2.1 The Impact of HS on the Aerial Parts of Rice

Under HS, inhibition of photosynthesis is one of the primary factors limiting crop growth (Garcia et al. 2023). HS (42°C, 14 h) leads to the collapse of thylakoid structures and damage to stromal lamellae, resulting in significant chloroplast structural damage (Figure 1A) (Zhang et al. 2022). This structural damage to the thylakoids directly disrupts the light reaction process, weakening interactions between lipid molecules and electron carrier proteins (Prasertthai et al. 2022). Consequently, this leads to the degradation of core subunits of Photosystem II (PSII), such as the D1 protein, thereby reducing PSII's maximum photochemical efficiency, maximum quantum yield, and steady-state quantum yield (Prasertthai et al. 2022). Simultaneously, HS (45°C, 6 h) reduces the effectiveness of photoprotection mechanisms, as evidenced by a weakened non-photochemical quenching capacity and a reduced ability to dissipate excess light energy (Prerostova et al. 2022). Ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) is a key enzyme in the Calvin cycle that catalyzes the fixation of carbon dioxide (Makino 2021). HS reduces the activity and efficiency of Rubisco, thereby reducing the rate of CO₂ assimilation, which reduced carbon fixation efficiency in the Calvin cycle and ultimately impacts overall photosynthetic productivity (Qu et al. 2021).

The reproductive growth stages of rice, including booting, heading, flowering, and grain filling, are more sensitive to HS than the vegetative growth stages (Arshad et al. 2017). HS primarily reduces spikelet fertility and causes yield loss by affecting pollen viability and longevity (Karwa et al. 2020; Shrestha et al. 2022), pollen tube growth and orientation (Shi et al. 2018), stigma and ovary function (Qi and Wu 2022), flowering time (Liu, Zhou, and Sun 2023a), and carbohydrate metabolism, among other factors (Wang et al. 2019). For example, heat-sensitive rice varieties such as US-312, IET23296, PHB-71, and PR-113 show a 40%–51% reduction in pollen viability under HS (Karwa et al. 2020). Both the heat-tolerant rice variety N22 and the heat-sensitive IR64 experience inhibited pollen tube growth due to HS, preventing effective pollen delivery to the embryo sac and resulting in reductions in spikelet fertility by 12.4% and 64.9%, respectively (Shi et al. 2018). Additionally, HS reduces the activity of the Ghd7 repressor protein, which is involved in the photoperiodic regulation of flowering (Nagalla et al. 2021). This leads to premature flowering, negatively affecting pollination and fertilisation of rice pollen and stigmas (Nagalla et al. 2021). Short-term HS increases the concentration of non-structural carbohydrates (NSCs) in the stem while decreasing NSC concentration in the panicle (Mahmood et al. 2024). The reduced efficiency of NSC transfer from the stem to the panicle, along with abnormal carbon allocation, severely impairs nutrient transport to the reproductive organs (Mahmood et al. 2024).

In addition to affecting rice growth, HS also affects grain quality (Zhang et al. 2023; Zhou, Xia, and He 2020). Starch structure and composition are crucial determinants of grain quality (Zhao et al. 2019). The amylose content typically affects the hardness of the rice grain, while amylopectin influences grain cohesiveness (Zhao et al. 2019). HS during the grain-filling stage causes a sharp decline in amylose content, leading to an increase in chalky grains. This negatively affects the overall milling rate and head rice yield, directly reducing the economic value of the grain (Zhang et al. 2018; Siddik et al. 2019; Nakamura et al. 2021). The Wx gene is primarily responsible for the synthesis of amylose in starch (Zhang, Xu, and Jiang 2021a). High temperatures can suppress Wx gene transcription, thereby impairing starch synthesis (Zhang, Xu, and Jiang 2021a). Elevated temperatures also decrease the proportion of short-chain amylopectin molecules in the grain, while increasing the production of medium-chain and long-chain amylopectin molecules (Zhang et al. 2016). These changes result in higher crystallinity and altered gelatinisation properties (Zhang et al. 2016). Additionally, HS during the flowering and grain-filling stages can affect the accumulation of 2-acetyl-1-pyrroline, a key compound involved in aroma metabolism, resulting in changes in the grain's aromatic characteristics (Mishra et al. 2024).

