Important role of the type VI secretion system in the virulence of an emergent human pathogen Aeromonas hydrophila: Delineating the mechanism of action of its effectors
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Aeromonas hydrophila causes various human diseases, including gastroenteritis, wound infections and septicemia. In this study, we characterized the new type 6 secretion system (T6SS) from isolate SSU of A. hydrophila and demonstrated its role in bacterial virulence. We have provided evidence that the T6SS is independent of other secretion systems and that the vasH gene is essential for the expression of the genes encoding the T6SS gene cluster. We demonstrated T6SS’s ability to translocate effector proteins into the host cells and also showed that T6SS mutants were less toxic to human and murine cell lines, and more efficiently phagocytosed by macrophages. Importantly, bacterial-host cell interaction was needed for the T6SS to induce cytotoxicity in eukaryotic cells. Based on 2-dimensional gel electrophoresis and mass spectrometric analyses of bacterial supernatants, we identified a member of the VgrG protein family, (VgrG1), containing a vegetative insecticidal protein (VIP-2) domain at its carboxyl-terminal end, as well as the hemolysin co-regulated protein (Hcp). We provided evidence that VgrG1 showed ADP-ribosyltransferase activity associated with its VIP-2 domain. Our data indicated that episomal expression of the vgrG1 gene in HeLa Tet-Off cells disrupted the actin cytoskeleton, followed by a decrease in cell viability and an increase in apoptosis. We also provided evidence that the expression of the hcp gene in HeLa cells resulted in apoptosis of the host cells without changes in their morphology. In addition, we showed that the addition of exogenous recombinant Hcp (rHcp) reduced bacterial uptake by macrophages. These results were substantiated by the in vivo analysis of cytokine profiling and activation of host immune cells after infection with the ÄvasH mutant supplemented with rHcp indicating that this T6SS effector inhibited production of pro-inflammatory cytokines and induced production of immunosuppressive cytokines, such as IL-10 and TGF-â. In summary, we showed that the T6SS represent an important virulence determinant of A. hydrophila.