牛源多杀性巴氏杆菌交叉保护性抗原的筛选

Selection of cross-protective antigens from bovine Pasteurella multocida

为筛选牛源多杀性巴氏杆菌(Pm)交叉保护性抗原,本研究以A型Pm CQ2株基因组为模板,对其omp A、omp H、plp E、pm0979、P6和pfh B基因分别进行扩增,克隆于相应表达载体并进行原核表达,同时对表达产物进行纯化及western blot鉴定。将纯化的重组蛋白免疫小鼠,分别测定其对2 LD50剂量的A型和B型Pm的攻毒保护效果。结果表明,重组蛋白r Omp A、r Omp H、r Plp E、r Pm0979、r P6和r Pfh B对A型Pm CQ2株的攻毒保护率分别为20%、60%、70%、40%、20%和10%,对B型Pm CVCC450株的攻毒保护率分别为0、30%、40%、20%、10%和0,表明重组蛋白r Plp E、r Omp H和r Pm0979可以针对不同血清型的Pm提供较好的交叉保护作用。本研究为牛源Pm新型疫苗的研制奠定了基础。

To screen cross protective antigens from bovine Pasteurella multocida, in this study, six genes （ompA, ompH, pipE, pm0979, P6 and pthB） amplified from the genomic DNA of P.multocida capsular serotype A strain CQ2 were cloned into prokaryotic expression vector pET-30a or pET-32a and expressed in E.coli BL21, respectively. The purified recombinant proteins confirmed by westem blot were emulsified with an equal volume of Freund＇s incomplete adjuvant and immunized mice, and then the cross-protections of the recombinant proteins immunized mice were evaluated by challenging with 2 LDs0 bovine P.multocida capsular serotype A strain CQ2 or capsular serotype B strain CVCC450, respectively. The results showed that recombinant proteins of rOmpA, rOmpH, rPlpE, rPm0979, rP6 and rPfhB provided 20%, 60%, 70%, 40%, 20% and 10% cross protection for mice challenging with P.multocida strain CQ2 and 0, 30%, 40%, 20%, 10% and 0 for mice challenging with serotype B strain CVCC450, respectively. In conclusion, these data suggested that these proteins of rPlpE, rOmpH and rPm0979 could be applied as subunits or muti-component vaccine candidates for further development of new cross protective vaccine against bovine P.multocida infections.