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目的 探讨北京地区流行的埃可病毒18型（Echovirus 18, E18）基因组、病毒进化及体外感染特征。方法 对2010年3月至2019年10月收集的首都医科大学附属首都儿童医学中心诊断为手足口病、疱疹性咽峡炎等的患儿咽拭子样本通过实时荧光定量PCR(rRT-PCR)进行了通用型肠道病毒（pan-Enterovirus, pan-EV）及肠道病毒A组71型（Enterovirus A71, EV-A71）、柯萨奇病毒A(Coxsackievirus A,CVA)组16型（CVA16）、CVA6及CVA10回顾性筛查；对确定为pan-EV阳性但不能确定血清型的样本，经RT-PCR扩增VP1编码区，通过序列分析进行分型；确定为埃可病毒18型（E18）阳性的样本接种RD细胞，获得分离株，进行宏基因组测序获得全长基因组序列，确定基因型；多方法（RDP4、SimPlot、系统进化树）进行重组事件分析。通过rRT-PCR绘制生长曲线，比较RD与Hep-2细胞中代表株感染特征。结果 7652例样本中，E18阳性17例（0.20%,17/7652）;RD细胞中成功分离到14株，VP1编码区全序列分析显示所有分离株均属C2基因亚型，与我国既往流行株聚为一簇。以s6868为代表株，基因组全长7412 nt，与原型株（Metcalf）核苷酸序列相似性为80.30%,7种算法均提示其在P3编码区（4006-6592 nt）与CVB5重组。增殖实验表明，在RD细胞中E18 RNA拷贝数在感染12h后从lg6.5934±0.1714拷贝/μL增加至lg8.1852±0.2712拷贝/μL，在48h(lg9.3356±0.3829拷贝/μL)达到平台期，高于其在感染Hep-2细胞12h后的lg5.7121±0.1585拷贝/μL。结论 2010-2019年北京地区流行的E18分离株均属于C2基因亚型，s6868毒株在P3编码区与CVB5可能存在重组。E18在RD细胞中的增殖效率高于Hep-2细胞，RD细胞可作为E18体外研究细胞模型。
Abstract：Objective To investigate the genomic features, evolutionary characteristics, and in vitro infection properties of echovirus 18(E18) circulating in Beijing, China. Methods Throat swab specimens were retrospectively collected from pediatric patients diagnosed with hand, foot, and mouth disease(HFMD), herpangina, or related illnesses at the Capital Institute of Pediatrics, Capital Medical University, between March 2010 and October 2019. Samples were screened for pan-enterovirus(pan-EV), EV-A71, coxsackievirus A16(CVA16), CVA6, and CVA10 using real-time reverse transcription PCR(rRT-PCR). Pan-EV-positive samples that could not be serotyped were further subjected to RT-PCR amplification of the VP1 coding region followed by sequencing for genotyping. E18-positive samples were inoculated into RD cells for virus isolation. Full-length genome sequences were obtained by metagenomic sequencing to determine genotypes. Recombination events were analyzed using multiple approaches, including RDP4, SimPlot, and phylogenetic analysis. Viral growth kinetics were assessed by rRT-PCR to compare replication characteristics of a representative strain in RD and Hep-2 cells.Results Among 7,652 clinical specimens, 17 were positive for E18(0.20%, 17/7,652). Fourteen E18 isolates were successfully obtained in RD cells. VP1 sequence analysis demonstrated that all isolates belonged to the C2 subgenotype and clustered with previously reported strains circulating in China. Using strain s6868 as a representative, the complete genome was 7,412 nt in length and shared 80.30% nucleotide identity with the prototype strain Metcalf. Recombination analysis using seven algorithms consistently indicated a potential recombination event with coxsackievirus B5(CVB5) within the P3 coding region(nt 4,006 – 6,592). Replication kinetics showed that in RD cells, E18 RNA copy numbers increased from lg 6.5934 ± 0.1714 copies/μL at 12 h post-infection to lg 8.1852 ± 0.2712 copies/μL, reaching a plateau at 48 h(lg 9.3356 ± 0.3829 copies/μL), which were markedly higher than those observed in Hep-2 cells at 12 h post-infection(lg 5.7121 ± 0.1585 copies/μL). Conclusion E18 strains circulating in Beijing from 2010 to 2019 predominantly belonged to the C2 subgenotype, and the representative strain s6868 may have undergone recombination with CVB5 in the P3 coding region. E18 exhibits more efficient replication in RD cells than in Hep-2 cells, indicating that RD cells are a suitable in vitro model for E18 infection studies.
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基本信息:
DOI：10.13242/j.cnki.bingduxuebao.250332
中图分类号:R373.2
引用信息:
[1]韩振志,贾立平,蔺晨博,等.北京地区埃可病毒18型基因组及细胞感染特征分析[J].病毒学报,2026,42(01):14-22.DOI:10.13242/j.cnki.bingduxuebao.250332.
基金信息:
北京市自然科学基金资助项目（项目号：L242052）,题目：脑炎/脑膜炎患者中肠道病毒病原学及疾病负担研究; 国家自然科学基金青年科学基金项目（项目号：82402599）,题目：埃可病毒11型3A蛋白通过拮抗RNAi抗病毒天然免疫参与脑膜炎发生机制的研究; 首都儿科研究所青年优才专项（项目号：YCYJ-2025-05）,题目：北京市儿童中多血清型肠道病毒B组的共同流行和进化机制研究; 北京市卫生健康委员会高层次公共卫生技术人才建设项目（项目号：学科带头人-02-20）,题目：儿童重症呼吸道感染病毒病原谱变迁及新发病毒病原预警体系建立~~