Mutualistic interaction between Herbaspirillum seropedicae and Trichoderma longibrachiatum enhances maize plant growth

The soil biota community is predominantly composed of bacteria and fungi that modulate key processes in the biosphere. Mutualistic microbial interactions represent a promising strategy for developing biotechnological products. We investigated the mutualistic association between the diazotrophic bacterium Herbaspirillum seropedicae and the saprophytic fungus Trichoderma longibrachiatum, assessing structural compatibility, diazotrophic activity, and plant growth promotion. Microscopical evaluation revealed compatible cell-to-cell interaction, characterized by bacterial attachment to and dispersal along fungal hyphae, followed by biofilm formation. In co-culture studies using a nitrogen-free semi-solid medium, the bacterial population increased, accompanied by a 500% increase in acetylene reduction, a 120% increase in nifH gene expression, and a 50% increase in protein content compared to monoculture. Immunogold-labeling coupled with transmission electron microscopy using polyclonal antibodies against nitrogenase (nifH-subunit) and bacterial cell surface domains revealed, respectively, that nitrogenase per bacterial cell was not affected by hyphae presence, but cell-wall surface epitopes were enhanced. Inoculation of maize with this consortium increased shoot biomass by up to 226%, root biomass by up to 250%, and doubled the photosynthetic rate relative to controls. Confocal imaging of maize-inoculated plants confirmed the presence of bacterial aggregates and biofilms on root surfaces and in intercellular spaces, often associated with fungal hyphae and mineral particles, which supports the "fungal highway" model for bacterial dispersion. These findings demonstrate that T. longibrachiatum not only supports H. seropedicae survival and spatial distribution but also increases diazotrophic activity, benefiting plant growth. This work highlights the potential of the targeted bacterial-fungal consortium as a biotechnological tool to improve nutrient acquisition, reduce chemical fertilizer use, and enhance crop productivity in agroecosystems.