Ambient temperatures are increasing at a considerable rate as part of the current climate change, and agriculture has been impacted through losses in crop yields that compromise food security (Nadeem et al. 2018; Carmody et al. 2020). Our results demonstrate that soybean development was affected by heat stress at an average temperature of 40 ºC throughout the soybean cycle.
Temperatures are categorized as high for soybeans when they are above 27°C in the long term, both in the vegetative and reproductive phases (Kai and Iba 2014). In high temperature situations the photosynthetic process is impaired due to the activity of oxidative stress. Severe heat stress induces programmed cell death by enzymatic denaturation (Hassan et al. 2020).
The increase in temperature directly affected the photosynthetic metabolism, however, under the application of the biostimulant, especially at the 1 L ha-1 dose, there was mitigation of the harmful effect of stress, which may be related to the action of A. nodosum extract in soybean. Algae extract promotes an increase in the absorption and assimilation of nutrients, with emphasis on nitrogen resulting from increased activity of enzymes such as nitrate reductase and glutamine synthetase (Fan et al. 2013). It can also increase up to 30.5 and 20% the concentration of nutrients such as phosphorus and potassium, respectively, in plant tissue (Di Stasio et al. 2017). In this way, the organic composition of biostimulants contributes to improving physiological processes, inducing tolerance to abiotic stress, and improving production quality (Di Stasio et al. 2017; Langowski et al. 2019). The reduction of A in the other treatments evaluated may be due to the photoprotection mechanism, since, in Arabidopsis model plants, a reduction in the expression of the AtRBCS1A and AtRCA genes, responsible for catabolism and activation of the RubisCO enzyme complex, was observed (Santaniello et al. 2017), which does not favor the assimilation of CO2, evidenced by the high Ci in these treatments.
Higher gs and E in plants under application of 1 L ha-1 of the biostimulant demonstrate a positive effect. The increase in gs favored A and CE. In addition, greater E provided a reduction in LT, indicating photochemical efficiency of the photosynthetic apparatus even under heat stress, corroborated by the low Ci (Martynenko et al. 2016; Santaniello et al. 2017; Ergo et al. 2018). In fact, biostimulants induce photoprotective plant defence systems under periods of drought stress (Goñi et al. 2018). However, there was no influence of the biostimulant added to the heat stress in the WUE, indicating that in these conditions, soybean maintained the metabolic processes essential for its development (Flexas et al. 2016).
In fact, the application of seaweed extract contributed to attenuating the effect of stress due to the maintenance of the gene expression of RBCS1A related to photosynthesis (At1g67090) and RCA (At2g39730) and PIP1; 2 (At2g45960) and βCA1 (At3g01500) that are involved in controlling the diffusion of CO2 in the mesophile in Arabidopsis thaliana (De Saeger et al. 2020).
Under conditions of heat stress, the high temperature can decrease the internal concentration of CO2, causing the blocking of enzymes of photosynthetic activity and ATP synthesis (Iqbal et al. 2019). However, our results demonstrate that differences in photosynthetic efficiency between the evaluated doses are likely to be large enough to be the main factor related to increases in P.
Among the harmful effects of heat stress, the reduction in membrane permeability is noteworthy, as it promotes drastic changes in the process of photosynthesis and respiration, which come from the increase in the energetic kinetics of the molecules, resulting in the denaturation of proteins, porosity of the cell membrane and increased fluidity (Jedmowski et al. 2015; Hassan et al. 2020).
The relative chlorophyll content of plants under heat stress did not differ from control plants. This maintenance of chlorophyll concentration may be related to the plant's self-protection system, as stress due to high temperatures induces the reduction of chlorophyll biosynthesis, which may have the effect combined with the degradation of chlorophyll molecules (Vass 2012; Fahad et al. 2017), however chlorophylls perish have not been influenced by the factors studied in this research, unlike the antioxidant enzymes SOD and CAT.
SOD activity increased due to the dose of the biostimulant, enabling the attenuating effect of heat stress, as it is the first enzyme to act in the defense of the antioxidant system. SOD acts in the dismutation of the superoxide anion to form H2O2, thus minimizing the damage caused by ROS through the breakdown of superoxide radicals that are generated by oxidative stress (Shafi et al. 2015; Caverzan et al. 2016).