2.2 The Impact of HS on the Underground Parts of Rice

Compared to the above-ground parts of plants, the response of roots to HS has been less extensively studied, with most existing research focusing on thermomorphogenesis (González-García et al. 2023). Thermomorphogenesis refers to the morphological and developmental changes that plants undergo in response to non-lethal temperature increases, allowing them to adapt to environmental temperature fluctuations (Casal and Balasubramanian 2019). Different species exhibit varying optimal growth temperatures for root structures (Fonseca de Lima et al. 2021). HS induces structural changes in plant roots, including reduced primary root length, decreased lateral root density, altered root growth angles, an increased proportion of fine roots, reduced root hair density and average length, increased membrane permeability and fluidity, and reduced water viscosity (Fan et al. 2022; Luo et al. 2021; Lam et al. 2020). These structural changes severely impair root functions, notably reducing their ability to absorb water and nutrients (Rehman et al. 2019). Furthermore, HS elevates the oxygen demand of plant roots while simultaneously reducing oxygen availability, leading to root hypoxia (Fonseca de Lima et al. 2021).

The growth and development of rice roots are influenced by various environmental factors, with the optimal temperature range for root development being 25°C–28°C (Fonseca de Lima et al. 2021; Sharifi et al. 2018). A temperature increase from 23°C to 27°C enhances the translocation of sodium (Na) and cadmium (Cd) from the rice roots to the above-ground parts, potentially exacerbating Cd toxicity (Feng et al. 2023). When HS (40°C) occurs, excessive Cd accumulation in rice roots inhibits crop growth by disrupting cell structure and reducing nitrogen uptake, which interferes with the nitrogen assimilation process (Munir et al. 2023). HS also negatively affects rice root morphology, including root length, surface area, and dry weight, thereby reducing the contact area between roots and soil. This reduction leads to lower water and nutrient absorption efficiency (Figure 1B) (Huang et al. 2021). Additionally, temperatures above 35°C significantly decrease nitrogen fixation efficiency in rice roots, while at 44°C, nitrogenase activity in the rice root system ceases entirely (Trolldenier 1982). HS at 45°C further disrupts cytokinin synthesis in the root system and its transport to the leaves, compounding the physiological stress on the plant (Prerostova et al. 2022).

3.1 Perception and Signal Transduction: Perceiving High Temperatures and Transmitting the Signal From the External Environment to the Cell Interior

Cell wall-modifying enzymes, such as pectin methylesterases (PMEs), play a crucial role in regulating cell wall plasticity (Pelloux, Rusterucci, and Mellerowicz 2007). During HS, PMEs positively regulate rice thermotolerance by maintaining appropriate Ca²⁺ levels in the intercellular spaces (Figure 2) (Ezquer et al. 2020; Gall et al. 2015). In rice, 43 different types of PMEs have been identified, but their physiological functions remain largely unexplored (Jeong, Nguyen, and Lee 2015). Rhizobia and arbuscular mycorrhizal fungi secrete bioactive substances, such as lipo-chitooligosaccharides (LCOs) and other signalling molecules (Kan, Fang, and Jia 2017; Jalmi and Sinha 2022). These substances induce the expression of plant cell wall-related genes, enhancing plant stress resistance and symbiotic relationships by regulating pectin demethylation (Kan, Fang, and Jia 2017; Jalmi and Sinha 2022). Microorganisms may reshape plant cell walls through various mechanisms, suggesting that cell wall modulation could be a critical strategy for rice to cope with HS (Wu, Bulgakov, and Jinn 2018).

