Construction and immunological identification of lactic acid bacteria vaccine to Helicobacter pylori HspA

Objective To explore the expression and immune activity of Helicobacter pylori(Hp) heat shock protein A (HspA) in Lactococcus lactis(L.lactis)NZ3900,a food-grade delivery vehicle.Methods NZ3900/pNZ8110-hspA was constructed using gene recombination technique.Growth curves were plotted to study whether the growth of recombinant bacteria was affected after exogenous gene hspA cloned into L.lactis and whether the growth of empty bacteria,empty plasmid bacteria and recombinant bacteria was affected after treatment of nisin at different concentrations.Sodium dodecylsulfate polyacrglaide gel electrophoreses (SDS-PAGE) and tricine SDS-PAGE were used to detect HspA expressed in recombinant bacteria.Western blot was adopted to confirm whether the HspA expressed by transformant bacteria having immunoreactivity.Results The hspA gene was amplified from genome DNA of Hp MEL-Hp27 and then cloned in the E.coli-L.lactis shuttle vector pNZ8110 and transformed into E.coli MC1061.The positive recombinant plasmid was electro-transformed into L.lactis NZ3900.There was no obvious affection observed from the growth curve after exogenous gene hspA cloned into L.lactis NZ3900.The growth of recombinant bacteria was affected after Nisin treatment at different concentrations except 10 ng/ml Nisin treatment,while the growth of empty bacteria,empty plasmid bacteria was not affected by different concentrations of nisin.No HspA strip was observed in SDS-PAGE and tricine SDS-PAGE analyses.Western blot analysis showed that HspA expressed by recombinant bacteria had good immunoreactivity.Conclusion The growth of recombinant bacteria was affected by the expressioin of HspA.