Evaluation of INSeq To Identify Genes Essential for Pseudomonas aeruginosa PGPR2 Corn Root Colonization

Abstract

The reciprocal interaction between rhizosphere bacteria and their plant hosts results in a complex battery of genetic and physiological responses. In this study, we used insertion sequencing (INSeq) to reveal the genetic determinants responsible for the fitness of Pseudomonas aeruginosa PGPR2 during root colonization. We generated a random transposon mutant library of Pseudomonas aeruginosa PGPR2 comprising 39,500 unique insertions and identified genes required for growth in culture and on corn roots. A total of 108 genes were identified as contributing to the fitness of strain PGPR2 on roots. The importance in root colonization of four genes identified in the INSeq screen was verified by constructing deletion mutants in the genes and testing them for the ability to colonize corn roots singly or in competition with the wild type. All four mutants were affected in corn root colonization, displaying 5- to 100-fold reductions in populations in single inoculations, and all were outcompeted by the wild type by almost 100-fold after seven days on corn roots in mixed inoculations of the wild type and mutant. The genes identified in the screen had homology to genes involved in amino acid catabolism, stress adaptation, detoxification, signal transduction, and transport. INSeq technology proved a successful tool to identify fitness factors in P . aeruginosa PGPR2 for root colonization.

Bacterial Analogs of Plant Tetrahydropyridine Alkaloids Mediate Microbial Interactions in a Rhizosphere Model System

“…P. koreensis CI12 and Escherichia coli S17-1λpir with pSAM_BT20 (27) supplemented with ampicillin (100 μg ml −1 ) were first grown individually for 16 h in LB at 28°C and 37°C, respectively, with agitation. Cells were washed and resuspended in LB to an optical density at 600 nm of 2.0.…”

High-throughput identification of genes influencing the competitive ability to obtain nutrients and performance of biocontrol in Pseudomonas putida JBC17

“…The argG (Tn18-D10) gene encodes argininosuccinate synthase, an arginine biosynthetic enzyme in Pseudomonas 46 . Mutation in the argG gene led to reduced fitness of the P. aeruginosa strain PGPR2 during corn root colonization 47 , whereas a mutant of the P. fluorescens strain WCS365 auxotrophic for arginine had reduced root-colonizing ability in tomatoes 48 . These previous reports indicated the importance of arginine in surface attachment, which is likely crucial for biocontrol.…”

The Regulatory Hierarchy Following Signal Integration by the CbrAB Two-Component System: Diversity of Responses and Functions

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