Improving safety of a live attenuated Edwardsiella ictaluri vaccine against enteric septicemia of catfish and evaluation of efficacy.

Edwardsiella ictaluri is a Gram-negative facultative intracellular pathogen causing enteric septicemia of channel catfish (ESC). Our recent work indicated that tricarboxylic acid cycle and one-carbon metabolism are critical pathways for E. ictaluri virulence. Although single and double gene deletions in these pathways resulted in safe and efficacious vaccines for use in catfish fingerlings, vaccine trials in catfish fry showed safety concerns. Therefore, we aimed to improve the safety of these mutants by constructing two triple mutant combinations. ESC-NDKL1 (ΔgcvPΔsdhCΔfrdA) was constructed by introducing an in-frame deletion of frdA in a gcvP-sdh mutant. ESC-NDKL2 (ΔgcvPΔsdhCΔmdh) was constructed in a similar manner. ESC-NDKL1 strain was a better vaccine candidate compared to ESC-NDKL2, providing better safety and efficacy in catfish fry and catfish fingerlings. Field trials in earthen ponds under three vaccination conditions showed that survival was significantly higher in catfish vaccinated with ESC-NDKL1 by immersion at the fry stage, oral vaccination in ponds, and fry immersion-pond oral combination (86.74%, 81.67%, and 95.22%, respectively) compared to sham-vaccinated (42.75%), and Aquavac-ESC fry immersion vaccinated (61.51%) catfish. Our findings indicate that ESC-NDKL1 is a good candidate for further development as a vaccine for ESC.