We used wastewater-based epidemiology and amplicon-based long-read high-throughput sequencing for surveillance of enteroviruses (EVs) in Maricopa County, Arizona, Southwest United States. We collected 48 samples from 13 sites in three municipalities between 18 June and 1 October 2020, and filtered (175 mL each; 0.45 µm pore size) and extracted RNA from the filter-trapped solids. The RNA was converted to cDNA and processed through two workflows (Sanger sequencing (SSW) and long-read Illumina sequencing (LRISW)) each including a nested polymerase chain reaction (nPCR) assay. We subjected the ~350 bp amplicon from SSW to Sanger sequencing and the ~1900–2400 bp amplicon from LRISW to Illumina sequencing. We identified EV contigs from 11 of the 13 sites and 41.67% (20/48) of screened samples. Using the LRISW, we detected nine EV genotypes from three species (Enterovirus A (CVA4, EV-A76, EV-A90), Enterovirus B (E14) and Enterovirus C (CVA1, CVA11, CVA13, CVA19 and CVA24)) with Enterovirus C representing approximately 90% of the variants. However, the SSW only detected the five Enterovirus C types. Similarity and phylogenetic analysis showed that multiple Enterovirus C lineages were circulating, co-infecting and recombining in the population during the season despite the SARS-CoV-2 pandemic and the non-pharmaceutical public health measures taken to curb transmission.

中文翻译：

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2020 年 6 月至 10 月美国西南部亚利桑那州马里科帕县三个城市的基于废水的流行病学和长读长测序确定肠道病毒循环

我们使用基于废水的流行病学和基于扩增子的长读长高通量测序来监测美国西南部亚利桑那州马里科帕县的肠道病毒 (EV)。我们在 2020 年 6 月 18 日至 10 月 1 日期间从三个城市的 13 个地点收集了 48 个样本，并过滤（每个 175 毫升；0.45 微米孔径）并从过滤器捕获的固体中提取 RNA。将 RNA 转化为 cDNA 并通过两个工作流程（Sanger 测序 (SSW) 和长读长 Illumina 测序 (LRISW)）进行处理，每个流程都包括嵌套聚合酶链反应 (nPCR) 分析。我们将来自 SSW 的 ~350 bp 扩增子进行 Sanger 测序，并将来自 LRISW 的 ~1900-2400 bp 扩增子进行到 Illumina 测序。我们从 13 个位点中的 11 个和筛选样本的 41.67% (20/48) 中鉴定了 EV contigs。使用 LRISW，我们从三个物种（肠道病毒 A（CVA4、EV-A76、EV-A90）、肠道病毒 B（E14）和肠道病毒 C（CVA1、CVA11、CVA13、CVA19 和 CVA24））中检测到九种 EV 基因型，其中肠道病毒 C 约占 90%的变体。然而，SSW 只检测到五种肠道病毒 C 型。相似性和系统发育分析表明，尽管 SARS-CoV-2 大流行和采取了非药物公共卫生措施来遏制传播，但在本季节，多种肠道病毒 C 谱系仍在人群中传播、共同感染和重组。