Prediction and characterization of novel epitopes of serotype A foot-and-mouth disease viruses circulating in East Africa using site-directed mutagenesis.
Prediction and characterization of novel epitopes of serotype A foot-and-mouth disease viruses circulating in East Africa using site-directed mutagenesis.
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DOI：
10.1099/vir.0.000051
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发表时间：
2015-05
期刊：
影响因子：
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通讯作者：
Mahapatra M
中科院分区：
文献类型：
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作者：
Bari FD;Parida S;Asfor AS;Haydon DT;Reeve R;Paton DJ;Mahapatra M
通过单克隆抗体（mAb）逃逸突变体研究，鉴定了口蹄疫病毒（FMDV）衣壳表面的抗原表位，从而确定了血清型A型口蹄疫病毒（FMDV）的四个抗原位点。以前的工作主要集中在从亚洲、欧洲和拉丁美洲分离的病毒上。在这项研究中，我们报告了非洲血清型A型口蹄疫病毒的表位预测和使用反向遗传学检测选择的表位。利用计算机方法分析衣壳序列（n = 56），预测了24个衣壳氨基酸残基具有抗原性，通过将衣壳序列与血清病毒中和数据相关联，预测了6个衣壳氨基酸残基具有抗原性。预测残基分布在表面暴露的衣壳区VP1-VP3上。利用cDNA克隆进行位点定向诱变，产生了涉及7个位点的12个突变病毒，验证了其中8个预测表位残基变化的重要性。用病毒中和试验（VN）评价氨基酸取代对病毒抗原性的影响。VP1-43、VP1-45、VP2-191和VP3-132四个不同位置的突变导致VN滴度显著降低（P值分别为0.05、0.05、0.001和0.05）。据我们所知，这是首次预测含有VP1-43至-45氨基酸（相当于O型血清中的抗原位点3）、VP2-191和VP3-132的抗原区域为A型fmdv的表位并对其进行血清学评估。这鉴定了东非最近流行的血清A型口蹄疫病毒的新的衣壳表位。
Epitopes on the surface of the foot-and-mouth disease virus (FMDV) capsid have been identified by monoclonal antibody (mAb) escape mutant studies leading to the designation of four antigenic sites in serotype A FMDV. Previous work focused on viruses isolated mainly from Asia, Europe and Latin America. In this study we report on the prediction of epitopes in African serotype A FMDVs and testing of selected epitopes using reverse genetics. Twenty-four capsid amino acid residues were predicted to be of antigenic significance by analysing the capsid sequences (n = 56) using in silico methods, and six residues by correlating capsid sequence with serum–virus neutralization data. The predicted residues were distributed on the surface-exposed capsid regions, VP1–VP3. The significance of residue changes at eight of the predicted epitopes was tested by site-directed mutagenesis using a cDNA clone resulting in the generation of 12 mutant viruses involving seven sites. The effect of the amino acid substitutions on the antigenic nature of the virus was assessed by virus neutralization (VN) test. Mutations at four different positions, namely VP1-43, VP1-45, VP2-191 and VP3-132, led to significant reduction in VN titre (P value = 0.05, 0.05, 0.001 and 0.05, respectively). This is the first time, to our knowledge, that the antigenic regions encompassing amino acids VP1-43 to -45 (equivalent to antigenic site 3 in serotype O), VP2-191 and VP3-132 have been predicted as epitopes and evaluated serologically for serotype A FMDVs. This identifies novel capsid epitopes of recently circulating serotype A FMDVs in East Africa.