Wastewater-Based Epidemiology and Long-Read Sequencing to Identify Enterovirus Circulation in Three Municipalities in Maricopa County, Arizona, Southwest United States between June and October 2020

• Main Authors: Temitope O. C. Faleye, Devin A. Bowes, Erin M. Driver, Sangeet Adhikari, Deborah Adams, Arvind Varsani, Rolf U. Halden, Matthew Scotch
• Format: Article
• Language: English
• Published: MDPI AG 2021-09-01
• Series: Viruses
• Subjects: wastewater-based epidemiological monitoring; Enterovirus C; human; high-throughput nucleotide sequencing; Arizona; environmental monitoring
• Online Access: https://www.mdpi.com/1999-4915/13/9/1803

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.