Plants continuously release exudates into their surroundings, in the form of liquid, solid, or gaseous compounds, through their leaves, shoots, or roots. Plants exuded c. 11% of the net fixed C or 27% of the C assigned to roots to the rhizosphere. The amount of these compounds varies depending on multiple factors, such as plant age, species, and nutritional performance (Bais et al., 2006; Jones et al., 2009; Nakayama and Tateno, 2018). Root exudates contain various substances, including primary and secondary metabolites and phytohormones, such as ET, JA, SA, indole 3-acetic acid (IAA), abscisic acid (ABA), gibberellic acid (GA), and cytokinins (CKs). Primary metabolites consist of sugars, amino acids, and organic acids, while secondary metabolites include flavonoids, glucosinolates, phytoalexins, triterpenes, and benzoxainoids (Badri et al., 2008; Pang et al., 2021). For instance, tomato plants exude a mixture of metabolites, including organic acids, steroidal glycoalkaloids, acylsugars, and hydroxycinnamic acid derivatives, into the rhizosphere (Korenblum et al., 2020).

Microbes diverge in their functioning and metabolism, and their sensitivity to water availability varies. Thus, drought can directly impact the assembly of plant microbiome. Plant surface microbiome, i.e., phyllosphere, is likely to be significantly altered by drought since environmental conditions vary more rapidly than those inside plant tissue (i.e., endosphere), which is more stable (Trivedi et al., 2020). Most of the microbes from the bulk soil (a source of potential microbes to colonize plant roots) are directly influenced by external climatic factors, including drought. On the other hand, rhizosphere microbiome are directly altered by these factors and indirectly by plant responses, such as changes in host morphology, physiology, immune system, and root exudation (Figure 2). Previous studies suggest a consistent response of the host-associated microbiome to water deficiency (Vescio et al., 2021; Wipf et al., 2021; Aslam et al., 2022; Tebele et al., 2023). Under water limitation, several plant species recruit monoderm bacteria that are resistant to dryness owing to their thicker cell walls and reduce diderm bacteria in the roots and rhizosphere (Naylor et al., 2017; Naylor and Coleman-Derr, 2018).

Understanding how drought stress affects plant–microbiome assemblage is still a challenging task owing to the multilayers and complex interconnections that orchestrate this ‘triangle’ interaction. Chemical signals exchange heavily influences plant-microbiome communication under water scarcity (Figure 2). For example, under stressful environments, plants have evolved a ‘cry for help’ mediated exudation, resulting in the recruitment of stress-mitigating microbes (Liu et al., 2020). Plants have a complex microbial recruiting system to select the most beneficial microorganisms to incorporate into plant tissues, discriminating between friendly and beneficial or hostile and harmful interactions (Hacquard et al., 2017; Teixeira et al., 2019; Zhang et al., 2023). The host plant possesses protective mechanisms to perform this selection: (i) pattern recognition receptors (PRRs), the initial mechanism of defense to identify microbe-associated molecular patterns (MAMPs), such as fungal chitin or bacterial flagellin, leading to MAMP-triggered immunity (MTI) and (ii) nucleotide-binding leucine-rich repeat (NLR) proteins as a second defense mechanism to identify pathogen effectors, inducing effector-triggered immunity (ETI). Changes in plant immunity caused by drought could structure the plant-microbiome assembly, especially inside host plant tissues. Suppression of ETI may interfere with host-mediated management of microbe colonization and may result in microbial dysbiosis inside plant tissues. The suppression of ETI can also be a new pattern used by plants to lessen their defense mechanisms, allowing beneficial microbes to colonize roots and promote stress mitigation-related gene transcripts.

The mutual communication involving the plant defense system and the microbiome structures the plant-microbiome assemblage. Plants control immune response in rapidly changing environmental conditions using active but tightly controlled modifications in different hormone pathways (i.e., ET, SA, ABA, and JA; Li et al., 2021; Ait-El-Mokhtar et al., 2022). Lebeis et al. (2015) reported that drought reduces SA production, which is involved in the assembly of both endophytic and epiphytic microbiome. SA can promote or inhibit microbial community growth directly on microbiome members or through established signaling pathways based on hormonal cross-talk. For instance, ABA-induced production under water scarcity interferes with the SA-mediated immune pathway. Drought-mediated alterations in plant hormones may differ depending on the plant developmental stage and tissue type. For example, maize grown under water limitation boosts benzoxazinoid defense system in plant aboveground part while stimulating terpenoid phytoalexins in the belowground tissues (Vaughan et al., 2018). Alterations in the distribution or allocation of diverse signaling molecules or defense metabolites as a result of drought may have an additional impact on microbiome assembly.

It is worth noting that in host-microbiome research, core-and-hub microbiota concepts are gaining traction (Singh B. K. et al., 2020). These refer to the microbiota that exist in a specific species regardless of environmental conditions, growing season, or management practices and perform critical host functions (Trivedi et al., 2020). Given their significance, it is crucial to understand how drought affects the ‘core-and-hub’ microbiota that may organize community-scale functions in plant-microbiome communications. An enlarged understanding of the ecological vectors that orchestrate microbiome response to water scarcity will promote our knowledge of microbiome traits that boost plant performances under changing environments.

