Modulation of biofilm-formation in Salmonella enterica serovar Typhimurium by the periplasmic DsbA/DsbB oxidoreductase system requires the GGDEF-EAL domain protein STM3615.

Naeem Anwar,Ute Römling,Syed Fazle Rouf,Mikael Rhen,Michael Hensel

doi:10.1371/journal.pone.0106095

Naeem Anwar, Ute Römling + Show 3 more

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https://doi.org/10.1371/journal.pone.0106095

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Abstract

In Salmonella enterica serovar Typhimurium (S. Typhimurium), biofilm-formation is controlled by the cytoplasmic intracellular small-molecular second messenger cyclic 3′, 5′-di- guanosine monophosphate (c-di-GMP) through the activities of GGDEF and EAL domain proteins. Here we describe that deleting either dsbA or dsbB, respectively encoding a periplasmic protein disulfide oxidase and a cytoplasmic membrane disulfide oxidoreductase, resulted in increased biofilm-formation on solid medium. This increased biofilm-formation, defined as a red, dry and rough (rdar) colony morphotype, paralleled with enhanced expression of the biofilm master regulator CsgD and the biofilm-associated fimbrial subunit CsgA. Deleting csgD in either dsb mutant abrogated the enhanced biofilm-formation. Likewise, overexpression of the c-di-GMP phosphodiesterase YhjH, or mutationally inactivating the CsgD activator EAL-domain protein YdiV, reduced biofilm-formation in either of the dsb mutants. Intriguingly, deleting the GGDEF-EAL domain protein gene STM3615 (yhjK), previously not connected to rdar morphotype development, also abrogated the escalated rdar morphotype formation in dsb mutant backgrounds. Enhanced biofilm-formation in dsb mutants was furthermore annulled by exposure to the protein disulfide catalyst copper chloride. When analyzed for the effect of exogenous reducing stress on biofilm-formation, both dsb mutants initially showed an escalated rdar morphotype development that later dissolved to reveal a smooth mucoid colony morphotype. From these results we conclude that biofilm-development in S. Typhimurium is affected by periplasmic protein disulphide bond status through CsgD, and discuss the involvement of selected GGDEF/EAL domain protein(s) as signaling mediators.

Highlights

Infections with Salmonella enterica (S. enterica) represent a major health problem and a significant burden on food industry [1]
Typhimurium affect rdar morphotype development through selected GGDEF/EAL domain proteins such as CsgD, and as the elevated rdar morphotype development could be downregulated by the c-di-GMP phosphodiesterases YhjH in both the DdsbA or DdsbB mutant, we investigated to identify GGDEF/EAL domain protein(s) are required for the up-regulated rdar morphotype in DdsbA/B mutant
We demonstrate a role of the periplasmic disulfide bond system (Dsb) oxidoreductase system in biofilm generation of S

Summary

Introduction

Infections with Salmonella enterica (S. enterica) represent a major health problem and a significant burden on food industry [1]. DdsbB ly, sterile filter paper discs soaked with 1M CuCl2 were put at the very edge of bacterial inoculum streaked on CR plates. We tested the reductant dithiothreitol (DTT) at a 5 mM final non-lethal concentration in the CR plates, after which we followed rdar morphotype development. When grown on such plates, the dsb mutants again showed enhanced rdar morphotype development at day one post inoculation, at which time the wild type strain still exhibited a rather smooth colony morphotype (Fig. 5A). At the second day post inoculation, both dsb mutants had lost their rdar morphotype, and converted into a highly mucoid morphotype three days post inoculation (Fig. 5A)

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