The plasma membrane is the primary structure for heat perception and sensing (Ohama et al. 2017). Cyclic nucleotide-gated ion channels (CNGCs) located on the plasma membrane act as non-selective cation channels, mediating Ca²⁺ signalling during HS (Ohama et al. 2017). In rice, OsCNGC14 and OsCNGC16 are significantly upregulated under HS, and mutants of these genes exhibit increased sensitivity to high temperatures (Figure 2) (Cui, Lu, and Li 2020). Annexins, a family of Ca²⁺-dependent phospholipid-binding proteins, are implicated in the regulation of HS responses in rice (Gerke and Moss 2002; Jami et al. 2012). Additionally, glutamate receptor-like channels on the plasma membrane play a role in heat signal transduction; exogenous application of glutamate enhances heat tolerance in maize seedlings by activating Ca²⁺ signalling pathways (Figure 2) (Li, Ye, and Qiu 2019; Yu et al. 2023). Microorganisms also influence plant Ca²⁺ signalling pathways by secreting metabolites or signalling molecules, thereby regulating various plant signal transduction pathways (Jha and Pandey 2021; Saand et al. 2015). Future research should investigate the impact of rice-microbe interactions on calcium signalling channels under HS to better understand their regulatory roles (Zamioudis and Pieterse 2012).

In addition to Ca²⁺, ROS serve as crucial signalling molecules in plants’ response to abiotic stress (Medina et al. 2021). Under HS, the expression of OsNox5-9 in rice is significantly upregulated, leading to increased ROS production (Wang et al. 2013). High-temperature conditions can trigger microorganisms to help plants maintain low levels of ROS, which function as signalling molecules that interact with calcium signalling pathways to regulate various physiological responses (Ma and Berkowitz 2007). In addition to Ca²⁺ and ROS, other components involved in heat signal transduction have been identified, including the phosphoinositide signalling system (Ren et al. 2017; Epand 2017). Phosphoinositide-specific phospholipase C9 (PLC9) is a key enzyme in the phosphoinositide signalling system (Liu, Liu, and Wang 2020c). Ectopic expression of the Arabidopsis AtPLC9 gene in rice has been shown to enhances the rice's heat tolerance (Liu, Liu, and Wang 2020c).

HSFs and HSPs play a crucial role in activating the HSR and regulating heat tolerance in rice (Hu, Hu, and Han 2009). For instance, in rice, OsHsfA2c and OsHsfB4b interact with the OsClpB-cyt promoter, which encodes the essential Hsp100 protein (Singh et al. 2012). Among the 25 identified HSFs in rice, 19 are positively associated with HS (Jin, Gho, and Jung 2013). Notably, OsHsfA2a exhibits the highest expression levels under HS conditions in rice (Chauhan et al. 2011; Malumpong et al. 2019; Shamshad et al. 2023). Current research indicates that microorganisms can regulate plant HSPs expression directly or indirectly through various mechanisms, thereby influencing plant heat tolerance (Khan, Asaf, and Khan 2020a). However, the specific mechanisms by which microorganisms regulate the expression of HSFs and HSPs in rice remain unclear (Malumpong et al. 2019). Further research is needed to explore the influence of microorganisms on OsHSFA2 and its downstream genes in rice (Wu et al. 2022). In addition to HSFs, other transcription factors (TFs) in rice also regulate downstream genes in response to HS (REN et al. 2021). For example, NAC family members ONAC127 and ONAC129 directly regulate OsMSR2 and OsEATB, while indirectly influencing the expression of OsHSP101, contributing to the HS response (Ren et al. 2021). The MYB family member OsPL, containing HS cis-regulatory elements, enhances rice heat tolerance (Akhter et al. 2019). Similarly, the bZIP family TF OsbZIP74 upregulates OsNTL3 expression in response to HS and endoplasmic reticulum stress (Liu et al. 2020a).

