Liang Weifeng, Zhou Guohui, Liu Wenming, Yang Baolin, Li Chaosi, Wang Haiwei, Yang Decheng, Ma Wenge, Yu Li
Division of Livestock Infectious Diseases, State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, No. 427 Maduan Street, Harbin, 150001, People's Republic of China.
Institute of Veterinary Medicine, Xinjiang Academy of Animal Science, 151 Eastern Kelamayi Street, Ürümqi, 830000, People's Republic of China.
Arch Virol. 2016 Oct;161(10):2705-16. doi: 10.1007/s00705-016-2984-7. Epub 2016 Jul 15.
Foot-and-mouth disease (FMD), caused by foot-and-mouth disease virus (FMDV), is a highly contagious infectious disease that affects domestic and wild cloven-hoofed animals worldwide. In recent years, a series of outbreaks of serotype A FMD have occurred in many countries. High-affinity neutralizing antibodies against a conserved epitope have the potential to provide protective immunity against diverse subtypes of FMDV serotype A and to protect against future pandemics. In this study, we produced an A serotype FMDV-specific monoclonal antibody (MAb) against the viral capsid protein VP1, designated 9A9, that potently neutralized FMDV A/JLYS/CHA/2014 with a 50 % neutralization titer (NT50) of 4,096. GST-fusion proteins expressing truncated peptides of VP1 were subjected to Western blot analysis using MAb 9A9, and it was found that the peptide (143)RGDLGPLAARL(153) of VP1 was the minimal epitope for MAb 9A9 binding. Western blot analysis also revealed that the epitope peptide could be recognized by positive sera from serotype A FMDV-infected pigs and cattle. Subsequent alanine-scanning mutagenesis analysis revealed that residues Gly(147) and Leu(149) of the 9A9-recognized epitope are crucial for MAb 9A9 binding. Furthermore, under immunological pressure selected by MAb 9A9, a single amino acid residue replacement (L149P) occurred in a viral neutralization-escape mutant, which verified the location of a critical residue of this epitope at Leu(149). Importantly, the epitope (143)RGDLGPLAARL(153) was highly conserved among different topotypes of serotype A FMDV strains in sequence alignment analysis. Thus, the results of this study could have application potential in the development of epitope-based vaccines and a suitable MAb-based diagnostic method for detection of type A FMDV as well as quantitation of antibodies against FMDV serotype A.
口蹄疫（FMD）由口蹄疫病毒（FMDV）引起，是一种高度传染性的传染病，影响着全球范围内的家养和野生偶蹄动物。近年来，许多国家发生了一系列A型口蹄疫疫情。针对保守表位的高亲和力中和抗体有可能提供针对A型FMDV不同亚型的保护性免疫，并预防未来的大流行。在本研究中，我们制备了一种针对病毒衣壳蛋白VP1的A型FMDV特异性单克隆抗体（MAb），命名为9A9，它能有效中和FMDV A/JLYS/CHA/2014，50%中和滴度（NT50）为4096。使用MAb 9A9对表达VP1截短肽的GST融合蛋白进行了蛋白质印迹分析，发现VP1的肽段（143）RGDLGPLAARL（153）是MAb 9A9结合的最小表位。蛋白质印迹分析还表明，该表位肽可被A型FMDV感染猪和牛的阳性血清识别。随后的丙氨酸扫描诱变分析表明，9A9识别表位的甘氨酸（Gly）（147）和亮氨酸（Leu）（149）残基对MAb 9A9结合至关重要。此外，在MAb 9A9选择的免疫压力下，病毒中和逃逸突变体中发生了单个氨基酸残基替换（L149P），这证实了该表位关键残基位于亮氨酸（Leu）（149）处。重要的是，在序列比对分析中，表位（143）RGDLGPLAARL（153）在A型FMDV菌株的不同拓扑型中高度保守。因此，本研究结果在基于表位的疫苗开发以及用于检测A型FMDV和定量抗A型FMDV抗体的合适的基于单克隆抗体的诊断方法方面可能具有应用潜力。
