The role of VxrB in Vibrio harveyi and evaluation of a VxrB knock-out as a vaccine candidate.

Abstract

Vibrio harveyi is a major bacterial pathogen in marine aquaculture and can cause serious disease outbreaks in fish and invertebrates. Two-component systems help bacteria sense environmental change and adjust gene expression, including genes linked to virulence. VxrB is a response regulator that has been connected to virulence control in other Vibrio species, but its role in V. harveyi is not well defined. In this study, we deleted VxrB in a virulent V. harveyi strain and named the mutant VHΔVxrB. We compared it with the wild-type strain (W-VH) using in vitro assays, gene expression analysis, and infection and vaccination tests in pearl gentian grouper. Deleting VxrB did not change cell morphology by Gram staining or scanning electron microscopy. However, VHΔVxrB grew more slowly and showed reduced hemolytic activity, while swarming motility increased. RT-qPCR showed that VxrB deletion reduced expression of several virulence-associated genes and lowered expression of the type VI secretion system (T6SS) genes hcp and vgrG. In fish, VHΔVxrB was less virulent than W-VH, with an LD50 of 1.45 × 10 CFU/mL compared with 4.88 × 10 CFU/mL for the wild type. We also tested VHΔVxrB as a live attenuated vaccine. Vaccinated fish had higher survival after homologous challenge, with an RPS of 68.75%, and showed no obvious histopathological lesions. Vaccination increased spleen expression of mhc-Iα, myd88, and il-10. Together, these results show that VxrB supports virulence-related programs in V. harveyi and that VHΔVxrB is a promising attenuated vaccine candidate for pearl gentian grouper.