3.2 Strategies to Cope With HS: Maintaining Protein Homeostasis and Alleviating Oxidative Stress

Under HS, high molecular weight HSPs and small HSPs in rice function synergistically to protect the structure and function of intracellular proteins, thereby enhancing rice heat tolerance (Liu, Tseng, and Wu 2023a; Do et al. 2023; Kim and An 2013). During HS, the accumulation of misfolded proteins in the endoplasmic reticulum activates the expression of OsNTL3 and OsbZIP74, improving rice's ability to respond to HS (Figure 2) (Liu et al. 2020a). Proteins such as OsHTAS, OsHIRP1, and TT1, which possess E3 ligase activity, contribute to heat tolerance in rice by facilitating the rapid and efficient removal of damaged proteins (Figure 2) (Liu et al. 2015; Kim, Lim, and Jang 2019; Li et al. 2015). These proteins play a crucial role in maintaining cellular protein homeostasis during HS conditions. Additionally, beneficial microorganisms may aid in maintaining intracellular protein homeostasis by secreting signalling molecules and metabolites, enhancing the plant's antioxidant system, and indirectly modulating the UPR and UPS (Godara and Ramakrishna 2023; Eid, Salim, and Hassan 2019). These processes collectively improve the clearance of misfolded and damaged proteins, further supporting rice under HS (Godara and Ramakrishna 2023; Eid, Salim, and Hassan 2019).

HS not only disrupts protein homeostasis but also leads to a sharp increase in ROS levels within plant cells (YANG, CHEN, and QI 2024a). To mitigate the effects of HS, rice enhances the activity of antioxidant enzyme systems and non-enzymatic antioxidants to eliminate excess ROS (Shekhawat et al. 2022). During HS, OsHSP17.9, OsANN1, HES1, MSD1, and OsNCED1 in rice regulate the expression of superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPX), which directly participate ROS scavenging and oxidative stress response (Qiao et al. 2015; Do et al. 2023; Xia et al. 2022; Shiraya 2015; Zhou et al. 2022). In addition to ROS elimination, the accumulation of osmoprotectants serves as another critical defence mechanism for enhancing rice heat tolerance (Bamagoos, Alharby, and Fahad 2021; Kavi Kishor et al. 2022). For instance, in the rice OsProDH mutant, increased proline levels are associated with improved tolerance under high-temperature conditions (Guo et al. 2020).

3.3 Strategies to Cope With HS: Physiological and Developmental Adjustments in Rice

Stomatal regulation is a crucial mechanism by which plants respond to environmental changes, particularly temperature fluctuations (Wang and Chang 2024; Haworth et al. 2021; Islam et al. 2018). During HS, the opening and closing of stomata affect rice's transpiration rate and CO₂ assimilation capacity (Figure 1D) (Caine et al. 2023). High-yielding rice varieties generally exhibit higher transpiration rates under various weather conditions (Aryan et al. 2022; You et al. 2023; Kondo et al. 2021). Under HS, rice increases stomatal conductance to accelerate the transpiration rate, thereby lowering leaf temperature (Luan and Vico 2021). Remarkably, even at night, rice utilises transpiration cooling regulated by its circadian rhythm to maintain optimal temperature (Zhang, Yang, and Peng 2021b). The overexpression of OsMDHAR4 in rice leads to reduced intracellular H₂O₂ levels, keeping stomata open and resulting in increased water loss (Liu et al. 2018). Conversely, the overexpression of OsHTAS significantly elevates abscisic acid (ABA) levels, which stimulates H₂O₂ production, promotes stomatal closure, reduces water evaporation, and mitigates water stress induced by HS (Liu et al. 2015). Microorganisms can regulate stomatal dynamics by modulating plant hormones levels, such as ABA (Egamberdieva et al. 2017). For example, the beneficial root endophytic fungus Piriformospora indica enhances rice survival under drought stress by promoting stomatal closure and reducing water loss (Tsai et al. 2020). This microbial modulation of stomatal regulation may also aid plants in coping with HS (Matkowski and Daszkowska-Golec 2023). However, further research is needed to elucidate the mechanisms by which microorganisms affect stomatal regulation in rice under HS and how these interactions contribute to HS alleviation (Matkowski and Daszkowska-Golec 2023).

