Lin Xiaojuan, Xu Mingyi, Han Yonglin, Ji Feng, Wang Suting, Chen Meng, Wang Mengmeng, Xu Aiqiang, Tao Zexin
Shandong Center for Disease Control and Prevention, Jinan, 250014, China.
Shandong Academy of Occupational Health and Occupational Medicine, Jinan, 250062, China.
Food Environ Virol. 2025 May 27;17(2):32. doi: 10.1007/s12560-025-09646-2.
Since molecular typing method of enterovirus was introduced, many new types have been discovered. However, due to the low epidemic potential, the information on newer enteroviruses remains limited globally. This study aims to investigate the diversity and phylogeny of newer enterovirus types through environmental surveillance utilizing next-generation sequencing (NGS) technology. Sixty-six wastewater samples were collected from two cities in eastern China between 2019 and 2021 and concentrated 100-fold using a negatively charged membrane method. After cell culture, 388 enterovirus isolates representing 14 serotypes were recovered, including Sabin-like poliovirus type 1 (n = 22) and type 3 (n = 57). Concurrently, RNA extraction was performed on all 66 sewage concentrates, and VP1 semi-nested RT-PCR yielded 56 amplicons, which were subsequently subjected to NGS. The NGS analysis identified a total of 33 serotypes, with echovirus 11, coxsackievirus A10, echovirus 18, coxsackievirus B4, and coxsackievirus B5 being the most prevalent, accounting for 29.11%, 9.87%, 8.27%, 8.14%, and 6.10% of the total reads, respectively. Newer identified enterovirus types A76, A89, A90, and C113 were detected in 1 (1.52%), 17 (25.76%), 5 (7.58%), and 6 (9.09%) sewage samples, respectively. Phylogenetic analysis based on the partial VP1 coding region indicated that these local enteroviruses formed distinct lineages separate from previously identified strains. These findings demonstrate that sewage contains enteroviruses with considerable diversity. NGS-based sewage surveillance offers a significant advantage in data output compared to the cell culture method and can be effectively utilized for monitoring newer enterovirus strains.
自肠道病毒的分子分型方法被引入以来，已发现了许多新类型。然而，由于流行潜力较低，全球范围内关于更新型肠道病毒的信息仍然有限。本研究旨在利用下一代测序（NGS）技术通过环境监测来调查更新型肠道病毒类型的多样性和系统发育。2019年至2021年期间，从中国东部两个城市收集了66份废水样本，并使用带负电荷的膜方法浓缩了100倍。细胞培养后，回收了代表14种血清型的388株肠道病毒分离株，包括1型萨宾样脊髓灰质炎病毒（n = 22）和3型（n = 57）。同时，对所有66份污水浓缩物进行了RNA提取，VP1半巢式RT-PCR产生了56个扩增子，随后对其进行了NGS分析。NGS分析共鉴定出33种血清型，其中埃可病毒1,1、柯萨奇病毒A10、埃可病毒18、柯萨奇病毒B4和柯萨奇病毒B5最为常见，分别占总读数的29.11%、9.87%、8.27%、8.14%和6.10%。新鉴定的肠道病毒类型A76、A89、A90和C113分别在1份（1.52%）、17份（25.76%）、5份（7.58%）和6份（9.09%）污水样本中被检测到。基于部分VP1编码区的系统发育分析表明，这些本地肠道病毒形成了与先前鉴定的菌株不同的谱系。这些发现表明污水中含有具有相当多样性的肠道病毒。与细胞培养方法相比，基于NGS的污水监测在数据输出方面具有显著优势，可有效用于监测更新型肠道病毒株。
