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目的 近年来，柯萨奇病毒A组6型(CoxsackievirusA6, CVA6)成为国内手足口病(Hand, foot, and mouth disease, HFMD)的主要血清型，2013年起，中国CVA6流行基因型已由D2基因亚型演变成D3a基因亚型，本研究旨在分析这两种基因亚型在乳鼠体内致病力差异及潜在分子机制，为疾病防控与疫苗研发提供依据。方法 研究选取代表性的两株D2和四株D3a基因亚型CVA6毒株，以106半数组织培养细胞感染剂量(50%Tissue Culture Infectious Dose, TCID50)肌肉注射感染1日龄ICR乳鼠，观察六株病毒对小鼠的生存曲线、体重、临床评分，检测乳鼠脑组织病毒滴度，结合组织病理学与免疫组化的结果，分析P1编码区氨基酸序列并预测蛋白结构。结果 D2和D3a基因亚型CVA6在乳鼠脑组织的病毒滴度最高分别为103.875 TCID50和102.25TCID50，存在显著统计学差异(P<0.05);233/HeB/CHN/2011株(D2基因亚型)感染组乳鼠脑组织出现神经元变性坏死、尼氏体丢失及明显炎性反应，同时免疫组化检测显示VP2抗原阳性；26/YC/CHN/2023株(D3a基因亚型)感染组乳鼠脑组织中未见明显损伤，无抗原检出。氨基酸序列比对结果发现，两株病毒在VP2-172、VP3-49和VP1-242等关键位点存在异亮氨酸(I)/缬氨酸(V)替换；单点氨基酸替换后预测其蛋白结构，结果显示，VP1-54和VP1-90位点氨基酸替换改变了VP1环区氢键网络与表面电荷分布，可能影响病毒衣壳结构稳定性、受体亲和力及致病力。结论 本研究结果显示CVA6不同基因亚型(D2与D3a)毒株间的神经致病力差异，并借助蛋白结构预测，筛选出影响病毒致病力的关键氨基酸位点，这些位点可作为CVA6分子流行病学监测与流行风险预警的重要位点。
Abstract：Objective In recent years, Coxsackievirus A6(CVA6) has emerged as the predominant serotype causing hand, foot, and mouth disease(HFMD) in China. Since 2013, the dominant circulating genotype of CVA6 has shifted from the D2 sub-genotype to the D3a sub-genotype. This study aimed to compare the neuropathogenicity of these two sub-genotypes in suckling mice and to explore potential molecular determinants underlying their pathogenic differences, thereby providing a scientific basis for disease control and vaccine development. Methods Two representative CVA6 strains of the D2 sub-genoype and four strains of the D3a sub-genotype were selected. One-day-old ICR suckling mice were infected via intramuscular injection with a dose of 106 50% tissue culture infectious doses(TCID50). Survival rates, body weight changes, and clinical scores were monitored. Viral titers in brain tissues were determined, and neuropathological changes were evaluated by histopathological examination and immunohistochemical analysis. Amino acid sequences of the P1 coding region were compared, and protein structural changes associated with key amino acid substitutions were predicted. Results The peak viral titers of CVA6 D2 and D3a sub-genotypes in suckling mouse brains reached 103.875 TCID50 and 102.25 TCID50, respectively, with a statistically significant difference(P < 0.05). In the 233/HeB/CHN/2011 strain(D2 sub-genotype) infection group, brain tissues exhibited neuronal degeneration and necrosis, loss of Nissl bodies, and pronounced inflammatory responses, accompanied by positive immunohistochemical staining for the VP2 antigen. In contrast, no apparent histopathological lesions or detectable viral antigens were observed in the 26/YC/CHN/2023 strain(D3a sub-genotype) infection group. Amino acid sequence alignment revealed isoleucine(I)/valine(V) substitutions at several key positions, including VP2-172, VP3-49, and VP1-242. Protein structure prediction following single-site amino acid substitutions indicated that substitutions at VP1-54 and VP1-90 altered the hydrogen-bonding network and surface charge distribution within the VP1 loop region, which may affect capsid stability, receptor binding affinity, and viral pathogenicity. Conclusion This study demonstrates marked differences in neuropathogenicity between CVA6 strains of the D2 and D3a sub-genotypes. Through protein structure prediction, key amino acid residues potentially contributing to CVA6 pathogenicity were identified. These sites may serve as important molecular markers for CVA6 molecular epidemiological surveillance and epidemic risk assessment.
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基本信息:
DOI：10.13242/j.cnki.bingduxuebao.250366
中图分类号:R373.23
引用信息:
[1]赵营颖,王蕊,段伟,等.六株不同基因型CVA6对1日龄ICR乳鼠的致病力差异研究[J].病毒学报,2026,42(01):37-50.DOI:10.13242/j.cnki.bingduxuebao.250366.
基金信息:
国家疾病预防控制局公共卫生人才培养支持项目（项目号：GJJKJ-2024-ZY）; 传染病溯源预警与智能决策全国重点实验室（NITFID）资助项目（项目号：ZDGWNLJS25-36）,题目：未知及新发病原体监测及预测预警技术研究; 国家重点研发计划项目（项目号：2021YFC2302003）,题目：病毒监测网络数据标准及数据平台建设~~