases cell damage and electrolyte leakage, adversely affecting LWP. The water in the leaf, which is important to both food production and photosynthesis, also indirectly affects the GY of corn varieties. Alam et al. (2017) also found that ‘HS’ decreased leaf water content and increased leaf senescence, resulting in significant maize yield loss.

Chl-a pigments are also adversely affected by stress and undergo structural deterioration (Karim, Fracheboud & Stamp, 1999; Mohammed & Tarpley, 2010). Therefore, in addition to the defects in the photosynthesis mechanism seen with stress, leaves cannot sufficiently absorb the light energy to reduce carbon dioxide, sugar, and other plant substances. Sensitive corn varieties cannot tolerate high ‘HS.’ The formation of reactive oxygen species such as O₂, OH and H₂O₂ in the cells of sensitive corn varieties causes damage to organelles such as chloroplasts and mitochondria in the plant cells (Guo, Zhou & Zhang, 2006). In parallel with these findings, many researchers have reported less photosynthesis and less assimilate accumulation in the leaves, shortened grain filling periods, and decreased corn grain yield due to dry matter loss under ‘HS’ conditions (Yan et al., 2017; Baiyeri et al., 2019). Low chlorophyll sub-pigments such as chlorophyll-a and b formation under ‘HS’ conditions also lowers GY (Yamori, Hikosaka & Way, 2014). Consistent with this study, Marchand et al. (2005) reported that the activity of photosystem II was significantly reduced or stopped under ‘HS,’ which disrupted the functioning of photosynthetic pigments such as chl-a and b.

Cell membrane damage (%) and total phenol content (µg g⁻¹)

The CMD is the most practical and accurate indicator of the plant’s stress level at the cellular level. When the temperature rises significantly, reactive oxygen species accumulate in the cells of sensitive corn varieties, and these harmful oxygen species cause peroxidation in the membranes. A significant negative correlation was recorded between CMD and other parameters. Without water, the membranes lose flexibility and are damaged over time. Zhang et al. (2005) also reported that ‘HS’ increased membrane permeability and damaged mesophyll cells. In addition, decreases in other physiological and morphological characteristics, especially GY, were reported as the damage rate of membranes increased (Zhang et al., 2005). In contrast, resistant corn varieties have tolerance mechanisms such as the ability to make ionic and osmotic adjustments, membrane permeability adjustments, and these varieties can even protect and repair the proteins and cell membranes damaged by ‘HS’ (Vinocur & Altman, 2005).

The TPC accumulates more in plant tissues up to a certain stress level in ‘HS’ compared to ‘C’ conditions. The photosynthesis and gas exchange in the leaves were disrupted during ‘HS’ conditions due to a decrease in Chl-a pigment. The phenolic sub-pigments deteriorated and decreased accordingly. Environmental factors and the characteristics of the corn variety both significantly affect TPC. Intense ‘HS’ may adversely affect the structure of enzymes such as antioxidants and phenols, leaving plants vulnerable (Yamakawa et al., 2007). Consistent with these findings, Shahzad et al. (2011) reported that the total phenol content of corn plants decreased under high-temperature conditions.