# Transposon sequencing reveals Burkholderia gene fitness in a spaceflight-relevant plant-pathogen interaction

## TL;DR

This study explores how a spaceflight-isolated Burkholderia bacterium behaves in tomato roots when challenged by a fungal pathogen, revealing its stress-adaptation mechanisms.

## Contribution

First evaluation of Burkholderia contaminans gene fitness in a spaceflight-relevant plant-pathogen interaction.

## Key findings

- B. contaminans establishes in the tomato root zone but does not promote plant growth in tissue culture.
- The type II secretion system is critical for root zone establishment, and a Nudix hydrolase is important for responding to FOL infection.
- B. contaminans persists in the root zone through stress adaptation rather than direct antifungal activity.

## Abstract

The spaceflight environment imparts unique selective pressures on the plants and microbes of plant growth chambers on the International Space Station (ISS), which generally manifests through genetic signatures associated with a heightened response to stress. Terrestrially, a baseline understanding of the gene fitness response for any plant growth-promoting microbe when in a tripartite relationship with host and pathogen is currently unknown and is important to characterize before closed-environment spaceflight implementation. To that end, this study evaluated the behavior of an ISS plant habitat isolate of Burkholderia contaminans as tomato seeds transitioned to seedlings and assessed gene fitness during challenge with Fusarium oxysporum f. sp. lycopersici (FOL), the causal agent of Fusarium wilt. Using a seed film delivery method vetted for spaceflight, B. contaminans was applied to Solanum lycopersicum cv. Red Robin seeds. Green fluorescent protein (GFP)-tagged B. contaminans was primarily found to localize at the shoot-root junction and was detected on shoots. Upon FOL challenge, B. contaminans population levels remained stable, and despite harboring antifungal and plant growth-promoting capacity, these properties were not conferred in response to FOL in the tissue culture environment. To probe mechanisms underlying the bacterial-fungal interaction between B. contaminans and FOL in the tomato root zone, a genome-wide transposon mutant library was developed for the B. contaminans isolate. Transposon sequencing (Tn-Seq) analysis revealed that the type II secretion system (T2SS) was critical for root zone establishment, whereas a Nudix hydrolase was specifically important for responding to FOL infection and provided further confirmation that antifungal and siderophore-producing gene clusters were not.

This study is the first to evaluate the genetic fitness of a Burkholderia contaminans International Space Station (ISS) isolate in the plant root zone in association with the obligate pathogen Fusarium oxysporum f. sp. lycopersici (FOL). This isolate of B. contaminans establishes in the tomato root zone, does not confer plant growth promotion in tissue culture, but is persistent in the tomato root zone when challenged with FOL through stress-adaptation mechanisms rather than direct antifungal antagonism. The response of B. contaminans in the host root zone when in the presence of the pathogen suggests the microbe is primed to counter stress, which may further confer an advantage in the spaceflight environment.