Background: The recently raised awareness of the threat of a new influenza pandemic has stimulated the interest in detection of influenza A viruses in animal secretions. Virus isolation alone is ...unsatisfactory for this purpose because of the inherent limited sensitivity and the lack of host cells that are universally permissive to all influenza A viruses. Previously described PCR methods are more sensitive, but are targeted predominantly at virus strains currently circulating in humans, since the primer sets display considerable numbers of mismatches to animal influenza A viruses.
Methods: Classical virus isolation approaches were compared with an RT-PCR based screening approach for the detection of influenza A virus in cloacal swabs and droppings collected from wild birds.
Results: A new set of primers, based on highly conserved regions in the matrix gene, was designed for single tube reverse transcription PCR for the detection of influenza A viruses from multiple species. This PCR proved to be fully reactive with a panel of genetically diverse virus isolates obtained from birds, humans, pigs, horses and seals and including all known subtypes of influenza A virus. It was not reactive with other RNA viruses tested.
Conclusions: Comparative tests using fecal and cloacal swab samples from birds confirmed that the new PCR is faster and up to 100-fold more sensitive than classical virus isolation procedures. PCR-based pre-testing of specimens enables high throughput screening for influenza A virus in wild animals.
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