Developing a platform to monitor Highly Pathogenic Avian Influenza (HPAI) virus sequences in wastewater and environmental samples

Since its introduction to North America in late 2021, highly pathogenic avian influenza (HPAI) H5N1 clade 2.3.4.4b has had serious and widespread effects on wild and domesticated birds. In early 2024, spillover infections were detected for the first time in dairy cattle, leading to an ongoing outbreak affecting ~1000 herds across 16 states. Spillover infections into mammalian hosts like cats, foxes and humans can lead to very severe symptoms including death, although human-to-human transmission is fortunately inefficient at present. To overcome some of the limitations of current approaches to monitor HPAI infections in wild bird, domesticated animal and human populations, we have developed a novel and comprehensive platform to quantify and sequence HPAI viruses from wastewater and environmental samples. Our interdisciplinary team of ornithologists, environmental microbiologists and biochemists have created and optimized a pipeline that (1) identifies productive sites and methods for sampling, (2) efficiently concentrates and extracts viral RNA from complex samples, (3) quantifies viral RNA copies using RT-qPCR and (4) sequences amplicons targeting the genes encoding hemagglutinin, neuraminidase and polymerase subunits. Preliminary results indicate that the HPAI virus sequences detected in the wastewater samples from Detroit, as well as from rural sites frequented by waterfowl share genetic features characteristic of viruses adapted to avian hosts. As the HPAI pandemic continues to develop, we expect that our surveillance platform will play an important role to monitor the evolution of genomic sequences in HPAI viruses associated with higher levels of pathogenicity and transmissibility in avian and mammalian hosts.