Plant and microbiota have a two-way communication, both underground and aboveground, that enables them to sense and respond to stress conditions. In response to stress, plants release a range of metabolites that attract specific microorganisms capable of enhancing their tolerance to stress (Bai et al., 2022). According to the ‘cry for help’ hypothesis, plants recruit particular microbiome communities that aid them in managing stress (Liu et al., 2020). This concept was first observed when plants grown in soils deficient in phosphorus and nutrient-supplying arbuscular mycorrhizal fungi (AMF), and nitrogen recruited nitrogen-fixing rhizobia (Carbonnel and Gutjahr, 2014; Nishida and Suzaki, 2018). The ‘cry for help’ assumption is also applicable to plants experiencing drought stress, as the microbiome composition in roots significantly changes by promoting actinobacteria and other gram-positive bacteria over gram-negative ones (Timm et al., 2018). Plants may selectively recruit drought-tolerant microbes that have evolved from repetitive drought periods, resulting in beneficial and efficient plant-microbiome interactions that enhance the performance of both host and microbes (Naylor and Coleman-Derr, 2018). Terhorst et al. (2014) demonstrated that drought-stressed Brassica rapa plants exhibit higher and more diverse bacterial richness around their roots in comparison to the controls.

Santos-Medellín et al. (2021) revealed that the above- and below-ground microbiome undergo a plant-driven change in response to water limitation, resulting in an increase in drought-tolerant endophytic monoderm bacteria that may help mitigate drought. The same authors demonstrated how persistent drought can permanently impede plant endophytic microbiome growth. This effect persists even after the drought constraint is alleviated. Their research revealed how long-lasting drought stress may shift microbial community composition, which may affect plant fitness. They also identified promising candidates for microbiome engineering to create performant microbial assemblages against water deficit, including drought-resistant endophytic microorganisms that increased in abundance in the endosphere after drought stress. Active microorganism recruitment under stressful conditions appears to be a common evolutionary mechanism to promote plant performances. Nonetheless, the strategies that allow plant host to incorporate external signals during symbiotic microbes’ recruitment and the host genetic characters controlling this recruitment are still under investigations. These processes are orchestrated by multilayer components involving plants, microorganisms, soil, and environmental traits that shape the final result. The accumulation of stress-related factors in plant roots during drought, including G3P and pipecolic acid, has been linked to actinobacteria enrichment in the rhizosphere (Knight et al., 2018; Caddell et al., 2020; Table 2). Understanding the underground signals communication is still in its early stages, particularly in drought-stressed hosts that shape their tolerant microbes.

A dynamic understanding of the role of metabolites and their genomic features is essential to comprehend the multilayered complexity of the ‘cry for help’ theory in host microbe assembly before and after drought conditions. This will provide new insight into creating drought-tolerant microbial consortia for sustainable agriculture. Recently, Bai et al. (2022) proposed a model based on the ‘cry for help’ hypothesis for plant microbiome recruitment under water scarcity (Figure 2). Under drought conditions, plants undergo molecular and metabolic readjustments and produce specific root metabolites. The root exudates may promote the reprogramming and restructuring of the microbiome by recruiting selective drought-tolerant microorganisms with a vast arsenal of functional enzymes. Subsequently, drought-resistant microbiota can mitigate stress impact and deliver nutrients to host plants via multiple direct and indirect mechanisms.

The plant and its associated microbiome form a harmonized and functional entity known as a holobiont (Bordenstein and Theis, 2015), in which evolutionary selection occurs not only between the host and the associated microbes but also among the microbes (Ait-El-Mokhtar and Baslam, 2023). To maintain the harmony of this association, systems of coordination among the microbial communities and with the host plant are necessary. Our understanding of the acquisition of microbes in the environment and the rules governing their association in the plant holobiont is very limited. It is widely believed that during root penetration into bulk soil, the soil microbiome progressively differentiates into the rhizosphere microbiome via contact with rhizodeposits, which have a significant effect on the microbiota composition (Tian et al., 2020; Figure 2). After this initial community shift, the microbiota composition is finely adjusted in specific compartments (rhizosphere, rhizoplane, or endosphere; Bulgarelli et al., 2013). Plant genetic factors, including root exudate quality and quantity and root morphology, have been known to shape the rhizospheric microbial communities (Sasse et al., 2018). Other factors, including the plant developmental stage, plant immune system, and season, have also been shown to play a significant role in shaping the rhizosphere microbiome (Hassani et al., 2018). The presence of a core microbiota in some crops, irrespective of fertilization management or soil origin, suggests that these communities are partly assembled and selected by plants. In general, it is believed that the plant selects its microbial associates via the action of its root exudates. Yet, this unidirectional recruitment is being questioned (Uroz et al., 2019) because the co-evolution of the plant-microbe holobiont suggests bidirectional interactions, particularly in terms of rhizosphere microbiome shaping (Figure 3).

