百日咳杆菌粘附素对猪源支气管败血波氏杆菌的保护性抗原的筛选

Immunogenicity and protective efficacy of rertactin recombinants against Bordetella bronchiseptica challenge

摘要:

摘要：【目的】本研究通过百日咳杆菌黏附素(PRN)基因的分段克隆表达及其在BALB/c小鼠的主动和被动免疫保护试验筛选PRN中的保护性抗原肽。【方法和结果】利用大肠杆菌进行PRN的完整蛋白、N端和C端多肽及其RI和RII区域多肽（双拷贝）的表达，命名为GST-PRN、GST-PN、GST-PC、GST-2PRI和GST-2PRII。Western blot检测证实5种表达产物均具有良好的反应原性。在主动免疫保护试验中，5种表达产物均能诱导小鼠产生较高的PRN抗体水平；当使用3 LD50的支气管败血波氏杆菌

Abstract:

Abstract: [Objective] In this study the immunogenicity and protective efficacy of 5 pertactin recombinants against Bordetella bronchiseptica (Bb) challenge is shown. [Methods and results] The complete coding sequence (2 040 bp) of the prn gene (PRN) and its fragments, 5’-terminal 1 173 bp fragment (PN), 3’-terminal 867 bp fragment (PC), two copies of region I (654 bp; PRI) in PN, and two copies of region II (678 bp; PRII) in PC, were separately cloned into the prokaryotic expression vector pGEX-KG, and expressed in the Eschierichia coli BL21 (DE3) using induction by IPTG. The recombinant proteins were named GST-PRN, GST-PN, GST-PC, GST-2PRI and GST-2PRII. All 5 recombinant proteins showed immunological reactivity in the Western-blot analysis. Mice, immunized subcutaneously with two doses of the purified proteins mixed with an equal volume of Freund’s adjuvant, produced robust PRN-specific IgG antibody levels. When challenged, 6 of 9 mice in GST-2PRI group and all 9 mice in the other groups survived intranasal challenge with three times the 50% lethal dose (LD50) of virulent Bb HH0809. After challenge with 10 LD50 7/9, 3/9, 6/9, 1/10 and 6/10 of the mice survived. Furthermore, complete protection against intraperitoneal (i.p.) challenge with 10 LD50 of HH0809 was observed in mice that were injected i.p. with 0.5 ml rabbit anti-GST-PRN, GST-PN, GST-PC or GST-2PRII serum. Only 1 of 10 mice survived in the group of mice that received anti-GST-2PRI, and no survivors were noted in the group of mice that received PRN-absorbed rabbit antiserum (0/5). [Conclusion] In this study, we showed the immunogenicity and protective efficacy of 5 pertactin recombinants against Bb challenge, with GST-PC>GST-PN, and GST-2PRII>GST-2PRI. These findings built a good foundation for the further research into highly efficient vaccine against bordetellosis.