Introduction
Soil, rhizosphere, and the rhizoplane, including also internal root tissue, represent complex ecosystems where various organisms interact and influence positively, neutrally, or negatively each other’s growth, development, and ecological functions. These interactions are essential for ecosystem functioning and have profound implications for plant health and productivity. Microbes in the roots and rhizosphere can directly and indirectly affect plant growth and development by providing several services to both plants and ecosystems, such as facilitating the nutrient acquisition, promoting hormone synthesis, suppressing pathogens, inducing systemic resistance, contributing to the cycling of organic matter, nitrogen fixation, and degradation of pollutants, thus influencing overall soil fertility and ecosystem sustainability. Such microbes can have various degrees of biotic interaction with plants from free-living commensal to plant symbionts, and include different groups, from bacteria to fungi. Within the plant symbionts are rhizobia, a group of nitrogen-fixing bacteria that can establish mutualistic symbiotic associations with leguminous plants. These bacteria reside within specialized structures called nodules on plant roots and convert atmospheric nitrogen into ammonia, which can be utilized by the host plant. Sinorhizobium meliloti , a model rhizobial species, exhibits a broad host range and forms symbiotic associations with leguminous (Fabaceae ) genera such as Medicago ,Melilotus , and Trigonella . S. meliloti possesses peculiar genetic and physiological traits, allowing it to efficiently colonize plant roots, establish nodules, and fix nitrogen . Several studies have shown the large genetic and symbiotic diversity of S. meliloti strains, highlighting the importance of rhizobial genotype x plant genotype partnership for successful symbiotic plant growth promotion and nitrogen fixation . Tough the attention has mainly been focused on discovering the genomic determinants of such symbiotic diversity , recently the focus has shifted to transcriptional variation, as closer proxy to phenotypic differences . Here, for S. meliloti , we detected the transcriptomic signatures of wide genotype x genotype interaction in response to the treatment with the symbiotic inducer luteolin and with plant root exudates from three alfalfa (Medicago sativa L.) varieties. In addition, to decipher the extent of epistatic interaction among the main set of symbiotic genes and the rest of the genome, a hybrid S. meliloti strain was constructed by mobilizing the megaplasmid harbouring most of symbiotic genes .This hybrid strain allowed to discover a large number of unique transcriptional signatures related to intragenomic regulatory interactions which can explain part of strain and plant genotype-specific features of symbiosis.
While many details are known on rhizobia-plant interaction, few studies have investigated the interaction between rhizobia and the other relevant components of the root microbiome, i.e. fungi . Fungi play a crucial role in plant-microbe interactions and soil ecosystem functioning. They form intricate associations with plant as free-living organisms in the rhizosphere or, forming interactions with plant roots as endophytes. Rhizospheric fungi contribute to nutrient cycling by decomposing organic matter, releasing nutrients, and enhancing nutrient uptake by plants . They can also act as biocontrol agents, suppressing plant pathogens and promoting plant growth. Furthermore, rhizospheric fungi influence soil structure and stability, affecting water retention and root penetration. Their interactions with plants and soil make them key players in ecological processes and agricultural sustainability.
Concerning nitrogen-fixing symbiosis, arbuscular mycorrhizal fungi have shown multiple mutualistic effects with S. meliloti and the host plant M. truncatula and extensive effects of microbial mutualists on gene expression. Among the most studied fungi related to plant growth, used also as bioinoculant, Trichoderma spp. hold a special attention. They are a group of filamentous fungi, known for their multifunctional roles in the soil ecosystem. They are widely recognized as biocontrol agents against plant pathogens due to their ability to produce a diverse array of antimicrobial compounds and compete for resources. Trichoderma spp. can also promote plant growth by enhancing nutrient availability, stimulating root development, and inducing systemic resistance. Additionally, they contribute to the decomposition of organic matter, thereby influencing nutrient cycling and soil structure. The biocontrol and growth-promoting properties ofTrichoderma spp. make them valuable components of sustainable agriculture and integrated pest management strategies . Unlike arbuscular mycorrhizal fungi, as R. irregularis , free-living filamentous fungi as Trichoderma can be easily grown invitro and they are also well-established models with available genome sequences, as well as methods for the study of their secretome .
The deciphering of the relationships and molecular signals taking place in the rhizosphere among rhizobia and Trichoderma will then offer in perspective numerous potentialities in providing testable experimental models for understanding and exploiting plant-microbiome interaction and dissecting mutualistic and antagonistic effects down to the molecular level.
However, it is becoming more and more clear that mutualistic effects have a strong intraspecific genotype component, i.e. different combinations of partner’s genotypes may lead to significant changes in the outcome of the association (e.g. quality of the symbiosis in rhizobium-legume association) . No data have been reported so far on genotype-by-genotype interactions between rhizobia and rhizospheric fungi and how this can affect the quality of the symbiosis withM. sativa as host plant .
Here, we report the results of a study aimed at determining the effect different Trichoderma species may have on the symbiotic nitrogen-fixer S. meliloti and the presence of genotype-by-genotype interaction between strains of S. melilotiand species of Trichoderma . By combining microbiological observation, physiological, metabolic, and transcriptomic analyses, we show the existence of rhizobial strain-specific response toTrichoderma species, with the presence of either synergistic, neutral, or antagonistic interactions depending on the rhizobium-Trichoderma combination. These interactions determine changes in gene expression of a large fraction of the rhizobial genomes (up to 23.45 % of total genes) and are also reflected in host plant growth-promoting phenotypes. These results provide novel insights into the well-known mutualism between rhizobia and host legumes, emphasizing the role of the molecular dialogue taking place in the plant rhizosphere. Moreover, evidence presented here give a proof-of-concept that careful analysis of microbial interactions can be key for successful development of community-mimicking bioinoculants (also referred to as synthetic communities) in agriculture.