Journal of Microbiology and Biotechnology

The Korean Society for Microbiology and Biotechnology (한국미생물·생명공학회)
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The Attenuation Mechanism and Live Vaccine Potential of a Low-Virulence Edwardsiella ictaluri Strain Obtained by Rifampicin Passaging Culture

  • Shuyi Wang (State Key Laboratory of Freshwater Ecology and Biotechnology, Institute of Hydrobiology, Chinese Academy of Sciences) ;
  • Jingwen Hao (State Key Laboratory of Freshwater Ecology and Biotechnology, Institute of Hydrobiology, Chinese Academy of Sciences) ;
  • Jicheng Yang (Dalian Ocean University) ;
  • Qianqian Zhang (State Key Laboratory of Freshwater Ecology and Biotechnology, Institute of Hydrobiology, Chinese Academy of Sciences) ;
  • Aihua Li (State Key Laboratory of Freshwater Ecology and Biotechnology, Institute of Hydrobiology, Chinese Academy of Sciences)
  • Received : 2022.10.11
  • Accepted : 2022.12.02
  • Published : 2023.02.28
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Abstract

The rifampicin-resistant strain E9-302 of Edwardsiella ictaluri strain 669 (WT) was generated by continuous passage on BHI agar plates containing increasing concentrations of rifampicin. E9-302 was attenuated significantly by 119 times to zebrafish Danio rerio compared to WT in terms of the 50% lethal dose (LD50). Zebrafish vaccinated with E9-302 via intraperitoneal (IP) injection at a dose of 1 × 103 CFU/fish had relative percentage survival (RPS) rates of 85.7% when challenged with wild-type E. ictaluri via IP 14 days post-vaccination (dpv). After 14 days of primary vaccination with E9-302 via immersion (IM) at a dose of 4 × 107 CFU/ml, a booster IM vaccination with E9-302 at a dose of 2 × 107 CFU/ml exhibited 65.2% RPS against challenge with wild-type E. ictaluri via IP 7 days later. These results indicated that the rifampicin-resistant attenuated strain E9-302 had potential as a live vaccine against E. ictaluri infection. A previously unreported amino acid site change at position 142 of the RNA polymerase (RNAP) β subunit encoded by the gene rpoB associated with rifampicin resistance was identified. Analysis of the whole-genome sequencing results revealed multiple missense mutations in the virulence-related genes esrB and sspH2 in E9-302 compared with WT, and a 189 bp mismatch in one gene, whose coding product was highly homologous to glycosyltransferase family 39 protein. This study preliminarily explored the molecular mechanism underlying the virulence attenuation of rifampicin-resistant strain E9-302 and provided a new target for the subsequent study of the pathogenic mechanism of E. ictaluri.

Keywords

Acknowledgement

This study was supported by the National Key Research and Development Program of China (No. 2020YFD0900300), the Natural Science Foundation of China (No. 32073023), the Key Project of Scientific & Technological Innovation of Hubei Province (2018ABA101), and Wuhan Science and Technology Project (No. 2019020701011480). We also thank Sichuan Agriculture University and Yangtze River Fisheries Research Institute of Chinese Academy of Fishery Sciences for donating E. ictaluri strains.

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