Identifying the species-origin of faecal droppings used for avian influenza virus surveillance in wild-birds

Abstract

Background: Avian influenza virus (AIV) surveillance in birds is important for public health. Faecal droppings from wild-birds are more readily available for such studies, but the inability to identify the species-origin of faecal samples limits their value.

Objectives: To develop, optimise, and field-test a method to simultaneously detect AIV and identify the species-origin from faecal samples.

Study design: Analytical sensitivity of the species-identification RT-PCR was assessed on serial dilutions of faecal droppings. Overall sensitivity of the methods for species-identification and AIV detection was assessed on 92 faecal and cloacal samples collected from wildlife, poultry markets, and experimentally H5N1-infected birds.

Results: All 92 samples were correctly identified to 24 different species, with a detection limit of 2.8mug of faecal material. All 20 specimens previously shown by virus culture to be positive for influenza virus were correctly identified by RT-PCR for influenza A using the same nucleic-acid extracts used for species-identification.

Conclusion: We have optimised and evaluated a method for identifying the species of origin and detecting AIV from bird faecal droppings that can be applied to routine surveillance of influenza viruses in wild-birds.

Figure 1

Comparison of the sensitivities of the single (panel A) and nested PCR (panel B) methods: Agarose gel electrophoresis of products from PCR amplification of partial fragments of mitochondrial COI from fecal samples. Serial dilutions were done to determine the minimal amount of initial fecal sample needed for successful species identification. Fecal sample in each reaction was respectively 352μg (representing a dilution of 1/625 of fecal swab material), 70.4μg, 14.1μg, 2.8μg, 0.6μg and 0.1μg. Negative control was distilled water.