Stomatal regulation under HS often reduces photosynthetic efficiency, compelling rice to adjust its metabolic strategies to optimise resource allocation (Li et al. 2020). Adapting to high temperatures demands substantial energy and metabolic resources (Bamagoos, Alharby, and Fahad 2021; Hou et al. 2021; Suprasanna, Nikalje, and Rai 2016). For example, rice accelerates the grain-filling rate under HS to complete its growth cycle more rapidly, thereby shortening the effective grain-filling period (Liu et al. 2021). To maintain energy homeostasis, rice modulates hormone levels under HS, enhancing the expression of genes associated with sucrose transport and metabolism, thereby ensuring the availability of essential energy and metabolic intermediates (Rezaul et al. 2019). Microorganisms contribute to the optimisation of plant carbon and nitrogen metabolism through mechanisms such as nitrogen fixation, improving nutrient absorption efficiency, regulating carbon metabolism, secreting plant hormones, and strengthening antioxidant systems (Maitra, Pramanick, and Dey 2021). For example, the nitrogen-fixing strain Azotobacter vinelandii AV7, isolated from the rhizosphere soil of tomatoes and capable of withstanding temperatures up to 50°C, promotes root development and overall plant growth by solubilizing phosphates, producing indole-3-acetic acid (IAA), and generating siderophores (Sev et al. 2020). Microorganisms that enhance plants energy metabolism capacities are vital for maintaining energy homeostasis under HS (Maitra, Pramanick, and Dey 2021).

Plants can also adjust root growth and development to cope with HS (Koevoets, Venema, and Elzenga 2016; Vu, Stuerz, and Asch 2020). In rice, changes in microRNA (miRNA) expression in roots are more pronounced than in shoots under HS, indicating the root system's response mechanisms to HS are more sensitive and active (Mangrauthia et al. 2017). Additionally, at high temperatures, rice roots induce coiling movements by promoting ethylene biosynthesis and signalling, thereby continuously adjusting their growth trajectory (Cai, Dai, and Jin 2024b). Notably, rice roots can independently perceive and respond to temperature changes, enabling them to rapidly adapt to HS, thereby enhancing the plant's overall resilience (Prerostova et al. 2022; Bellstaedt et al. 2019). The role of soil microorganisms in influencing plant responses to HS is also significant and warrants further exploration (Wang et al. 2024). Future efforts could focus on designing synthetic microbial communities to optimise rice root responses to HS. Such approaches hold the potential to simultaneously improve soil quality, increase rice yield, and enhance heat tolerance (Wang et al. 2024).

The molecular mechanisms underlying rice's response to HS suggest that microorganisms may directly or indirectly influence rice's heat tolerance through various pathways (Shekhawat et al. 2022). Under HS, microorganisms may secrete signalling molecules that regulate cell wall methylation and affect the activity of calcium signalling channels, thereby regulating Ca²⁺ concentrations (Wu, Bulgakov, and Jinn 2018; Zamioudis and Pieterse 2012). Additionally, microorganisms can regulate the antioxidant system, influencing the concentration of the second messenger ROS within rice cells, thereby participating in the response to heat signal transduction (Ismail et al. 2021). They also regulate the expression of rice HSPs through various direct or indirect mechanisms (Wu et al. 2022). Furthermore, microorganisms may indirectly regulate rice UPR and UPS, aiding in the maintenance of intracellular protein homeostasis (Godara and Ramakrishna 2023; Eid, Salim, and Hassan 2019). Ongoing exploration and innovation in these research areas hold great promise for advancing microbe-mediated thermotolerance in rice. Such advancements could significantly enhance the sustainability and resilience of agricultural practices in the face of rising global temperatures (D'hondt et al. 2021).

