Construction and evaluation of type III secretion system mutants of the catfish pathogen Edwardsiella piscicida

Abstract

Catfish is the largest aquaculture industry in the United States. Edwardsiellosis is considered one of the most significant problems affecting this industry. Edwardsiella piscicida is a newly described species within the genus Edwardsiella, and it was previously classified as Edwardsiella tarda. It causes gastrointestinal septicemia, primarily in summer months, in farmed channel catfish in the southeastern U.S. In the current study, we adapted gene deletion methods used for Edwardsiella to E. piscicida strain C07–087, which was isolated from a disease outbreak in a catfish production pond. Four genes encoding structural proteins in the type III secretion system (T3SS) apparatus of E. piscicida were deleted by homologous recombination and allelic exchange to produce in-frame deletion mutants ( EpΔ ssaV, EpΔ esaM, EpΔ yscR, and EpΔ escT). The mutants were phenotypically characterized, and virulence and vaccine efficacy were evaluated. Three of the mutants, EpΔ ssaV, EpΔ yscR, and EpΔ esaM, were significantly attenuated compared to the parent strain (P< 0.05), but EpΔ escT strain was not. Vaccination of catfish with the four mutant strains ( EpΔ ssaV, EpΔ esaM, EpΔ yscR, and EpΔ escT) provided significant protection when subsequently challenged with wild type strain. In conclusion, we report methods for gene deletion in E. piscicida and development of vaccine candidates derived from a virulent catfish isolate.