Chestley Taeyo, Sroga Patrycja, Nebroski Michelle, Hole Kate, Ularamu Hussaini, Lung Oliver, Nfon Charles
National Centre for Foreign Animal Disease, Canadian Food Inspection Agency, Winnipeg, MB, Canada.
National Veterinary Research Institute, Vom, Plateau State, Nigeria.
Front Vet Sci. 2022 Sep 20;9:977761. doi: 10.3389/fvets.2022.977761. eCollection 2022.
Foot-and-Mouth Disease Virus (FMDV), the causative agent of Foot-and-Mouth Disease, is a highly feared, economically devastating transboundary pathogen. This is due to the virus' extremely contagious nature and its ability to utilize multiple transmission routes. As such, rapid and accurate diagnostic testing is imperative to the control of FMD. Identification of the FMDV serotype is necessary as it provides the foundation for appropriate vaccine selection and aids in outbreak source tracing. With the vast genetic diversity, there is a desperate need to be able to characterize FMDV without relying on prior knowledge of viral serotypes. In this study, the Neptune bioinformatics tool was used to identify genetic signatures specific to each Southern African Territories (SAT) 1, 2 and 3 genomes but exclusionary to the other circulating FMDV serotypes (A, O, Asia1, and the heterologous SAT1, SAT2 and/or SAT3). Identification of these unique genomic regions allowed the design of TaqMan-based real-time reverse transcriptase PCR (rRT-PCR) primer/probe sets for SAT1, SAT2 and SAT3 viruses. These assays were optimized using prototypic FMDV cell culture isolates using the same reagents and thermocycling conditions as the FMDV pan-serotype 3D rRT-PCR assay. Cross-reactivity was evaluated in tandem with the FMDV pan-serotype 3D rRT-PCR utilizing representative strains from FMDV serotypes A, O, Asia1, SAT1, SAT2 and SAT3. The SAT1, SAT2, and SAT3 primer/probe sets were specific for the homologous serotype and exclusionary to all others. SAT1 and SAT3 primer/probe sets were able to detect several topotypes, whereas the SAT2 assay was revealed to be specific for topotype VII. The SAT2 topotype VII specificity was possibly due to the use of sequence data deposited post-2011to design the rRT-PCR primers and probes. Each assay was tested against a panel of 99 bovine tissue samples from Nigeria, where SAT2 topotype VII viruses were correctly identified and no cross-reactivity was exhibited by the SAT1 and 3 assays. These novel SAT1, SAT3 and SAT2 topotype VII rRT-PCR assays have the potential to detect and differentiate circulating FMD SAT viruses.
口蹄疫病毒（FMDV）是口蹄疫的病原体，是一种令人高度恐惧、具有经济破坏性的跨界病原体。这是由于该病毒具有极强的传染性，且能够利用多种传播途径。因此，快速准确的诊断检测对于口蹄疫的防控至关重要。鉴定FMDV血清型是必要的，因为它为选择合适的疫苗提供了基础，并有助于追踪疫情源头。鉴于FMDV存在巨大的遗传多样性，迫切需要能够在不依赖病毒血清型先验知识的情况下对其进行特征描述。在本研究中，使用海王星生物信息学工具来鉴定南非领土（SAT）1、2和3各基因组特有的遗传特征，但排除其他流行的FMDV血清型（A、O、亚洲1型以及异源的SAT1、SAT2和/或SAT3）。鉴定出这些独特的基因组区域后，得以设计出用于SAT1、SAT2和SAT3病毒的基于TaqMan的实时逆转录聚合酶链反应（rRT-PCR）引物/探针组。使用原型FMDV细胞培养分离株，采用与FMDV泛血清型3D rRT-PCR检测相同的试剂和热循环条件对这些检测方法进行了优化。利用FMDV血清型A、O、亚洲1型、SAT1、SAT2和SAT3的代表性毒株，与FMDV泛血清型3D rRT-PCR同时评估交叉反应性。SAT1、SAT2和SAT3引物/探针组对同源血清型具有特异性，对所有其他血清型均具有排他性。SAT1和SAT3引物/探针组能够检测多种拓扑型，而SAT2检测方法显示对拓扑型VII具有特异性。SAT2对拓扑型VII的特异性可能是由于使用了2011年后存入的序列数据来设计rRT-PCR引物和探针。针对来自尼日利亚的一组99份牛组织样本对每种检测方法进行了测试，其中SAT2拓扑型VII病毒被正确鉴定，SAT1和3检测方法未表现出交叉反应。这些新型的SAT1、SAT3和SAT2拓扑型VII rRT-PCR检测方法具有检测和区分流行的FMD SAT病毒的潜力。