Conventionally, root exudate secretion was thought to be a passive process facilitated by various pathways, such as diffusion across the root cell membrane, ionic channels, and vesicle transport (Baetz and Martinoia, 2014). The chemical characteristics of the exuded molecules determine the type of the secretion pathway. Diffusion is involved in the exudation of metabolites with low molecular weight, such as amino acids, sugars, carboxylic acids, and phenolics. This process is caused by the difference in the concentrations of compounds between root cell cytoplasm and rhizosphere, and it may be influenced by root membrane permeability, root cell integrity, and compound polarity (Badri and Vivanco, 2009). Ionic channel pathways are involved in the exudation of carbohydrates and particular carboxylates, including oxalate and malate (released in large amounts), which are transported through membranes via a transport mechanism performed by proteins rather than diffusion. Two types of anionic channels are involved in this process: SLow Anion Channels (SLACs), which are activated in many seconds, and QUick Anion Channels (QUACs), which take a few milliseconds to be activated (Dreyer et al., 2012). The third form of passive transport pathways is vesicle transport (exocytosis) responsible for the exudation of high molecular weight metabolites kept inside vesicles (Badri and Vivanco, 2009). Exuded compounds are produced by the endoplasmic reticulum or the Golgi apparatus and aid in pathogen defense (Weston et al., 2012). In contrast, the active transport pathway of root-secreted metabolites is mediated by plasmatic membrane proteins (Baetz and Martinoia, 2014): ATP-Binding Cassette (ABC) and Multidrug and Toxic Compound Extrusion (MATE) (Kang et al., 2011). Protein-mediated root exudation may take three forms and depends on their specificity: transporters exuding multiple compounds, compounds secreted via various membrane transporters in the rhizosphere, and metabolites secreted by a single transporter. ABC transporters are classified as primary transporters owing to the use of ATP hydrolysis for the necessary energy to transport varied solutes (Jones and George, 2002; Orelle et al., 2018). The gene names encoding these transporters have evolved over time. Nevertheless, another classification has been established based on the organization of TMD and NBD domains, assembling the various members into nine families termed with letters ranging from A to I, despite the fact that family H does not exist in plants (Verrier et al., 2008). Some studies have looked into the involvement of ABC transporters in root exudation and highlighted their significance in this process (Badri et al., 2008, 2009; Olanrewaju et al., 2019).

Recent studies have shown that shifting from naturally occurring microbiome to adapted microbial communities to the stressful condition enables the plant holobiont to quickly adapt to changing environments (Pantigoso et al., 2022; Faist et al., 2023; Li et al., 2023). Rice provides an example of plant-mediated microbial abundance modulation, where under water deficiency, the microbiome composition shifts to a greater abundance of drought-resistant plant growth promoting rhizobacteria (PGPR) strains (Santos-Medellín et al., 2017). Changes in root exudation caused by water shortage may be the source of the enhancement of several types of microorganisms. Stressful soil conditions, along with microbial composition changes, can influence rhizosphere microbial activity. The modulation of the rhizosphere microbiome is influenced not only by plants but also by microbes. By exuding phytohormones, antimicrobials, volatile compounds (VCs), and quorum-sensing, plants may regulate the plant environment and even benefit their hosts (Venturi and Keel, 2016). Under water limitation conditions, the rhizosphere microbiome is shaped by microbial interactions among members and their preferences for specific metabolites (Zhalnina et al., 2018). Microbial interactions may alter gene expression within communicating microbes (Sasse et al., 2018), indicating that microbial interactions affect both the function and shape of the microbiome. According to Uroz et al. (2019), both partners are ecological engineers of the holobiont because they regulate plant-associated microbes. Therefore, microbial communications (and plant-modulating microbiome) in the rhizosphere play a critical role in shaping plant-associated microbiome.

Using metatranscriptomic analysis to study the wheat holobiont under water deficiency, Pande et al. (2023) revealed that the microbial associates were the most responsive to water deficit. The majority of the differentially abundant (DA) genes were associated with bacteria in the rhizosphere and fungi in the roots when comparing drought-stressed wheat and control treatments. In the rhizosphere, Actinobacteria were overrepresented in positively regulated DA transcripts, while Acidobacteria and Proteobacteria were overrepresented in negatively regulated ones. These authors demonstrated that certain transcripts were more abundant in the roots and rhizosphere under severe water stress, including heat shock proteins (HSPs), as well as carbohydrate and amino acid transport and metabolism-related transcripts. Another metatranscriptomic study reported that drought stress promoted the transcripts of Proteobacteria and Bacteroidetes, while reducing those of Actinobacteria and Acidobacteria (Tartaglia et al., 2023). This study also indicated an over-expression of universal stress proteins by Proteobacteria and Bacteroidetes in water-stressed treatment compared to the control. Furthermore, Xu et al. (2018a,b) revealed greater enrichment in various Actinobacteria and Chloroflexi taxa and a decline in the abundance of different Acidobacteria and Deltaproteobacteria taxa under drought conditions. This change was coupled with an increase in G3P-related transcripts in Actinobacteria.