4.1 Microorganisms in HS Environments and Their Interactions With Plants

HS significantly impacts microbial communities in soil and on plant surfaces, typically resulting in reductions in microbial diversity, richness, and biomass, thereby disrupting community stability (Bashir Ahmed Siddique et al. 2023; Guillot et al. 2019; Guo et al. 2024). The natural environment is highly complex and microorganisms sense and respond to environmental temperature changes through various mechanisms (Samtani, Unni, and Khurana 2022). The impact of HS on soil microbial communities depends on factors such as heat intensity, exposure duration, soil type, and the adaptability of the microbial community (Samtani, Unni, and Khurana 2022). For example, under extreme HS (60°C), the relative abundance of α-Proteobacteria and γ-Proteobacteria increases significantly in low-nutrient soils but decreases markedly in high-nutrient soils (Ciss, Tall, and Sall 2023).

As for rhizosphere microorganisms, HS can indirectly increase the spatial heterogeneity of rhizosphere microbial habitats by altering soil conditions and plant growth patterns, which in turn affects the structure of rhizosphere microbial communities and the relative abundance of specific functional groups (Jia et al. 2020; Yu et al. 2024). For instance, studies have shown that HS reduces the levels of available nitrogen (e.g., nitrate nitrogen, ammonium nitrogen) and water-soluble organic carbon (WDOC) in soils (Lin et al. 2023). In response to these changes, microbial communities adapted their metabolic strategies and community structures, with specific genera such as Nitrospira and Penicillium becoming enriched (Lin et al. 2023). These adaptations play a critical role in stabilising microbial biomass carbon (MBC), enabling microbes to accelerate nitrogen cycling and enhance plant nitrogen uptake (Lin et al. 2023). This supports plant growth under HS conditions.

Plants actively recruit beneficial “stress-tolerant microbiomes” to help them cope with HS (Figure 1C) (Trivedi et al. 2020). They achieve this by altering the composition of root exudates, releasing specific signalling molecules, and modulating their immune systems (Chai and Schachtman 2022; Abhilash et al. 2016). For instance, under HS, sorghum selectively recruits heat-tolerant Actinobacteria, which produce various siderophores and other secondary metabolites to improve soil structure, water retention, iron availability, and organic matter decomposition, thereby optimising the rhizosphere environment (Wipf, Bùi, and Coleman-derr 2021). Similarly, during HS, the relative abundance of Basidiomycota in the rhizosphere of summer maize increases significantly, along with notable upregulation of specific metabolic pathways, including valine, leucine, and isoleucine biosynthesis, penicillin and cephalosporin biosynthesis, and ABC transporter pathways in the rhizosphere microbial community (Yuan et al. 2024). Under drought stress, the abundance of Streptomyces increases by 3.4 times in C₄ plants compared to a 2.4-fold increase in C₃ plants, suggesting that enriched Streptomyces may enhance the adaptability of C₄ plants in arid environments (Liu, Li, and Singh 2024). In rice, Actinobacteria and Chloroflexi are significantly enriched in the root microbial community under drought conditions, potentially contributing to enhanced drought resistance (Santos-Medellín et al. 2017). Because HS often co-occurs with drought, grasses (e.g., wheat, sorghum, and maize) and other major crops like tomatoes selectively recruit drought-tolerant monoderm (Gram-positive) bacteria while reducing the abundance of diderm (Gram-negative) bacteria in the rhizosphere and roots (Trivedi et al. 2022; Naylor et al. 2017). This selective microbial assembly may similarly occur under HS conditions.

