A genome-wide screen identifies genes in rhizosphere-associated pseudomonas required to evade plant defenses

Abstract

Pseudomonas fluorescens and related plant root (“rhizosphere”)-associated species contribute to plant health by modulating defenses and facilitating nutrient uptake. To identify bacterial fitness determinants in the rhizosphere of the model plant Arabidopsis thaliana, we performed a high-throughput transposon sequencing (Tn-Seq) screen using the biocontrol and growth-promoting strain Pseudomonas sp. WCS365. The screen, which was performed in parallel on wild-type and immuno-compromised Arabidopsis plants, identified 231 genes that increased fitness in the rhizosphere of wild-type plants. A subset of these genes decreased fitness in the rhi-zosphere of immunocompromised plants. We hypothesized that these genes might be involved in avoiding plant defenses and verified 7 Pseudomonas sp. WCS365 can-didate genes by generating clean deletions. We found that two of these deletion mutants, ΔmorA (encoding a putative diguanylate cyclase/phosphodiesterase) and ΔspuC (encoding a putrescine aminotransferase), formed enhanced biofilms and in-hibited plant growth. We found that mutants ΔspuC and ΔmorA induced pattern-triggered immunity (PTI) as measured by induction of an Arabidopsis PTI reporter and FLS2/BAK1-dependent inhibition of plant growth. We show that MorA acts as a phosphodiesterase to inhibit biofilm formation, suggesting a possible role in biofilm dispersal. We found that both putrescine and its precursor arginine promote biofilm formation that is enhanced in the ΔspuC mutant, which cannot break down pu-trescine, suggesting that putrescine might serve as a signaling molecule in the rhizo-sphere. Collectively, this work identified novel bacterial factors required to evade plant defenses in the rhizosphere. IMPORTANCE While rhizosphere bacteria hold the potential to improve plant health and fitness, little is known about the bacterial genes required to evade host immu-nity. Using a model system consisting of Arabidopsis and a beneficial Pseudomonas sp. isolate, we identified bacterial genes required for both rhizosphere fitness and for evading host immune responses. This work advances our understanding of how evasion of host defenses contributes to survival in the rhizosphere.