A Multiplex RT-PCR Assay for Detection and Differentiation of Avian-Origin Canine H3N2, Equine-Origin H3N8, Human-Origin H3N2, and H1N1/2009 Canine Influenza Viruses.

Summary

# Editorial Summary Influenza viruses of multiple origins—including equine H3N8, avian-origin canine H3N2, human H3N2, and pandemic H1N1—are now established in canine populations worldwide, creating diagnostic challenges for practitioners investigating respiratory disease in dogs. Wang and colleagues developed a multiplex reverse-transcriptase PCR assay capable of simultaneously detecting and differentiating all four virus subtypes by targeting conserved regions of the hemagglutinin gene, achieving high specificity and sensitivity across all variants. When applied to 420 clinical respiratory samples, the mRT-PCR results correlated perfectly with conventional virus isolation methods, confirming its reliability for field use. For equine professionals, this work is relevant as a benchmark for understanding cross-species influenza surveillance methods and the potential for equine H3N8 circulation beyond equine populations; whilst the assay targets canine disease, improved diagnostic tools across species help establish baseline epidemiological data that inform biosecurity protocols and disease management strategies. The multiplex approach offers a faster turnaround than traditional isolation methods, enabling more rapid identification of circulating strains and potentially supporting evidence-based decisions on vaccination and infection control in mixed-species facilities.

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Key Findings

• •A multiplex RT-PCR assay successfully detected and differentiated four influenza virus subtypes (H1N1/2009, cH3N2, hH3N2, H3N8) with high specificity and sensitivity
• •mRT-PCR results from 420 clinical samples showed 100% consistency with conventional virus isolation methods
• •The assay enables rapid simultaneous identification of circulating canine influenza viruses in clinical diagnostics

Conditions Studied