The rice rhizosphere represents a distinct ecological niche, characterised by an oxygenated zone closely surrounded by anoxic soil (Ding et al. 2019). In rice paddies, methanogenic archaea significantly influence soil health, rice growth, and greenhouse gas emissions by participating in organic matter decomposition, nutrient cycling, and methane production (Noll, Klose, and Conrad 2010). Notably, rice paddies are among the largest anthropogenic sources of atmospheric methane, with rice plants acting as conduits for methane release, a process highly sensitive to temperature fluctuations (Hosono and Nouchi 1997). Under HS at 40°C, the abundance of ammonia-oxidising bacteria (AOB) in the rice rhizosphere significantly decreases, leading to reduced nitrate availability in the soil (Munir et al. 2023). At 45°C, the structure and function of methanogenic archaeal communities are significantly affected, with thermophilic hydrogenotrophic methanogens (Methanocellales) exceeding 90% in abundance, while acetoclastic methanogens (Methanosaetaceae) decrease in relative abundance (Noll, Klose, and Conrad 2010). This shifts alters the metabolic pathways of acetate and propionate in paddy soils (Noll, Klose, and Conrad 2010), resulting in the accumulation of these organic acids and a corresponding increase in soil acidity during prolonged HS (Wu, Chin, and Conrad 2002). The resulting soil acidification may affect nutrient uptake by rice roots and microbial activity, but the precise implications remain unclear. Understanding how root-associated microbial communities influence ecosystem functions in rice under HS is essential for improving rice crop resilience and sustaining productivity in rice-based agroecosystems (Huang et al. 2021).

4.2 Microorganisms Alleviating Plant HS and Their Mechanisms: Hormone Regulation, Antioxidant Production, and Gene Expression Regulation

In recent years, the application of microorganisms has gained significant attention in advancing sustainable agriculture (Kumar and Verma 2018; Trivedi et al. 2021; Toju et al. 2018). Microorganisms play a crucial role in alleviating plant HS through diverse mechanisms (Table 1) (Carrell et al. 2022; Shaffique et al. 2022). The rhizobacterium, Pseudomonas brassicacearum, isolated from wheat rhizosphere soil, enhances antioxidant enzyme activity, increases protein synthesis, and improves water status in maize, thereby mitigating the effects of HS (Ashraf, Bano, and Ali 2019). Similarly, the endophytic fungus Penicillium glabrum alleviates HS in soybean and sunflower by reducing ABA and proline concentrations, enhancing antioxidant enzyme activity, and reducing ROS accumulation (Ismail et al. 2021). In rice, the bacterium Bacillus amyloliquefaciens SN13, originating from alkaline soil, significantly improves heat tolerance in seedlings by improving cell membrane integrity (reducing electrolyte leakage and malondialdehyde content), accumulating osmotic regulators (proline, sugars, and soluble proteins), activating the antioxidant defence system (increasing levels of superoxide dismutase, catalase, peroxidase, ascorbic acid, and glutathione), and upregulating the expression of stress-responsive genes (OsDHD, OsGST, OsLEA, OsNAM, OsGRAM, and OsNRAMP6) (Figure 3A) (Tiwari et al. 2017). Under HS conditions, the endophytic fungus Paecilomyces formosus LWL1 promotes rice growth by increasing fresh weight (50.32%), dry weight (47.52%), and chlorophyll content (26.81%) (Waqas et al. 2015). This fungus also reduces ABA and (jasmonic acid) JA levels, enhances the antioxidant system, and increases total protein content, thereby alleviating heat-induced damage in rice(Figure 3B) (Waqas et al. 2015).

