Development of a Novel High-Sensitivity Workflow for Detecting AIV in Aquatic Environments
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High pathogenicity avian influenza viruses (AIV) are a constant worldwide threat to animals and humans. The ongoing spread of the H5N1 virus underlines the pressing need for enhanced monitoring and containment strategies. Here, we report the development and validation of a novel workflow for investigating AIV genes in environmental water. This approach integrates three key components: concentration via QuickConc, extraction using a COPMAN kit, and detection through reverse transcription-preamplification-quantitative PCR. Spike experiments demonstrated that our workflow exhibits a 100-fold increase in sensitivity for AIV gene detection, compared to a combination of hemagglutination assay, QIAamp RNA Blood Mini, and one-step RT-qPCR as a control method. The efficacy of this approach was further corroborated by the successful identification of influenza M and H5 genes in field samples, specifically from surface water volumes not exceeding 500 mL collected from a body of water frequented by migratory birds. This workflow represents a significant advancement in environmental AIV surveillance, potentially augmenting our capacity to monitor and track AIV dynamics in wild bird populations using aquatic ecosystems.