Immunogenetic Analysis Of The House-keeping Glycolytic Enzymes In Edwardsiella Tarda

Except for metabolic functions,glycolytic enzymes have many biological activities,among of them,glyceraldehyde-3-phosphata dehydrogenase(GAPDH)was widely recognized to protect the host from infection.Recently,our lab also reported that fructose-1,6-bisphosphate aldolases(FBA)has a potential immunological protection against the infection of Edwardsiella tarda.The major aim of this work is to evaluate whether the other 8 glycolytic enzymes of E.tarda have the similar protection efficacy,including:hexokinase(HK),phosphoglucose isomerase(PGI),phosphofructokinase(PFK),triose phosphate isomerase(TPI),phosphoglycerate kinase(PGK),phosphoglycerate mutase(PGM),enolase(ENO)and pyruvate kinase(PK).The genes encoding these enzymes were cloned from E.tarda EIB202,a virulent strain isolated from diseased turbot in 2006 by our lab,then expressed in E.coli and recombinant proteins were finally purified for the following function assay.The potential immune.protection of these proteins was evaluated in animal model,zebrafish.The results showed that PGK,ENO and PGM had a higher relative percent survival(RPS),around 50%,than the others.Furthermore,turbots were used to evaluate the immunological protection effects,and high antibody titers were found in antisera of the vaccinated turbots.The antisera were able to react against the other marine pathogens,implying that these proteins are a kind of broad spectrum immunogen among different pathogens.Expressions of the immune-related genes were analyzed with RT-qPCR to elucidate the immune responses of the zebrafish vaccinated with the recombinant proteins.We found that the adaptive immunity in zebrafish vaccinated with recombinant PGK,ENO and PGM could be activated through Myd88-dependent signal pathway and the levels of IgM were significantly increased.These results showed that,just like GAPDH and FBA,PGK,ENO and PGM are also potential immunogens and could be used as broad-spectrum vaccines.