A Pseudomonas fluorescens type 6 secretion system is related to mucoidy, motility and bacterial competition

Abstract

BackgroundPseudomonas fluorescens strain MFE01 secretes in abundance two Hcp proteins (haemolysin co-regulated proteins) Hcp1 and Hcp2, characteristic of a functional type 6 secretion system. Phenotypic studies have shown that MFE01 has antibacterial activity against a wide range of competitor bacteria, including rhizobacteria and clinically relevant bacteria. Mutagenesis of the hcp2 gene abolishes or reduces, depending on the target strain, MFE01 antibacterial activity. Hcp1, encoded by hcp1, may also be involved in bacterial competition. We therefore assessed the contribution of Hcp1 to competition of P. fluorescens MFE01 with other bacteria, by studying MFE01 mutants in various competitive conditions.ResultsMutation of hcp1 had pleiotropic effects on the MFE01 phenotype. It affected mucoidy of the strain and its motility and was associated with the loss of flagella, which were restored by introduction of plasmid expressing hcp1. The hcp1 mutation had no effect on bacterial competition during incubation in solid medium. MFE01 was able to sequester another P. fluorescens strain, MFN1032, under swimming conditions. The hcp2 mutant but not the hcp1 mutant conserved this ability. In competition assays on swarming medium, MFE01 impaired MFN1032 swarming and displayed killing activity. The hcp2 mutant, but not the hcp1 mutant, was able to reduce MFN1032 swarming. The hcp1 and hcp2 mutations each abolished killing activity in these conditions.ConclusionOur findings implicate type 6 secretion of Hcp1 in mucoidy and motility of MFE01. Our study is the first to establish a link between a type 6 secretion system and flagellin and mucoidy. Hcp1 also appears to contribute to limiting the motility of prey cells to facilitate killing mediated by Hcp2. Inhibition of motility associated with an Hcp protein has never been described. With this work, we illustrate the importance and versatility of type 6 secretion systems in bacterial adaptation and fitness.

Contribution of the Pseudomonas fluorescens MFE01 Type VI Secretion System to Biofilm Formation

“…The expression of hcp2 contributes to the killing activity of MFE01 against many Gram-negative bacteria, whereas hcp1 expression is involved in the immobilisation of prey cells. The competitor population was nevertheless reduced slightly by the double mutant, MFE01 Δhcp1Δhcp2 , suggesting the involvement of another factor in T6SS-mediated MFE01 killing activity [36]. …”

“…Gene knockout was performed, as previously described [36], with the suicide plasmid pAKE604 [37]. In-frame hcp3 disruption, due to the deletion of a central region of about 300 bp, was achieved by PCR with the muta1-hcp3-F and muta2-hcp3-R primers (product of about 650 bp) or the muta3-hcp3-F and muta4-hcp3-R primers (product of about 600 bp) (Table 2).…”

“…The Hcp2 protein has been directly implicated in the killing activity of MFE01. Another Hcp protein, Hcp1, is encoded by hcp1 , mutation of which has pleiotropic effects on the phenotype of MFE01, affecting its mucoidy and motility [36]. However, hcp1 mutation has no effect on bacterial competition during incubation on solid medium.…”

“…fluorescens strain, MFN1032, under swimming and swarming conditions, whereas the hcp1 mutant of MFE01 cannot. Hcp1 appears to reduce the motility of prey cells, to facilitate killing by Hcp2 [36]. …”

The effect of light intensity and shear stress on microbial biostabilization and the community composition of natural biofilms

“…100 However, in contrast to Caulobacter, for Pseudomonas the flagellum is of major importance for surface adhesion as the initial step of biofilm formation. 101 Moreover, Decoin et al 102 demonstrated an expressional link between flagellum compounds, such as flagellin, and the production of antimicrobial agents, and suggested that parallel motility and antibiotic production enhances competitiveness. The well-described antibiotic production by different members of Pseudomonas 103 may effectively suppress bacterial competitors, and might be another explanation for the decreased stability of the biofilm matrix.…”