一株牛口咽液分离口蹄疫病毒的特性分析

Characteristics of a strain of foot-and-mouth disease virus isolated from bovine oropharyngeal fluid

摘要:

目的 分离、鉴定一株牛源口咽液中的口蹄疫病毒(foot-and-mouth disease virus, FMDV)，分析其抗原蛋白氨基酸变异及增殖特性。方法 采用RT-PCR及基因测序技术鉴定FMDV阳性牛口咽液样本，通过细胞接种从阳性样本中分离病毒，并利用间接免疫荧光法对分离病毒进行鉴定。通过PCR分段扩增及序列测定获得分离FMDV的全基因组，进一步通过序列比对分析分离FMDV的抗原蛋白氨基酸变异，通过蚀斑形成试验和一步生长曲线评价分离FMDV的增殖特性。结果 成功从牛口咽液中分离、鉴定出一株FMDV，命名为O/FMDV/OP/2022/B。该病毒属于O型Ind-2001谱系，其抗原蛋白VP1、VP2和VP3的氨基酸残基发生变异。在BHK-21细胞上，O/FMDV/OP/2022/B毒株形成的蚀斑显著小于疫苗毒株O/FMDV/HN/93的蚀斑，且其一步生长曲线明显低于O/FMDV/HN/93。结论 本研究从牛口咽液中分离出一株FMDV，发现其抗原蛋白氨基酸发生变异，且增殖能力显著低于疫苗毒株O/FMDV/HN/93。该研究有助于从病毒学角度深入理解FMDV持续感染牛口咽组织的机制，并为口蹄疫的防控提供了参考依据。

Abstract:

Objective To isolate and identify a strain of foot-and-mouth disease virus (FMDV) from bovine oropharyngeal fluid (OPF) and subsequently analyze the amino acid mutations in antigenic proteins and the proliferative properties of the isolated virus.Methods RT-PCR and sequencing were performed to identify the FMDV-positive OPF samples. The FMDV strain was isolated from bovine OPF samples by cell inoculation and identified by indirect immunofluorescence assay. The whole genome sequence of the strain was obtained by fragment amplification and sequencing. The amino acid sequences were compared to reveal the amino acid mutations in the antigenic proteins of the strain. The proliferative properties of the isolated FMDV strain were analyzed by the plaque forming assay and one-step growth curve.Results A FMDV strain O/FMDV/OP/2022/B was successfully isolated from bovine OPF, belonging to serotype O and Ind-2001 lineage. Amino acid mutations were identified in the antigenic proteins VP1, VP2, and VP3 of O/FMDV/OP/2022/B. In addition, the plaque of O/FMDV/OP/2022/B in BHK-21 cells was smaller than that of the vaccine strain O/FMDV/HN/93. The one-step growth curves showed that O/FMDV/OP/2022/B had significantly lower replication titer than O/FMDV/HN/93 in BHK-21 cells.Conclusion A strain of FMDV was isolated from bovine OPF, and it displayed amino acid mutations in antigenic proteins and had lower proliferative capacity than the vaccine strain O/FMDV/HN/93. This study contributes to a better understanding of the mechanism of persistent FMDV infection in the bovine oropharynx and provides a reference for the prevention and control of FMD.