CAT also showed an increase in activity due to the application of biostimulant and acts by removing H2O2, reducing it to two H2O molecules (Barbosa et al. 2014; Caverzan et al. 2016). According to the study by Chakraborty and Pradhan (2011), enzymes such as CAT, APX and SOD increased activity up to a temperature of 50 ºC. However, the activity of APX, POD, RN and Pro concentration were not influenced by the factors studied in this research, indicating that SOD and CAT were the main contributors in the elimination of ROS in soybean, which favored the maintenance of soybean development. In fact, SOD and CAT are directly related to productivity gains (Fig. 6).
The application of the biostimulant promoted an increase in HP, SD and LA, especially under the dose of 1 L ha-1, which may be related to the greater availability of molecules and nutrients from the algae extract. The larger the leaf area, the greater the photosynthetic area of the plant, with a direct influence on crop yield (Devi et al. 2015). Another factor that corroborates the stress-reducing effect was the increase in NNR, SDW and RDW.
Rosa et al. (2021) observed that the biostimulant based on A. nodosum (L.) seaweed extract and fulvic acids induced soybean plants better recovery after water deficit by providing faster reestablishment of cellular water potential, osmotic adjustment, increased stomatal conductance, photosynthetic activity and production of photoassimilates, higher efficiency in energy dissipating mechanisms, which reduced ROS generation, and higher activity of antioxidant enzymes. Similar results were obtained in this research on heat stress in soybean.
The seaweed A. nodosum has in its composition hormones such as auxins, cytokinins, gibberellins and abscisic acid that contribute to plant growth and development and adaptation to stress (Mori et al. 2017, Rouphael and Colla 2018, Ghaderiardakani et al. 2019, De Saeger et al. 2020). Studies suggest an effective participation in the regulation of homones using biostimulants based on algae extract, such as regulation in the promoters of auxin activity (AuxRE), activation of the cytokinin promoter (ARR5) and modulation of the response of the GA24 genes, GASA4, GASA1 of gibberellins (Wally et al. 2013; Goñi et al. 2016).
Our results suggest that increases in PP, G2, G3 and P under the dose of 1 L ha-1 are related to the attenuating effect of the biostimulant during the period of heat stress due to the maintenance of photosynthetic metabolism and antioxidant defense that allowed continuity of soybean development under stressful conditions.
As previously observed, the production components were positively influenced by most of the variables studied (Fig. 6). Thus, there are indications that the increase in LA provided greater surface for absorption of radiation necessary for CO2 assimilation, which was reflected in gains in dry matter mass in soybean plants and energy required for pod formation, mainly of 3 grains that directly influenced the return of culture. However, negative correlation between yield parameters and LT demonstrates the harmful effect of heat stress on soybean production.
Activity of proteins and molecules that respond to stress, directing less energy to defense mechanisms and increasing productive components under biostimulant action (Zang et al. 2010; Tandon and Dubey 2015; Hamed et al. 2018; Mukherjee and Patel 2020). All these ways of inducing different mechanisms promoted the process of adaptation to stress faster, which contributed to the gain in productivity even under conditions of thermal stress.
Synergistic effect of A. nodosum based biostimulants as mitigator of abiotic stress has been reported in the literature on heat stress in tomato (Carmody et al. 2020) and water deficit in soybean (Rosa et al. 2021). In fact, biostimulants with A. nodosum stimulate plant growth and adaptation to stress (Rouphael et al. 2017, De Saeger et al. 2020).
This research was focused on challenging soybean plants to several degrees of temperatures above their ideal conditions for development, i.e., 27°C (Kai and Iba 2014). Unlike thermal shocks applied for a short period of time (hours), this experimental design was more representative of naturally occurring stress conditions in the field, as moderate heat stress regimes can affect the function of vegetative tissues and further impair production parameters. Given the gap on the effect of this biostimulant on high temperatures in soybeans and its effects on crop yield, this research guides the search for a viable solution, creating more sustainable and environmentally acceptable agricultural practices.