A multiplex qRT-PCR assay for detection of Influenza A and H5 subtype targeting new SNPs present in high pathogenicity avian influenza Canadian 2022 outbreak strains

H5N1 is a highly pathogenic avian Influenza A subtype that has been known to also infect mammalian hosts and presents a potential public health risk. To address and mitigate the affects of new SNPs, found in recent Canadian outbreaks, on diagnostic detection we developed two qRT-PCR assays by modifying current probes to match sequences detected in the east and west coast of Canada. These assays were multiplexed with a third qRT-PCR assay targeting the M segment, allowing streamlined detection of Influenza A and subtyping for H5. This three-plex qRT-PCR was validated by assessing analytical specificity, limit-of-detection, precision, and accuracy. ### Competing Interest Statement The authors have declared no competing interest. ### Funding Statement This study was funded through British Columbia Centre for Disease Control operational funding sources. ### Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: The Clinical Research Ethics board of the University of British Columbia waived ethical approval for this work. I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines, such as any relevant EQUATOR Network research reporting checklist(s) and other pertinent material, if applicable. Yes All data produced in the present study are available upon reasonable request to the authors.