Characterization of OmpK, GAPDH and their fusion OmpK-GAPDH derived from Vibrio harveyi outer membrane proteins: their immunoprotective ability against vibriosis in large yellow croaker (Pseudosciaena crocea)

Abstract

Aim: To investigate the immunoprotection of three recombinant proteins derived from the Vibrio harveyi outer membrane proteins (OMPs) OmpK, glyceraldehyde‐3‐phosphate dehydrogenase (GAPDH) and their fusion OmpK–GAPDH as vaccine candidates from vibriosis of large yellow croaker (Pseudosciaena crocea). Methods: The ompK gene, of which the leader sequence was omitted, was fused with the gapdh gene. Three recombinant proteins r‐OmpK, r‐GAPDH and r‐OmpK–GAPDH were expressed and purified. Western blots were carried out to detect the specificity of the antibodies raised against the recombinant proteins; Fish were immunized with recombinant proteins and challenged by native V. harveyi. The immunoresponse to the recombinant proteins were determined by ELISA and phagocytic activity assay. Conclusions: The fusion protein r‐OmpK–GAPDH can afford greater protection against the wild V. harveyi than r‐OmpK or r‐GAPDH alone or their mixture in humoral and cellular immunity, indicating that OmpK and GAPDH could produce a synergistic immunoprotection against vibriosis of large yellow croaker (Pseudosciaena crocea) when fused into OmpK–GAPDH with a linker. Significance and Impact of the Study: It has been realized that a multi‐component OMP antigen can induce a higher frequency of immune effectors than a single OMP. The results presented here bring forth a good suggestion for the subunit vaccine design based on the OMPs of gram‐negative pathogens.

Vaccine efficacy of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) from Edwardsiella ictaluri against E. tarda in tilapia

“…Additionally, GAPDH is a common enzyme that exists in many organisms (Liu et al, 2005). Recent investigations of cell surface proteins of pathogenic streptococci and Gram-negative bacteria identified a cellwall-associated protein as structurally and functionally belonging to the GAPDH family of enzymes (Brassard et al, 2004;Hughes et al, 2002;Liu et al, 2005Liu et al, , 2007Sirover, 1999;Zhang et al, 2007). These studies demonstrated that GAPDH is well conserved among species.…”

“…Only the attenuated vaccine for E. ictaluri is commercially available in the USA. Moreover, recent reports indicated that an OMP, like GAPDH, from E. tarda is an effective vaccine candidate against E. tarda, V. anguillarum, and V. harveyi in fish (Liu et al, 2005Zhang et al, 2007). However, the efficacy of the GAPDH vaccine from E. ictaluri has not been investigated.…”

“…However, little is known about the virulence factors of E. ictaluri responsible for antibody effectiveness (Panangala et al, 2009b). Recent studies have revealed that OMPs of pathogenic bacteria may be effective vaccine candidates for protection against bacterial infections in fish (Kawai et al, 2004;Khushiramani et al, 2007;Li et al, 2010;Mao et al, 2007;Zhang et al, 2007). Notably, OMPs with epitopes exposed on the cell surface, such as fimbriae and porins, are easily recognized as foreign substances by the host immune system (Kawai et al, 2004).…”

“…Notably, OMPs with epitopes exposed on the cell surface, such as fimbriae and porins, are easily recognized as foreign substances by the host immune system (Kawai et al, 2004). Several studies have established that vaccines consisting of immunogenic fractions can induce higher levels of protection in fish (Salati, 1988) and other vertebrates (Zhang et al, 2007) than inactivated whole cell (WC) bacterial vaccines. However, to our knowledge, no polyvalent or cross-protective vaccines that are simultaneously active against E. tarda and E. ictaluri have been reported.…”

“…In group A streptococci, including Streptococcus pyogenes, S. equisimilis, S. agalactiae, S. suis, and S. pneumoniae, and in Staphylococcus aureus, a cell-surface GAPDH is involved in the interaction with, and adherence to, host cells and ligands (Brassard et al, 2004;Maeda et al, 2004;Pancholi and Fischetti, 1992). In the case of E. tarda, Vibrio anguillarum, Streptococcus iniae, Aeromonas hydrophila, and Vibrio harveyi, GADPHs have the ability to induce a protective immune response in fish (Zhang et al, 2007). This study aimed to identify and clone the gene encoding the 37-kDa E. ictaluri GAPDH, compare the nucleotide and amino acid sequences of GAPDH with those of other related bacterial species that infect fish, express the cloned GAPDH in large quantities to purify the protein, and investigate the immunoprotective activity of this recombinant GAPDH (rGAPDH) in tilapia (O. niloticus) in response to challenge with E. tarda.…”