GNPS - Elucidation of plant-microbe interactions mechanisms in the rhizosphere

Scientific rationale. The plant microbiome includes beneficial microbiota that provide the plant host protection from pathogens, increased nutrient availability and resilience against abiotic stress. It is well established that the well characterized plant root-colonizing, growth-promoting rhizobacterium Pseudomonas simiae WCS417 conveys benefit to plant host Arabidopsis thaliana, but the mechanisms of this microbially mediated enhancement remains underexplored. In a pilot in vitro investigation, we observed that only providing root exudates in the absence of the plant failed to result in bacterial growth, suggesting that a key component of an in vivo plant-microbe interaction is essential. Here, we propose to use novel plant propagation techniques, namely fabricated ecosystems via ecoBoxes, to facilitate controlled and reproducible plant-microbe interaction over a four-week period. Through metabolomics analyses, we aim to identify the presence/absence of key metabolites observed in experimental treatments.
Specific aim. Experimental characterization of root colonization mechanisms. Col-0 seeds will be surface sterilized and plated on mesh and ½ MS medium in square plates and left at 4oC for approximately 48 hours to allow for stratification. Two to 3-day old seedlings will be added to ecoBoxes and let to grow for 1-2 weeks with 60 ml 1/5 MS media (pH adjusted to ~ 5.7). Pseudomonas simiae WCS417 fluorescent strain, SB642 will be added to ‘with bacterium’ treatment and all samples will be subsequently collected at ~ 4 weeks. Each treatment will have 5 corresponding replicates. Exudate media will be processed for metabolomic analysis (15 samples total) while Arabidopsis will be used for confocal microscopy of plant root colonization (15 plants total).

The work (proposal:https://doi.org/10.46936/10.25585/60008794) conducted by the U.S. Department of Energy Joint Genome Institute (https://ror.org/04xm1d337), a DOE Office of Science User Facility, is supported by the Office of Science of the U.S. Department of Energy operated under Contract No. DE-AC02-05CH11231.