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Title: Optimal specimen collection and transport methods for the detection of avian influenza virus and Newcastle disease virus

item Spackman, Erica
item PEDERSEN, JANICE - Animal And Plant Health Inspection Service (APHIS)
item McKinley, Enid
item GELB, JACK - University Of Delaware

Submitted to: BioMed Central (BMC) Veterinary Research
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 2/15/2013
Publication Date: 3/15/2013
Citation: Spackman, E., Pedersen, J.C., Mckinley, E.T., Gelb, J. 2013. Optimal specimen collection and transport methods for the detection of avian influenza virus and Newcastle disease virus. BioMed Central (BMC) Veterinary Research. 9(35). DOI: 10.1186/1746-6148-9-35.

Interpretive Summary: Rapid and sensitive detection of avian influenza virus and other respiratory diseases in poultry is vital to maintaining the health of domestic poultry. Current diagnostic tests have excellent specificity and sensitivity, however methods for collecting and transport samples have never been fully optimized. Sample collection and transport are known to be critical for the accurate diagnosis of viral diseases, therefore we evaluated several aspects of sample collection and transport: the type of material used to make the sample collection swab, the broth used to store the swab after sample collection, how much broth to use, and whether broth was necessary. Finally, we evaluated how many samples could be transported in one tube. All of these elements were selected because they are practical to implement in the field and modifications could results in cost savings for the poultry industry. It was found that the optimal method for virus sample collection and transport were flocked nylon swabs in a protein rich, pH balanced broth. Up to 11 samples could be pooled together in one tube without adverse affects. Also, samples can not be transported dry, as virus detection was substantially reduced when broth was not used. This information will result in cost savings for the poultry industry and veterinary diagnostics by improving the accuracy of current tests and by testing more efficiently.

Technical Abstract: Active and passive surveillance for avian influenza virus (AIV) and Newcastle disease virus (NDV) is widespread in commercial poultry worldwide, therefore optimization of sample collection and transport would be valuable to achieve the best sensitivity and specificity possible, and to develop the most accurate and efficient testing programs. A H7N2 low pathogenicity (LP) AIV strain was selected and used as an indicator virus because it is present in lower concentrations in swabbings and thus requires greater sensitivity for detection compared to highly pathogenic (HP) AIV or virulent NDV. Using oro-pharyngeal and cloacal swabs collected from chickens experimentally exposed to the virus, we evaluated the effects of numerous aspects sample collection and transport: 1) swab construction material (flocked nylon, wound/non-flocked nylon, or urethane foam), 2) transport media (brain heart infusion broth [BHI] or phosphate buffered saline [PBS]), 3) media volume (2ml or 3.5ml), 4) transporting the swab wet in the vial or removing the swab prior to transport, or transporting the swab dry with no media, and 5) single swabs versus pooling 5 or 11 swabs per vial. The three most common diagnostic methods for AIV were used with each condition; real-time RT-PCR (rRT-PCR), virus isolation (VI) and commercial antigen detection immunoassays. The effects of pooling 5 or 11 individual swabs on the detection of AIV and NDV by VI and rRT-PCR was also evaluated. Statistically significant differences and consistent trends were observed with some elements; flocked and foam swabs were superior to wound nylon, BHI was better than PBS, and transporting swabs wet was better for virus recovery and detection than transporting them dry. There was no observable difference whether the swab was removed prior to transport or left in the vial. Also, with both AIV and NDV, there was no observed difference in virus detection between pools of 5 and 11 swabs.