In this cross-kingdom communication, there has been a recent surge of interest in small RNAs, specifically microRNAs, as active ‘hormone-like’ mediators in cell-to-cell communication (Leitão et al., 2020). Arabidopsis and Botrytis cinerea, a fungal pathogen, were the first to demonstrate the bi-directional cross-kingdom RNAi via sRNA trafficking (Wang et al., 2016). Still, multiple studies are emphasizing the function of microRNA exchanges in non-pathogenic associations with a focus on the host-microbiota communication in the gut (Liu et al., 2016; Bi et al., 2020; Casado-Bedmar and Viennois, 2022). Intestinal epithelial microRNAs have been reported to influence the composition of the gut microbiome by penetrating specific bacteria and controlling the transcription of a large number of genes involved in sugar degradation and housekeeping, thereby influencing the bacterial fitness (Gao et al., 2019). After their ingestion, plants secrete extracellular vesicles (EVs) in the gut, where specific bacteria can uptake them with their microRNA, causing an alteration in their gene expression (Mu et al., 2014; Teng et al., 2018). As a result, their metabolite production and secretion could be modified, resulting in differential growth of bacterial strains that interact with these specific bacteria. Following these advances, Middleton et al. (2021) propose that plants and their associated rhizosphere microbes interact through microRNAs, that modulate the composition and activity of rhizospheric microbiota. Strikingly, hundreds of microRNAs have been found in host root tissues (Breakfield et al., 2012). It is therefore assumed that some microRNAs, selected through co-evolution with nearby microorganisms, could be secreted through EVs structure in the rhizosphere.

Taking a step further, Escudero-Martinez et al. (2022) applied metagenomics data to map the plant genetic factors regulating microbiota in the rhizosphere of domesticated and wild barley genotypes. The authors identified a small number of loci that have a significant impact on the rhizospheric microbiome composition. One of those loci, called QRMC-3HS, emerged as a major determining factor of microbiota composition. A comparative root RNA-seq profiling of soil-grown sibling lines with contrasting QRMC-3HS alleles and presenting distinct microbiotas enabled the identification of three primary candidate genes: Nucleotide-Binding-Leucine-Rich-Repeat (NLR) gene and two other genes from QRMC-3HS. The NLR gene encodes an NLR protein, one of two types of immune system receptors involved in the microbe proliferation recognition through effector recognition (Jones and Dangl, 2006). The first gene of QRMC-3HS is differently regulated in sibling lines with different microbiotas, and it is unclear how it mechanistically contributes to rhizospheric plant-microbiota interactions because it encodes an unknown protein. The second gene encodes a xyloglucan endotransglucosylase/hydrolase (XTH) enzyme involved in the cleavage and/or rearrangement of xyloglucans (Yang et al., 2019), which are the most abundant hemicellulosic compounds in plant primary cell walls (Ezquer et al., 2020). The XTH gene may still be involved in microbiota shaping through cell wall polysaccharide modification closely related to plant-microbe interactions (Vorwerk et al., 2004). In Arabidopsis, cell wall traits serve as recruitment cues for almost 50% of the endogenous root microbial communities (Bulgarelli et al., 2012). In addition, cell wall alterations underpin some of the gene ontology classes identified in genome-wide association mapping experiments carried out using this plant (Horton et al., 2014). Improved adaptation to soil physicochemical conditions may represent further involvement of XTH genes in plant-microbiome interactions. Han et al. (2017) previously reported that XTH genes are involved in drought stress tolerance.

As previously discussed, soil microbes provide various benefits to plants, with root-associated microbiome playing a key role in determining host fitness and performance under various environments, including drought. Root microbiome composition is influenced by both plant and environmental factors. Plant-associated microbiome may boost host development under drought stress by stimulating different layers of plant tolerance mechanisms (Table 1). Among these microbes, PGPRs are abundant in the rhizospheric area and have been shown to be one of the most successful strategies for mitigating the adverse effect of water shortage on the host plant (Etesami and Jeong, 2018). PGPRs enhance crop plant resistance by increasing the production of osmolytes (including glycine betaine and proline), accumulating secondary metabolites, and modulating the expression of a myriad of host drought-related genes.

