Ren Jian-Le, Zhu Yuan-Mao, Zhou Yue-Hui, Lv Chuang, Yan Hao, Ma Lei, Shi Hong-Fei, Xue Fei
Division of Livestock Infectious Diseases, State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute of Chinese Academy of Agricultural Sciences, No. 427 Maduan Street, Nan Gang District, Harbin 150001, Heilongjiang Province, PR China.
Division of Livestock Infectious Diseases, State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute of Chinese Academy of Agricultural Sciences, No. 427 Maduan Street, Nan Gang District, Harbin 150001, Heilongjiang Province, PR China.
Vet Microbiol. 2015 Jul 9;178(1-2):61-9. doi: 10.1016/j.vetmic.2015.04.016. Epub 2015 Apr 25.
Bovine parainfluenza virus type 3 (BPIV3) is an important respiratory tract pathogen for both young and adult cattle. So far, three genotypes A, B and C of BPIV3 have been described on the basis of genetic and phylogenetic analysis. But fine mapping of epitopes of BPIV3 is scant and the antigenic variations among the three genotypes of BPIV3 have not been reported. Nucleocapsid protein (NP) is the most abundant protein in the virion and highly conserved in BPIV3, which is crucial for the induction of protective immunity in host. To identify antigenic determinants of BPIV3 NP, a panel of monoclonal antibodies (mAbs) was tested against a series of overlapping recombinant NP fragments expressed in Escherichia coli. Firstly, six monoclonal antibodies (mAbs) against NP of the genotype C of BPIV3 (BPIV3c) were generated by using the purified BPIV3c strain SD0835 as immunogen and the recombinant NP of SD0835 as screening antigen. Then three antigen epitopes were identified with the six mAbs. One epitope (91)GNNADVKYVIYM(102) was recognized by mAb 5E5. The mAbs 7G5, 7G8, 7G9, and 7H5 were reactive with another epitope (407)FYKPTGG(413). The third epitope (428)ESRGDQDQ(435) was reactive with mAb 6F8. Further analysis showed that the epitope (91-102 amino acids [aa]) was the most conserved and reactive with mAb 5E5 for all three genotypes of BPIV3 and HPIV3. The epitope (407-413 aa) was relatively conserved and reactive with mAbs 7G5, 7G8, 7G9, and 7H5 for BPIV3a, BPIV3c and HPIV3, but not reactive with BPIV3b. The epitope (428-435 aa) was less conserved and was reactive only with mAb 6F8 for BPIV3a and BPIV3c. These results suggested that there were evident antigenic variations among the three genotypes of BPIV3 and HPIV3. The mAb 6F8 could be used to detect BPIV3a and BPIV3c. The mAbs 7G5, 7G8, 7G9, and 7H5 might be used for differentiate BPIV3a, BPIV3c and HPIV3 from BPIV3b. The mAb 5E5 might be used for detecting all three types of BPIV3 and HPIV3. The results in this study would have potential applications in the development of suitable diagnostic techniques for BPIV3, which was prevalent in China.
牛副流感病毒3型（BPIV3）是幼牛和成年牛重要的呼吸道病原体。迄今为止，基于基因和系统发育分析已描述了BPIV3的三种基因型A、B和C。但BPIV3表位的精细定位很少，且尚未报道BPIV3三种基因型之间的抗原变异。核衣壳蛋白（NP）是病毒粒子中含量最丰富的蛋白，在BPIV3中高度保守，对宿主诱导保护性免疫至关重要。为鉴定BPIV3 NP的抗原决定簇，用一组单克隆抗体（mAb）检测了一系列在大肠杆菌中表达的重叠重组NP片段。首先，以纯化的BPIV3c株SD0835作为免疫原，以SD0835的重组NP作为筛选抗原，产生了6株针对BPIV3基因型C（BPIV3c）NP的单克隆抗体。然后用这6株单克隆抗体鉴定出3个抗原表位。一个表位（91）GNNADVKYVIYM（102）被单克隆抗体5E5识别。单克隆抗体7G5、7G8、7G9和7H5与另一个表位（407）FYKPTGG（413）反应。第三个表位（428）ESRGDQDQ（435）与单克隆抗体6F8反应。进一步分析表明，表位（91-102个氨基酸[aa]）是最保守的，并且对BPIV3和HPIV3的所有三种基因型都与单克隆抗体5E5反应。表位（407-413 aa）相对保守，对BPIV3a、BPIV3c和HPIV3与单克隆抗体7G5、7G8、7G9和7H5反应，但与BPIV3b不反应。表位（428-435 aa）保守性较差，仅对BPIV3a和BPIV3c与单克隆抗体6F8反应。这些结果表明BPIV3和HPIV3的三种基因型之间存在明显的抗原变异。单克隆抗体6F8可用于检测BPIV3a和BPIV3c。单克隆抗体7G5、7G8、7G9和7H5可用于区分BPIV3a、BPIV3c和HPIV3与BPIV3b。单克隆抗体5E5可用于检测所有三种类型的BPIV3和HPIV3。本研究结果在开发适合中国流行的BPIV3诊断技术方面具有潜在应用价值。
