Proteomics Analysis Reveals Previously Uncharacterized Virulence Factors in Vibrio proteolyticus.

Ann Ray,Lisa N Kinch,Kim Orth,Nick V Grishin,Marcela De Souza Santos,Dor Salomon,Margaret J Mcfall-Ngai

doi:10.1128/mbio.01077-16

Abstract

ABSTRACTMembers of the genus Vibrio include many pathogens of humans and marine animals that share genetic information via horizontal gene transfer. Hence, the Vibrio pan-genome carries the potential to establish new pathogenic strains by sharing virulence determinants, many of which have yet to be characterized. Here, we investigated the virulence properties of Vibrio proteolyticus, a Gram-negative marine bacterium previously identified as part of the Vibrio consortium isolated from diseased corals. We found that V. proteolyticus causes actin cytoskeleton rearrangements followed by cell lysis in HeLa cells in a contact-independent manner. In search of the responsible virulence factor involved, we determined the V. proteolyticus secretome. This proteomics approach revealed various putative virulence factors, including active type VI secretion systems and effectors with virulence toxin domains; however, these type VI secretion systems were not responsible for the observed cytotoxic effects. Further examination of the V. proteolyticus secretome led us to hypothesize and subsequently demonstrate that a secreted hemolysin, belonging to a previously uncharacterized clan of the leukocidin superfamily, was the toxin responsible for the V. proteolyticus-mediated cytotoxicity in both HeLa cells and macrophages. Clearly, there remains an armory of yet-to-be-discovered virulence factors in the Vibrio pan-genome that will undoubtedly provide a wealth of knowledge on how a pathogen can manipulate host cells.

Highlights

Members of the genus Vibrio include many pathogens of humans and marine animals that share genetic information via horizontal gene transfer
Vprh deletion had no effect on bacterial growth, as shown in Fig. S1 in the supplemental material. These results indicate that V. proteolyticus secretes VPRH, a leukocidin-like toxin that mediates actin cytoskeleton rearrangements followed by lysis of the infected host cell
Bacteria that are not known to be pathogenic to humans carry putative virulence determinants

Summary

Introduction

Members of the genus Vibrio include many pathogens of humans and marine animals that share genetic information via horizontal gene transfer. Further examination of the V. proteolyticus secretome led us to hypothesize and subsequently demonstrate that a secreted hemolysin, belonging to a previously uncharacterized clan of the leukocidin superfamily, was the toxin responsible for the V. proteolyticus-mediated cytotoxicity in both HeLa cells and macrophages. We identified a previously unstudied member of the leukocidin family of pore-forming toxins as the virulence factor responsible for the observed cytotoxicity in eukaryotic cells, as well as a plethora of additional putative virulence factors secreted by this bacterium. Analysis of the V. proteolyticus secretome revealed several potential virulence factors, and upon further examination, we identified a secreted hemolysin belonging to the leukocidin superfamily of pore-forming toxins that was required for the V. proteolyticus-mediated cytotoxic effects. Bioinformatic analysis indicated that this secreted toxin represents a previously unstudied clan of the leukocidin pore-forming toxin superfamily and is found in several marine bacteria, including strains pathogenic to humans and marine animals

Methods

Results

Conclusion