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Genes of two iron-regulated outer membrane proteins of Vibrio parahaemolyticus zj2003, a pathogenic strain isolated from large yellow croaker (Pseudosciaena crocea), psuA and pvuA, were cloned and expressed as N-terminal His6-tagged proteins in Escherichia coli BL21(DE3). The recombinant fusion proteins were purified with nickel chelate affinity chromatography. To analyze the immunogenicity of the proteins, groups of large yellow croaker were immunized with the purified recombinant psuA, pvuA or both, by intraperitoneal injection. Antibody response was assessed by enzyme-linked immunosorbent assay. Titers to the recombinant proteins increased from log2 3.25 to log2 9.80, 4-8 weeks following immunization. The relative percent survival of the groups vaccinated with psuA, pvuA, or a combination of the two, reached 50%, 62.5% and 75%, respectively. Western blot analysis was carried out with the serum from unvaccinated survival fish after infection. Both recombinant proteins were detected, indicating that these two proteins of V. parahaemolyticus zj2003 were immunogenic and could produce synergistic effects during in vivo infection, and they might be considered as important components for developing an aquaculture vaccine against this pathogen.

Vibrio alginolyticus is one of the Vibrio pathogens common to humans and marine animals. During infection and induction of the host immune response, outer membrane proteins of bacteria play an important role. In this study, an outer membrane protein gene (ompW) was cloned from V. alginolyticus and expressed in Escherichia coli. The 645 bp open reading frame (ORF) encodes a protein of 214 amino acid residues with a predicted molecular weight of 23.3 kDa. The amino acid sequence showed a high identity with that of Photobacterium damselae (96.2%) and Vibrio parahaemolyticus (94.4%). The alignment analysis indicated that OmpW was highly conserved. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that the gene was over-expressed in E. coli BL21(DE3). Western blot analysis revealed that the expressed protein had immunoreactivity. The recombinant protein was purified by affinity chromatography on Ni-NTA Superflow resin. Large yellow croaker vaccinated with the purified OmpW showed significantly increased antibody to OmpW, which could resist the infection by V. alginolyticus. A specific antibody was detected by enzyme-linked immunosorbent assay. This study suggested that the conserved OmpW could be an effective vaccine candidate against infection by V. alginolyticus.

Genes of two iron-regulated outer membrane proteins of Vibrio parahaemolyticus zj2003, a pathogenic strain isolated from large yellow croaker (Pseudosciaena crocea), psuA and pvuA, were cloned and expressed as N-terminal His(6)-tagged proteins in Escherichia coli BL(21)(DE(3)). The recombinant fusion proteins were purified with nickel chelate affinity chromatography. To analyze the immunogenicity of the proteins, groups of large yellow croaker were immunized with the purified recombinant psuA, pvuA or both, by intraperitoneal injection. Antibody response was assessed by enzyme-linked immunosorbent assay. Titers to the recombinant proteins increased from log(2) 3.25 to log(2) 9.80, 4-8 weeks following immunization. The relative percent survival of the groups vaccinated with psuA, pvuA, or a combination of the two, reached 50%, 62.5% and 75%, respectively. Western blot analysis was carried out with the serum from unvaccinated survival fish after infection. Both recombinant proteins were detected, indicating that these two proteins of V. parahaemolyticus zj2003 were immunogenic and could produce synergistic effects during in vivo infection, and they might be considered as important components for developing an aquaculture vaccine against this pathogen.

Vibrio alginolyticus is one of the Vibrio pathogens common to humans and marine animals. During infection and induction of the host immune response, outer membrane proteins of bacteria play an important role. In this study, an outer membrane protein gene (ompW) was cloned from V. alginolyticus and expressed in Escherichia coli. The 645 bp open reading frame (ORF) encodes a protein of 214 amino acid residues with a predicted molecular weight of 23.3 kDa. The amino acid sequence showed a high identity with that of Photobacterium damselae (96.2%) and Vibrio parahaemolyticus (94.4%). The alignment analysis indicated that OmpW was highly conserved. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that the gene was over-expressed in E. coli BL21(DE3). Western blot analysis revealed that the expressed protein had immunoreactivity. The recombinant protein was purified by affinity chromatography on Ni-NTA Superflow resin. Large yellow croaker vaccinated with the purified OmpW showed significantly increased antibody to OmpW, which could resist the infection by V. alginolyticus. A specific antibody was detected by enzyme-linked immunosorbent assay. This study suggested that the conserved OmpW could be an effective vaccine candidate against infection by V. alginolyticus.