Vurukonda et al. (2016) revealed that PGPRs associated with stressed plants are crucial for inducing systemic tolerance (IST). A metabolome study of drought-stressed Sorghum bicolor primed with Bacillus and Pseudomonas isolates showed an induction of signature metabolic profiles and biomarkers related to IST in the host (Carlson et al., 2020). The findings revealed substantial treatment-related differential metabolic reprogramming among rhizobacteria-treated and control plants. This was correlated to the ability of the selected isolates to preserve host plants against drought stress by up-regulating IAA, CK, SA, GA, JA, brassinosteroïds (BRs), sphingosine, psychosine, osmolytes and antioxidants, and down-regulating ET production. Raheem et al. (2018) found that IAA and other auxins produced by PGPR bacteria enhance plant performance under drought stress, as demonstrated by Bacillus amyloliquefaciens, an auxin-producing bacteria isolated from Acacia arabica rhizospheric areas in arid climate. The production of exopolysaccharide by PGPRs is another cue inducing plant drought tolerance. Using a Bacillus amyloliquefaciens exopolysaccharide-deficient mutant (epsC), Lu et al. (2018) demonstrated that the epsC gene is a key gene involved in drought tolerance in Arabidopsis (Table 3). The authors highlighted the effect of ET and JA in inducing IST and up-regulating many drought-resilient genes (including ERD1, LEA14, RD17, and RD29A) in leaves. Staudinger et al. (2016) used proteomics to show that Sinorhizobium sp -inoculated Medicago truncatula grown under drought induced hormonal crosstalk and increased JA translational regulation, playing a role in increased leaf maintenance in nodulated plants during drought. PGPRs have been shown to accelerate the antioxidant enzymes and osmoprotectants biosynthesis in drought-stressed plants. By using various formulations of PGPRs isolated from Megathyrsus maximus rhizosphere in dry areas, Moreno-Galván et al. (2020) showed that the isolates improved Megathyrsus drought tolerance by up-regulating proline biosynthesis and down-regulating malondialdehyde (MDA) concentration and glutathione reductase activity. Recently, Singh D. P. et al. (2020) revealed that the overexpression of genes encoding enzymes responsible for phenylpropanoid has been linked to ROS scavenging in rice plants inoculated with Pseudomonas and Trichoderma and grown under water scarcity conditions. Similarly, upregulation of genes encoding PiP, DHN, and DREB contributes to the improvement of drought resistance in plants treated with PGPRs. Drought-stressed tomato plants treated with PGPRs yielded enhanced growth traits, water status, proline biosynthesis, antioxidant enzyme defense (i.e., ascorbate peroxidase, catalase, and glutathione peroxidase), and H₂O₂ accumulation compared to the controls (Abbasi et al., 2020). The authors demonstrated that PGPR-treated tomato plants showed altered stress signaling and regulatory networks controlling the expression of target genes related to plant response to drought, such as ERF1 and WRKY70. Woo et al. (2020) reported that Bacillus subtilis boosted drought resilience in Arabidopsis and Brassica by up-regulating the expression of drought-sensor genes, including NCED3, RAB18, RD29B, and RD20 in Arabidopsis and WRKY7, CSD3, and DREB1D in Brassica.

Several studies have shown that specialized microbiome may alleviate plant drought stress (Mathur and Roy, 2021; Aslam et al., 2022; Singh et al., 2023). Endophytes, microorganisms living in the endosphere without inducing disease symptoms, confer stress resilience to host species and perform a significant function in the survival of some plants in high-stress environments. Various bacterial and fungal endophytes have been reported to increase plant drought resilience. Bacterial endophytes isolated from drought-stressed maize roots improved plant fitness due to biosynthesis of IAA, GA, and CK, 1-aminocyclopropane-1-carboxylate (ACC) deaminase activity and siderophore biosynthesis (Sandhya et al., 2017). Drought-stressed sorghum plants treated with a consortium of root bacterial endophytes (Enterobacter cloacae, Enterobacter sp., Ochrobactrum sp., and Microbacterium sp.) resulted in higher growth and osmotic adjustment (Govindasamy et al., 2020). Drought tolerance mediated through microbial inoculation has been associated with enhanced proline accumulation and proline-related biosynthesis genes, SbP5CS 1 and SbP5CS 2. Interestingly, Vigani et al. (2019) found that root endophytic bacteria conferred drought resistance in Capsicum annuum by modifying vacuolar H⁺ pyrophosphatases (VPP) expression, which aided in preserving osmotic balance. Strains of Bacillus endophytes found in Lepidium perfoliatum roots were responsible for the formation of biofilm on roots, resulting in drought resistance and seedling germination (Li et al., 2017). Piriformospora indica, a well-known root fungal endophyte, colonizes the roots of many plant species and provides multifaceted amenities, including drought alleviation (Gill et al., 2016). Metabolome and proteome profiling analysis of barley plants inoculated with P. indica- grown under drought stress showed that inoculation redistributes resources, maintains the aquaporins (AQPs) presence, and promotes energy modulation, photorespiration protective proteins and transporters production, primary metabolism, and autophagy in water-stressed plants (Ghaffari et al., 2019). Inoculation of rice roots with P. indica increased growth, biomass accumulation, mineral nutrition (Zn and P), and upregulated P5CS genes in drought-stressed plants (Saddique et al., 2018). González-Teuber et al. (2018) reported that inoculating the roots of Chenopodium quinoa with the fungal endophyte Penicillium minioluteum primarily conferred drought tolerance through significant adjustments in below-ground biomass, photosynthesis, water-use efficiency (WUE), and photochemical efficiency.

