The highly conserved a-enolase stimulats cross-protective immunity against serotype I and II Streptococcus iniae infection in channel catfish (Ictalurus punctatus)

Streptococcus iniae is an important fish pathogen causing substantial losses to aquaculture. The emergence of new serotype isolates brought greater challenge for the control of streptococcosis, thus screening more highly conserved surface antigens is especially important for cross-protective vaccines to control S. iniae infection. In this study, the serotype of S.iniae DGX07 was identified by biochemical tests and random amplified polymorphic DNA (RAPD) assay. The identity/similarity of alpha-enolase (ENO) amino acid sequences and the cross reactions of lysates with rabbit anti-rENO serum were analysed in different aquatic pathogenic bacteria. Furthermore, the vaccines of recombinant ENO (rENO) proteins and inactivated S. iniae with or without MontanideTM ISA 763 AVG (AVG) were prepared to vaccinate channel catfish by intraperitoneal injection. Then, the cross-protective immunity of rENO and immunostimulatory effect of each vaccine were evaluated in channel catfish. The re-sults showed that the serotype of S.iniae DGX07 was serotype I. The ENO amino acid sequences were highly conserved in different serotypes of S.iniae and S. agalactiae. And the rabbit anti-rENO serum could react with various Streptococcus lysates. Moreover, rENO conferred cross-protective immunity against S.iniae serotype I and II infection with the relative percent of survival (RPS) of 45.0% and 44.4% in channel catfish, respectively. Besides, AVG as an oil adjuvant, could increase the antibody levels, the serum lysozyme, ACH50 activity and RPS induced by rENO or inactivated S. iniae vaccines. And the expression levels of IL-1 beta 1, TNF-alpha and MHC II genes were significantly enhanced in AVG + rENO vaccinated channel catfish. Therefore, alpha-enolase had the potential to provide cross-protective immunity against different serotypes of S.iniae infection, and AVG was an intense stimulator to improve the immune response in channel catfish.