PS 12-170 - Following Fungal Farts: Using random barcoded transposon-site sequencing (Rb-TnSeq) bacterial libraries to explore the effects of volatiles from the filamentous fungus Trichoderma atroviride

Abstract: Plant-associated fungi provide their hosts with many important health related benefits, and can even protect the plant from invading microbial pathogens. Trichoderma atroviride is a plant root-associated filamentous fungus with potent antimicrobial effects, and volatile organic compounds (VOCs) from T. atroviride have been shown to discourage growth of a range of pathogenic microbes. However, few studies have explored how these VOCs may impact beneficial root-associated microbes. We predict that both volatiles and secreted metabolites will significantly shape fungal-bacterial interactions in the rhizosphere.

To test this hypothesis, we used a co-culture method we call a “Thunderdome” to assess how VOCs from T. atroviride impact the physiology of six Plant Growth Promoting Bacteria (PGPB) selected from across the proteobacteria: Azospirillum brasilense Sp245 and Sinorhizobium meliloti 1021 (alpha-proteobacteria), Burkholderia phytofirmans PsJN and Herbaspirillum seropedicae SmR1 (beta), and Klebsiella michiganensis M5al and Pseudomonas simiae WCS417(gamma). We also tested whether the VOCs significantly impacted 15 additional bacterial species that were isolated from the switchgrass rhizosphere.

We found fungal volatiles inhibited the growth of many bacteria, even when HEPES buffer was added to the media to prevent lowering of pH by CO2 or other acidic volatiles. A crystal violet staining assay showed that, in Azospirillum brasilense Sp245, fungal volatiles specifically inhibited biofilm formation. Using bacterial Random Barcode Transposon-site Sequencing (Rb-TnSeq) libraries, we are further investigating which bacterial genes in the different rhizosphere species are relevant for coping with specific VOCs produced by this fungus.

By elucidating the system wide effects of fungal derived VOCs in the rhizosphere, we can begin to design microbially driven strategies to enhance beneficial plant-rhizosphere relationships, and improve overall plant health.