(609c) Screening Rhizobacteria Interactions Using a High-Throughput Microwell Array Platform

Authors: 
Hansen, R., Kansas State University
Barua, N., Kansas State University
van der Vlies, A., Kansas State University
Fattahi, N., Kansas State University
Platt, T., Kansas State University
Plant root-associated microbial communities consist of thousands of unique species that function together to offer beneficial properties to the host. These benefits include nutrient uptake, plant hormone regulation, and pathogen protection. The structure and function of these communities is shaped by microbe-microbe interactions, most of which are unknown or poorly characterized because traditional co-culture methods can only examine a few interactions in a single assay. In addition, traditional methods typically probe interactions between colonies or bulk cell populations, overlooking single-cell interactions that often play a pivotal role in community function. To improve these characterization capabilities, we are developing a unique screening tool termed the “microbe array”, designed to discover unknown inter-cellular interactions that exist between plant pathogens and root microbiome members. By implementing previously developed techniques in microfabrication, the platform uses micro-scale wells to form thousands of unique combinations between small numbers of fluorescently-labeled plant pathogens and isolates from a root microbiome. Each well serves as a unique and independent co-culture site, and the growth of the pathogen in each well of the array can be monitored in parallel. This allows for identification of wells where abnormal pathogen growth occurs. We are currently developing materials and methods that enable extraction of cells out of individual wells of interest, allowing for the sequencing and discovery of microbiome members that promote or inhibit pathogen growth. Current studies are focused on screening interactions between Agrobacterium tumefaciens, the agent of Crown Gall Disease, and microbiome members from Heliantus annuus root. Identification of the critical species that inhibit plant pathogen growth and colonization will inform the development of constructed microbial communities for use in new biocontrol strategies. Successful development will also result in a new and powerful screening tool for rapidly uncovering microbe-microbe interactions that occur in any microbiome of interest.