Emergence of #Influenza #H1N1pdm09 6B.1 A.5a.2a and 6B.1 A.5a.2a.1 Subclades Leading to Subtyping #Failure in a Commercial Molecular #Assay
Highlights • Impact of genetic evolution in influenza A(H1N1)pdm09 on subtyping assay performance. • Influenza A subtyping assays are susceptible to primer- or probe-binding mismatches. • Subclades 6B.1 A.5a.2a.1 and 6B.1 A.5a.2a harbour mutations that caused subtyping failures in some specimens. • Sequencing confirmed all specimens were H1N1pdm09, within recognized subclades. Abstract Background During the 2023–2024 and early 2024–2025 influenza seasons , several influenza A-positive specimens in our laboratory failed subtyping for H1 , H1pdm09, and H3 using the Allplex Respiratory Panel 1 (Allplex RP1) (Seegene Inc.). This study aimed to identify the cause of these subtyping failures. Materials and Methods Between August 2023 and December 2024 , 23 nasopharyngeal specimens tested positive for influenza A but were unsubtypeable for H1, H1pdm09, and H3. Confirmatory testing by the manufacturer included target-specific PCR for the M and HA genes, followed by seque...