Pseudomonas chlororaphis Produces Multiple R-Tailocin Particles That Broaden the Killing Spectrum and Contribute to Persistence in Rhizosphere Communities

Abstract

R-tailocins are high-molecular-weight bacteriocins resembling bacteriophage tails. Pseudomonas chlororaphis 30-84 is a plant growth-promoting rhizobacterial (PGPR) strain that produces two distinct R-tailocin particles with different killing spectra. The two R-tailocins have different evolutionary histories but are released by the same lysis cassette. A previous study showed that both tailocins are important for pairwise competition with susceptible rhizosphere-colonizing strains; however, the broader role of tailocins in competition with the native rhizosphere microbiome was not tested. Genomic analysis of the P. chlororaphis 30-84 R-tailocin gene cluster uncovered the presence of three tail fiber genes in the tailocin 2 genetic module that could potentially result in tailocin 2 particles having different tail fibers and thus a wider killing spectrum. In this study, the tail fibers were found to incorporate onto different tailocin 2 particles, each with a distinct killing spectrum. A loss of production of one or both tailocins resulted in decreased P. chlororaphis 30-84 persistence within the wheat rhizosphere when in competition with the native microflora but not bulk soil. The capacity to produce three different versions of a single tailocin, each having one of three different types of tail fibers, is a previously unreported mechanism that leads to a broader R-tailocin killing spectrum. This study also provides evidence for the function of R-tailocins in competition with rhizosphere microbiome communities but not in bulk soil.IMPORTANCE Although R-tailocin gene clusters typically encode one tail fiber protein, three tail fiber-resembling genes were identified in association with one of the two sets of R-tailocin genes within the tailocin cluster of P. chlororaphis 30-84 and other sequenced P. chlororaphis strain genomes. This study confirmed that P. chlororaphis 30-84 not only produces two distinct tailocins, but that one of them is produced with three different types of tail fibers. This is a previously unreported strategy to increase the breadth of strains targeted by an R-tailocin. Our finding that R-tailocins produced by a PGPR Pseudomonas strain enhanced its persistence within the wheat rhizosphere microbiome confirms that R-tailocin production contributes to the population dynamics of rhizobacterial communities.

Keywords: R-tailocins; bacterial ecology; phage tail-like bacteriocins.

FIG 1

Bacterial populations in the wheat rhizosphere after repeated harvest cycles in autoclaved and natural soil. Inoculum of the wild type (WT) and the tailocin baseplate mutants having the empty vector as a control (ΔBP1, ΔBP2, and ΔBP1/2 mutants) or having the vector carrying the complement of the mutation (ΔBP1/pBP1 and ΔBP2/pBP2 mutants) inoculated into natural or autoclaved wheat field soil (Uvalde, TX). (A and B) Plants grown in autoclaved soil (A) or natural soil (B). Data points represent the means from 10 biological replicates, and error bars indicate standard errors. The experiment was repeated twice with similar results. For each harvest cycle, values with different letters are significantly different.

FIG 2

Bacterial populations in the bulk soil after repeated harvest cycles in autoclaved and natural soil. Bacterial populations were collected from the bulk soil from the same repeated harvest assays used for rhizosphere populations (see Fig. 1). (A and B) Bacterial populations in autoclaved soil (A) and natural soil (B). Data points represent the means from 10 biological replicates, and error bars indicate standard errors. The experiment was repeated twice with similar results. For each harvest cycle, values with the same letters are not significantly different.