AM symbiosis is one of the most complex and mutualistic interactions that plants have evolved to cope with droughts. Following a molecular dialog between the two partners involving “branching factors,” “mycorrhizal factors,” and common symbiotic signaling pathways, the AMF enters the plant roots and establish arbuscule formations ensuring nutrient exchange. Plant hormones act as central regulators of the development of the plant-AMF interaction (Santner et al., 2009; Charpentier et al., 2014; Etemadi et al., 2014), and auxins have potential key role in this interaction (Takeda et al., 2015; Jin et al., 2016). The modulation of AMF-mediated host drought tolerance is an extremely intricate process implicating multiple metabolites and pathways. Particularly, AMF can directly improve nutrients and water absorption and transport, boost host osmotic regulation, and increase plant gas exchange capacity, WUE, and antioxidant defense (Osakabe et al., 2014; Ruiz-Lozano et al., 2016). Zhang et al. (2019) found that a combination of Funneliformis mosseae, Rhizoglomus intraradices, and Diversispora versiformis improved the survival of Zenia insignis subjected to water scarcity through regulating osmolytes accumulation, antioxidant enzyme activity, and plant N and P uptake. Drought-stressed trifoliate oranges inoculated with F. mosseae have less oxidative stress via the increase of H₂O₂ efflux from the root system (Santner et al., 2009; Jin et al., 2016). In addition, the regulation of polyamine metabolism-associated gene transcripts by AMF has been identified to play a pivotal role in drought resilience (Zhang et al., 2020). Taking a step further downstream, several AM-specific host genes and proteins have been reported to play a key role in promoting plant water stress resilience. The different mechanisms involved in conferring drought resilience to mycorrhizal plants have been previously reviewed (Cheng S. et al., 2021; Wang et al., 2023). Drought-induced genes and compounds can be divided into: functional genes playing a direct role in stress [i.e., late embryogenesis abundant (LEA) proteins, AQPs, sugar, and proline] and regulatory genes implicated in the regulation of gene expression and the transduction signals (i.e., stress-related TFs) and signal molecules, such as calmodulin-binding protein. Many aspect of the molecular regulatory network linking AM symbiosis and drought stress have been elucidated, including the identification of several gene/protein functions (Bahadur et al., 2019; Ho-Plágaro and García-Garrido, 2022). Various TFs, such as AP2/ERF family, GRAS family, and MYB family have been found to be specifically induced by AMF under drought conditions, thereby contributing to the modulation of stress by phytohormones and other molecular signaling pathways (Wang et al., 2023). Jia-Dong et al. (2019) revealed that AM symbiosis significantly activated the expression of root tip AQPs PtTIP1;2, PtTIP1;3, and PtTIP4;1 in drought-stressed trifoliate orange. In the same vein, it has been reported that drought stress induced the up-regulation of eight AQP-associated genes in AMF-treated plants (Recchia et al., 2018). An interactive impact of AMF and drought was recorded on the over-expression of MAPK pathway gene that triggers an improvement in photosynthetic efficiency, osmolyte production, and antioxidant defense (Huang et al., 2020). Testerink and Munnik (2005) showed the role of ABA in up-regulating resistance genes expression to mitigate drought-induced damages. Under water limitation, lettuce inoculated with G. intraradices exhibited high expression of the Lsnced gene, which encodes a pivotal enzyme in the production of ABA. In contrast, the expression of this gene in roots was not affected by the exogenous application of ABA. This suggests that mycorrhizal plants adjust the endogenous ABA levels more efficiently and quickly than non-AM plants, leading to a more appropriate equilibrium between water acquisition and leaf transpiration under water deficit (Aroca et al., 2008). In a proteomic study on wheat, differential expression proteins involved in cell wall integrity and carbohydrate production were observed, while most stress-associated molecules, including enzymes involved in ET biosynthesis, were downregulated (Bernardo et al., 2017).

Recently, Poveda (2020) found an increase in the secretion of the fungal enzyme chorismate mutase by using mutant strains of Trichoderma parareese. This increase conferred tolerance to rapeseed plants grown under drought conditions by increasing gene expression (i.e., NCED3 and PYL4) related to the hormonal pathways of ABA. Notably, colonization of Trichoderma sp. on rapeseed roots increased under stressed condition, initiating a myriad of host metabolic pathways. Comparative qRT-PCR analyzes with the chorismate mutase-silenced strain connected this enzyme to drought-tolerant mechanisms owing to its involvement in ACCO1 (1-Aminocyclopropane-1-carboxylic acid oxidase) and ERF1 downregulation, and ABA pathway genes NCED3 upregulation. In addition, Bashyal et al. (2021) used transcriptome profiling of droughted- Trichoderma harzianum-inoculated rice and revealed the upregulation of 1,053 genes and the downregulation of 733 genes in stressed T. harzianum-inoculated plants. Most photosynthetic and antioxidative genes, including plastocyanin, PSI subunit Q, PSII subunit PSBY, small chain of Rubisco, proline-rich protein, osmoproteins, stress-induced proteins, AQPs, and chaperonins, were exclusively expressed in stressed T. harzianum-inoculated rice. Using the enrichment analysis, the same authors showed that the metabolic (38%) and pathways involved in the synthesis of secondary metabolites (25%), phenyl propanoid (7%), carbon metabolism (6%), and glutathione metabolism (3%) were the most enriched pathways.