Thermophilic microorganisms exhibit unique physiological and molecular mechanisms that enable them to adapt to extreme high-temperature environments (Urbieta et al. 2015). Their development and application in agricultural ecosystems have become a key focus of current research (Urbieta et al. 2015). The heat-tolerant Bacillus cereus SA1, capable of withstanding temperatures up to 45°C, significantly enhances the heat tolerance of tomato seedlings when inoculated (Khan, Asaf, and Khan 2020b). This is achieved through multiple mechanisms, including improved photosynthesis, reduced ABA levels, increased salicylic acid (SA) levels, enhanced antioxidant defences, and optimised nutrient uptake (khan, asaf, and khan 2020b). Similarly, Pseudomonas sp. AKM-P6, which grows at 50°C, enhances the heat tolerance of sorghum and wheat seedlings by increasing antioxidant enzyme activity, reducing ROS production, inducing the synthesis of high-molecular-weight proteins, and elevating levels of related cellular metabolites (Ali et al. 2009, 2011). Furthermore, Bacillus cereus TCR17, Providencia rettgeri TCR21, and Myroides odoratimimus TCR22, all capable of growth at 55°C, alleviate HS effects in sorghum (Bruno et al. 2020). These bacteria promote growth, strengthen the antioxidant system, reduce chromium accumulation, and regulate the expression of stress-related genes (Bruno et al. 2020). The thermophilic bacterium Ureibacillus sp. 18UE/10, isolated from hot springs, demonstrates additional potential in mitigating environmental stresses (Santana et al. 2020). It increases the nitrogen and phosphorus content in wheat stems under drought conditions, lowers the carbon-to-nitrogen (C/N) ratio in roots and stems, reduces the accumulation of toxic metals such as cadmium, lead, and nickel in wheat stems, and increases stomatal conductance (Santana et al. 2020). Interestingly, wheat root exudates regulate the adaptability of Ureibacillus sp. 18UE/10, fostering a more effective symbiotic relationship between the bacterium and the host plant (Santana et al. 2020). These traits, combined with its growth advantage in high-temperature environments, suggest that Ureibacillus sp. 18UE/10 holds significant potential for mitigating HS (Santana et al. 2020). In addition, the thermophilic endophytic fungus Thermomyces lanuginosus, isolated from extreme desert environments and capable of growing at 62°C, mitigates HS in cucumber (Ali et al. 2018). It preserves photosynthetic efficiency, improves water use efficiency, enhances metabolite accumulation, and reduces oxidative damage under high-temperature conditions in cucumber (Ali et al. 2018). In the future, microbe-based, eco-friendly tools are anticipated to enhance rice heat tolerance (Maldonado et al. 2022; Bardgett and Caruso 2020).

4.3 Microbial Assistance in Rice HS Resistance: Key Challenges to Overcome

The application of biocontrol agents in sustainable agriculture holds significant promise, particularly in minimising the reliance of chemical products (Velivelli et al. 2014). However, the colonisation ability of microorganisms on plants is crucial to their stable effectiveness in alleviating abiotic stresses such as HS (Gul et al. 2023; El-Saadony et al. 2022). Various application methods—such as seed treatment, foliar spraying, soil irrigation, and root soaking—as well as the timing of these applications, directly impact microbial colonisation and efficacy (Compant et al. 2019). A systematic investigation of the effects of microorganisms across different rice growth stages and application methods will help optimise their effectiveness (Gul et al. 2023).

However, even with successful initial colonisation, the long-term maintenance of microbial activity and function under HS remains an area requiring further investigation (Elnahal et al. 2022). Current research on alleviating plant HS primarily focuses on plants’ immediate responses to stress (Wei et al. 2013). Moreover, studies on microbial alleviation of HS in rice are largely concentrated on heat tolerance during the seedling stage, with relatively fewer investigations into the reproductive growth stages, which are directly linked to yield (Xu et al. 2020; Li, Cao, and Ma 2023b). More long-term and field studies related to HS are needed to bridge the gap between controlled experiments and natural field applications (Cox 2022).

In the future, it will be important to employ multiple strategies to enhance the colonisation and long-term effectiveness of microorganisms, thereby improving plant heat tolerance more effectively (D'hondt et al. 2021). Key approaches may include screening and developing microbial strains with strong survival abilities under high-temperature conditions, particularly those isolated from extreme environments such as hot springs, compost, or tropical soils (Jones, Marken, and Silver 2024). Moreover, constructing microbial communities that combine heat-tolerant and thermophilic microorganisms can leverage synergistic interactions to improve functional stability (Yang, Røder, and Wicaksono 2024b). It is also essential to study the specific molecular pathways and mechanisms by which microorganisms alleviate plant HS (Kan et al. 2023). Furthermore, incorporating new technologies, such as coating techniques or biofilm technology, may improve microbial colonisation efficiency and stability in the rhizosphere or plant tissues (Elnahal et al. 2022; Sohail et al. 2022). Finally, a comprehensive evaluation of the environmental risks and economic benefits of microbial inoculants is necessary to translate laboratory findings into practical applications (O'Callaghan, Ballard, and Wright 2022; Batista and Singh 2021).