Greenhouse gas methane in soil ecosystem adversely influences legume-rhizobia association in rhizosphere

Methane (CH₄), a potent greenhouse gas, is produced in anoxic zones of soil and released into the atmosphere. While CH₄’s role in global warming is well recognized, its influence on plant–microbe interactions in the rhizosphere is not well understood. This study aimed to evaluate how elevated CH₄ concentrations affect nodulation and nitrogen fixation in legumes, specifically in soybean ( Glycine max ) and pigeon pea ( Cajanus cajan ). A controlled experiment was conducted with three levels of rhizospheric CH₄ concentrations: ambient (control), 50 ppm, and 200 ppm. Parameters measured included CH₄ oxidation potential, shoot and root biomass, nodule number, acetylene reduction activity (ARA), and the abundance of functional microbial genes ( pmoA and nifH ). Rhizobial cell growth was also monitored to assess the direct effect of CH₄ on bacterial viability. CH₄ oxidation potential increased from 3.50 to 7.27 ng CH₄ g⁻¹ soil d⁻¹ in soybean. Relative to the ambient control, 50 ppm CH₄ reduced shoot biomass, root biomass, and nodule number by 24%, 26%, and 30%, respectively, in soybean, and by 27%, 23%, and 22%, respectively, in pigeon pea. These reductions were more pronounced at 200 ppm CH₄, reaching 35%, 54%, and 55% in soybean and 40%, 48%, and 45% in pigeon pea, respectively. ARA declined by 30% in soybean and 35% in pigeon pea at 200 ppm CH₄. CH₄ exposure increased pmoA gene abundance by 2.37-fold and reduced nifH by 0.53-fold. Rhizobial growth was unaffected, but nodulation was inhibited by 52% in soybean and 25% in pigeon pea. Elevated CH₄ in the rhizosphere can significantly impair legume–rhizobia symbiosis, reducing nodulation and biological nitrogen fixation.