Functional characterization of VscCD, an important component of the type III secretion system of Vibrio harveyi

Vibrio harveyi is a Gram-negative bacterium that occurs widely in the ocean and a kind of pathogenic bacteria associated with vibriosis in grouper. We investigated whether the VscCD protein of the type III secretion system (T3SS) was important for pathogenicity of V. harveyi. Mutations to the vscC and vscD genes (Delta vscCD) and complementation of the Delta vscCD mutant (C-Delta vscCD) were created. Moreover, the biological characteristics of the wild-type (WT) and mutant strains of V. harveyi 345 were compared. The results showed that deletion of the vscCD genes had no effect on bacterial growth, swimming/swarming ability, secretion of extracellular protease, or biofilm formation. However, as compared with the V. harveyi 345: pMMB207 (WT+) and complementary (C Delta vscCD) strains, the Delta vscCD: pMMB207 (Delta vscCD+) mutant displayed decreased resistance to acid stress, H2O2, and antibiotics. In addition, infection of the pearl gentian grouper (female Epinephelus fuscoguttatus x male Epinephelus lanceolatu) showed that as compared with the WT+ and C-Delta vscCD strains, the Delta vscCD+ strain significantly reduced cumulative mortality of the host. The colonization ability of the Delta vscCD+ mutant in the spleen and liver tissues of the pearl gentian grouper was significantly lower than that of the WT+ and C-Delta vscCD strains. In the early stage of infection with the Delta vscCD+ strain, the expression levels of IL-1 beta, IL-16, TLR3, TNF-alpha, MHC-I alpha, and CD8 alpha were up-regulated to varying degrees. As compared with the WT+ and C-Delta vscCD strains, luxR expression was significantly up-regulated in the Delta vscCD+ strain, while the expression of vcrH and vp1668 was significantly down-regulated. As an important component of the T3SS, VscCD seemed to play a significant role in the pathogenesis of V. harveyi.