The co-inoculation with different members of plant microbiome was also reported to enhance host drought tolerance. Eshaghi Gorgi et al. (2022) studied the effect of the dual application of PGPR and AMF on biomass accumulation, water status, photosynthetic pigments, and proline content of drought-stressed Melissa officinalis. They demonstrated that the combined microbial treatment increased all these parameters under drought stress. The same authors also reported that leaves chemical composition of secondary metabolites was altered by PGPR+AMF inoculation. Another study showed that the co-inoculation of Common myrtle with Funneliformis mosseae and Rhizophagus irregularis AMF strains and Pseudomonas fluorescens and P. putida PGPR strains boosted seedlings survival, growth fitness, and (non-)enzymatic antioxidant accumulation while reducing electrolyte leakage, and MDA and proline concentrations under drought stress (Azizi et al., 2021). In the same vein, the combined application of Glomus versiforme and Bacillus methylotrophicus recorded significant drought resistance through improving growth and photosynthetic performances, nutrition status, phenols and flavonoids accumulation, antioxidant enzymatic system and ABA and IAA concentrations in tobacco plants under drought stress (Begum et al., 2022). Singh D. P. et al. (2020) revealed that Trichoderma and Pseudomonas primed rice seeds induced significant increase in the transcript levels of multiple genes involved in the antioxidant enzymes and phenylpropanoid biosynthesis pathway in seedlings grown under drought stress.

Several studies have revealed the significant beneficial effects of combining microbes and others biostimulants in mitigating drought stress. The application of AMF and organic amendment improved the tolerance of pistachio seedlings to drought stress through enhancing soil physicochemical and biological traits, as well as plant nutrient uptake (Paymaneh et al., 2023). Soussani et al. (2023) demonstrated that the application of AMF and/or compost promoted tomato growth, yield and fruit bioactive compounds, while reducing oxidative stress and enhancing the efficiency of the antioxidant enzyme system under water stress. The combined effect of AMF, PGPR, and compost boosted tomato growth fitness, fruit yield and quality, while increasing drought stress tolerance (Tahiri et al., 2022). A two-year field experiment showed that the application of different biostimulants such AMF, PGPR, and seaweed extract increased wheat root volume, membrane stability index, leaf relative water content (RWC), and photosynthetic pigment content, which promoted plant resilience under water shortage conditions (Najafi Vafa et al., 2022).

The ubiquitous presence of the soil microbiome and its impact on various aspects of plant functioning under drought stress have made it increasingly challenging to isolate specific aspects of the microbiome-plant-drought axis. This highlights the need for multi-disciplinary interventions to uncover opportunities and strategies for enhancing crop drought resistance (Hartman and Tringe, 2019; Ali et al., 2022). Multidomain research approaches that combine plant resilience, microbiome recruitment, and the interactions among the different (a)biotic components of the ecosystem have proven to be valuable strategies. They establish connections that modulate microbiome composition and activity, leading to improved drought tolerance (Trivedi et al., 2022). Yet, the full potential of the host microbiome in sustainable agriculture is not yet fully realized due to multiple factors, including soil type, plant genotype, microbial interactions, agricultural practices, and the complex interplay among these components (Busby et al., 2017; Soman et al., 2017; Schmidt et al., 2019). Moreover, microbiome engineering faces challenges in achieving accurate and long-lasting positive effects on plants. Multiple interconnected factors, such as the richness and complexity of microbial communities and alterations in microbiota functioning during host development stages, contribute to the complexities that limit the effectiveness of microbiome engineering.

To maximize the benefits of plant microbiome assembly under drought stress, it is crucial to investigate the genetic complexity of both plants and their associated microbiome, the heterogeneity of their environment, and the metabolic patterns that influence the host microbiome. This requires the application of inclusive and systemic biological methods coupled with advanced multiomics techniques. While existing tools and methodologies have made significant advancement in understanding the effect of the microbiome on host resistance to water stress, there are still many gaps to be filled in order to make substantial progress toward microbiome-targeted approaches for enhancing crop production and resilience under drought stress.

Many approaches such as culture-dependent, culture-independent, and reductionist synthetic communities (SynCom) have been primarily used to study the root microbiome. However, reproducing a suitable natural conditions required for the development of different microbes in the laboratory is challenging, and a large number of them cannot be cultured, leading to a loss of microbiome information (Hill et al., 2000). However culture-dependent methods, including the reductionist SynCom method remain highly efficient for microbiome studies (Bai et al., 2015; Zhang et al., 2021). DNA fingerprinting and phospholipid fatty acid approaches, as culture-independent methods, focus on studying the composition and diversity of the entire microbiome community. However, these approaches provide less data in comparison to the current advances in metaomics, sequencing, and computational techniques (Jo et al., 2020). By utilizing advanced sequencing methods, studies have achieved unprecedented precision and comprehensiveness in understanding the composition of microbial communities under drought condition (Lundberg et al., 2012; Liu et al., 2019). Furthermore, the development of metagenomics, metatranscriptomics, and metaproteomics approaches enable a thorough understanding of microbiome function under drought stress (Liu et al., 2021).

