Simultaneous Discovery of Positive and Negative Interactions Among Rhizosphere Bacteria Using Microwell Recovery Arrays

Niloy Barua; Ashlee M. Herken; Kyle R. Stern; Sean Reese; Roger L. Powers; Jennifer L. Morrell-Falvey; Thomas G. Platt; Ryan R. Hansen

doi:10.3389/fmicb.2020.601788

Simultaneous Discovery of Positive and Negative Interactions Among Rhizosphere Bacteria Using Microwell Recovery Arrays

Niloy Barua, Ashlee M. Herken, Kyle R. Stern, Sean Reese, Roger L. Powers, Jennifer L. Morrell-Falvey, Thomas G. Platt, Ryan R. Hansen

Molecular and Cellular Imaging

Research output: Contribution to journal › Article › peer-review

17 Scopus citations

Abstract

Understanding microbe-microbe interactions is critical to predict microbiome function and to construct communities for desired outcomes. Investigation of these interactions poses a significant challenge due to the lack of suitable experimental tools available. Here we present the microwell recovery array (MRA), a new technology platform that screens interactions across a microbiome to uncover higher-order strain combinations that inhibit or promote the function of a focal species. One experimental trial generates 10⁴ microbial communities that contain the focal species and a distinct random sample of uncharacterized cells from plant rhizosphere. Cells are sequentially recovered from individual wells that display highest or lowest levels of focal species growth using a high-resolution photopolymer extraction system. Interacting species are then identified and putative interactions are validated. Using this approach, we screen the poplar rhizosphere for strains affecting the growth of Pantoea sp. YR343, a plant growth promoting bacteria isolated from Populus deltoides rhizosphere. In one screen, we montiored 3,600 microwells within the array to uncover multiple antagonistic Stenotrophomonas strains and a set of Enterobacter strains that promoted YR343 growth. The later demonstrates the unique ability of the platform to discover multi-membered consortia that generate emergent outcomes, thereby expanding the range of phenotypes that can be characterized from microbiomes. This knowledge will aid in the development of consortia for Populus production, while the platform offers a new approach for screening and discovery of microbial interactions, applicable to any microbiome.

Original language	English
Article number	601788
Journal	Frontiers in Microbiology
Volume	11
DOIs	https://doi.org/10.3389/fmicb.2020.601788
State	Published - Jan 5 2021

Funding

This manuscript has been released as a pre-print at BioRxiv, (Barua et al., 2020). We thank Priscila Guzman (Divsion of Biology, Kansas State Univeristy) for making the C58 and PAO1 strains used in experiments. Nanofabrication was conducted at Nebraska Nanoscale Facility at University of Nebraska Lincoln. Funding. We acknowledge support from the National Science Foundation (Award Number 1650187) and the U.S. Department of Energy (DE-SC0018579). A portion of this research was sponsored by the Genomic Science Program, U.S. Department of Energy, Office of Science, Biological and Environmental Research, as part of the Plant Microbe Interfaces Scientific Focus Area (http://pmi.ornl.gov). Oak Ridge National Laboratory is managed by UT-Battelle LLC, for the U.S. Department of Energy under contract DE-AC05-00OR22725. We acknowledge support from the National Science Foundation (Award Number 1650187) and the U.S. Department of

Funders	Funder number
Plant Microbe Interfaces Scientific Focus Area
U.S. Department of
National Science Foundation	1650187
U.S. Department of Energy	DE-SC0018579
Office of Science
Biological and Environmental Research
Oak Ridge National Laboratory
UT-Battelle	DE-AC05-00OR22725

Keywords

consortia
high throughput screening
microbial communities
microbial interactions
microbiome
microdevice
microwell
plant growth promoting rhizobacteria

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10.3389/fmicb.2020.601788

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title = "Simultaneous Discovery of Positive and Negative Interactions Among Rhizosphere Bacteria Using Microwell Recovery Arrays",

abstract = "Understanding microbe-microbe interactions is critical to predict microbiome function and to construct communities for desired outcomes. Investigation of these interactions poses a significant challenge due to the lack of suitable experimental tools available. Here we present the microwell recovery array (MRA), a new technology platform that screens interactions across a microbiome to uncover higher-order strain combinations that inhibit or promote the function of a focal species. One experimental trial generates 104 microbial communities that contain the focal species and a distinct random sample of uncharacterized cells from plant rhizosphere. Cells are sequentially recovered from individual wells that display highest or lowest levels of focal species growth using a high-resolution photopolymer extraction system. Interacting species are then identified and putative interactions are validated. Using this approach, we screen the poplar rhizosphere for strains affecting the growth of Pantoea sp. YR343, a plant growth promoting bacteria isolated from Populus deltoides rhizosphere. In one screen, we montiored 3,600 microwells within the array to uncover multiple antagonistic Stenotrophomonas strains and a set of Enterobacter strains that promoted YR343 growth. The later demonstrates the unique ability of the platform to discover multi-membered consortia that generate emergent outcomes, thereby expanding the range of phenotypes that can be characterized from microbiomes. This knowledge will aid in the development of consortia for Populus production, while the platform offers a new approach for screening and discovery of microbial interactions, applicable to any microbiome.",

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Simultaneous Discovery of Positive and Negative Interactions Among Rhizosphere Bacteria Using Microwell Recovery Arrays

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