One Health Genomic Surveillance of Avian and Human Influenza A Viruses Through Environmental Wastewater Monitoring

BackgroundInfluenza A viruses (IAV) are significant pathogens of humans and other animals. Although endemic in humans and birds, novel IAV strains can emerge, jump species, and cause epidemics, like the latest variant of H5N1. Wastewater-based epidemiology (WBE) has very recently been shown to detect human IAV but whether it can detect avian-origin IAV, and if whole genome sequencing (WGS) can be used to discriminate circulating strains of IAV in wastewater remains unknown. MethodsUsing a pan-IAV RT-qPCR assay, six wastewater treatment works (WWTWs) across Northern Ireland (NI), were screened from August to December 2022. A WGS approach using Oxford Nanopore technology was employed to sequence positive samples. Phylogenetic analysis of sequences relative to currently circulating human and avian IAVs was performed. FindingsWe detected a dynamic IAV signal in wastewater from September 2022 onwards across NI. "Meta" whole genome sequences were generated displaying homology to both human and avian IAV strains. The relative proportion of human versus avian-origin IAV reads differed across time and sample site. A diversity in subtypes and lineages was detected (e.g. H1N1, H3N2, and several avian). Avian segment 8 related to those found in recent H5N1 clade 2.3.4.4b was identified. InterpretationWBE affords a means to monitor circulating human and avian IAV strains and provide crucial genetic information. As such WBE can provide rapid, cost-effective, year-round "one-health" IAV surveillance to help control epidemic and pandemic threats. FundingThis study was funded by the Department of Health for Northern Ireland as part of the Northern Ireland Wastewater Surveillance Programme. HighlightsO_LIDynamic IAV RT-qPCR signal in wastewater detected across NI. C_LIO_LINanopore-based WGS reveals presence of both human and avian IAVs in wastewater. C_LIO_LIAvian IAV sequence similarity to gull-associated H13/H16 and recent H5N1 isolates. C_LIO_LICo-detection of distinct clades of human H1N1 and H3N2 subtypes. C_LI Author SummaryInfluenza A virus (IAV) is a major pathogen of humans and other animals and causes regular epidemics and devastating pandemics. Recently, a novel variant of highly-pathogenic H5N1 avian influenza has emerged spreading across the world killing millions of birds and infecting mammals, enhancing its pandemic potential. Strengthening global surveillance systems for human and animal IAV is thus a major priority. Wastewater-based epidemiology (WBE) has been applied to track SARS-CoV-2 and IAV in humans but whether this approach could work for avian IAV is not known. Here, we develop a "one-health" method to survey pan-IAV levels and genetically characterise the viruses. Through this we highlight co-detection of human and avian IAVs in wastewater, with homology to recent H5N1 isolates. Our work demonstrates the potential for WBE to help defend against not only human infections but emerging, zoonotic IAVs of pandemic potential. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=133 SRC="FIGDIR/small/23293833v1_ufig1.gif" ALT="Figure 1"> View larger version (26K): org.highwire.dtl.DTLVardef@a7b3c4org.highwire.dtl.DTLVardef@b2c12org.highwire.dtl.DTLVardef@17a81faorg.highwire.dtl.DTLVardef@1eb5c70_HPS_FORMAT_FIGEXP M_FIG C_FIG