The census method, which includes metagenomic and amplicons statistics, offers a comprehensive understanding of plant associated-microbes, which is essential for studying natural microbiome facts. On the other hand, the reductionist SynCom method links plant molecular biology to microbial ecology (Guttman et al., 2014; Mendes et al., 2014; Bulgarelli et al., 2015; Edwards et al., 2015; Liu et al., 2019). The census approach provides data for the reductionist investigation of plant-microbiome interactions. By combining this information with data on isolated strains, an extensive range of representative SynCom can be created, providing accurate genetic information (Bulgarelli et al., 2012; Vorholt et al., 2017). These SynComs are then used to simulate plant-microbiome interactions, allowing the study of the components that orchestrate microbial communities and validate their monitoring functions and molecular strategies throughout plant growth and development, even under stressful conditions, such as drought (Bai et al., 2015; Castrillo et al., 2017; Chen et al., 2020; Finkel et al., 2020).

Multiomics approaches offer a valuable tool for addressing the challenging task of translating plant alterations at the genetic, proteomic, or metabolomic levels. The integration of multiomics data-driven science has significantly advanced our understanding of microbiome composition and functional responses in intricate environments, such as the rhizosphere, where the complex network of microbial connections orchestrates plant behavior under stressful conditions. The combination of these approaches has led to remarkable progress in plant-microbiome-related research.

Each technique employed in multiomics approaches brings unique advantages, but no single approach can provide a comprehensive understanding of the mechanisms governing the assembly of plant-associated microbiome under drought stress. Therefore, it is crucial to integrate various research approaches to pave the way toward a comprehensive understanding and utilization of the microbiome in future studies, with the aim of developing targeted strategies to boost plant resilience to water deficit stress.

Unlocking the intricate interactions between plant and their microbiome under water limitation and their implications for plant fitness and productivity are crucial steps toward harnessing the potential of the microbiome to promote host adaptation to environmental perturbation. Exploring the extent of these interactions at evolutionary, ecological, biochemical, and molecular levels can provide valuable insights for advancing our system-level understanding and inform microbial approaches to improve host resistance and fitness. Several key research directions should be prioritized in the future:

Gain a complete comprehension of how drought impacts the assembly and functions of the plant microbiome across different temporal and spatial scales.

Investigate the effect of drought on plant fitness and defense system, elucidating the specific alterations in photosynthates, root exudates, and defense mechanisms, and their influence on the assembly and functions of the plant microbiome.

Determine the changes in major signaling cascades and metabolite profiles and their interactions, elucidating their roles in shaping host plant and microbial functions and fitness.

Increase our understanding of the biosynthetic pathways, genetics, and mechanisms of action of drought-responsive phytohormones, as well as their effects on plant-microbiome and microbe-microbe interactions.

Identify the frequency and duration of drought necessary for the eco-evolutionary adaptation of plant microbiome and the establishment of drought resilience in host plant.

Acquire cutting-edge knowledge about the molecular interactions that orchestrate plant-microbe interactions under drought conditions.

Develop methods for in situ manipulation of the plant and associated-microbiome to mitigate the detrimental effects of drought on crop productivity.

Microorganisms exhibit remarkable resilience to harsh environments, triggering various signaling cascades and metabolic processes that enable them to swiftly adapt to changing conditions. The soil microbiome, with its manifold benefits for plants, has the ability to mitigate the adverse impact of drought on crops (Anli et al., 2020; Boutasknit et al., 2020; Ben-Laouane et al., 2021; Meddich et al., 2021). For instance, certain ectomycorrhizal fungal (EMF) species in the rhizosphere have been found to colonize the roots of specific pinyon pine genotypes, enhancing their drought resistance (Lau and Lennon, 2012). The varying drought tolerance among different genotypes can be attributed to the presence of these EMF communities, which represent an extended genetic repertoire of the host plant and may serve as a key strategy for drought resilience. The association between host plant and diverse bacterial/fungal communities, possessing potent metabolic and biogeochemical functions, can facilitate plant adaptation to water scarcity. Through long-term field and greenhouse studies coupled with microbial community sequencing, researchers have uncovered links between host-determined EMF communities and disparities in plant performance. Moreover, they have discovered that drought-resistant genotypes exhibit more intense colonization by Geospora EMF species. Adaptation has been a subject of extensive discussion in evolutionary biology and ecology. Gehring et al. (2017) revealed that the adaptation does not necessarily involve drastic changes in phenotype but rather subtle variations that affect how the host forms associations with microscopic rhizospheric fungi. Many questions remain unanswered; (1) What are the host traits that control specific interactions with microbial communities? Are they morphological, chemical, or even phenological? Speculation can be made regarding the involvement of genes and chemical signaling molecules that underlie host-mycorrhizal interactions, (2) How significant are plant-associated soil microbial communities in drought adaptation? The fact that genotypic difference in drought resistance were observed only in pinyon pine associated with microbes suggests that the plant’s ability to form specific microbial associations is the primary driver of drought resistance.

Given that host genotypes shape distinct soil microbial communities that perform various biogeochemical and metabolic functions (Edwards et al., 2015; Wagner et al., 2016), the potential adaptation of plant traits controlling plant-microbe interactions becomes significant. A comprehensive understanding of host-microbiome interactions at biochemical and genomic levels will enhance our knowledge of drought adaptation and contribute to improved models of host responses to water deficiency. Recognizing that genotypes exhibit differential interactions with aboveground and belowground microbes may revolutionize the strategies employed to enhance plant agronomic traits and sustain global food security in the face of increasing drought trends.