Use of FTA® filter paper for the molecular detection of Newcastle disease virus

The feasibility of using Flinders Technology Associates filter papers (FTA cards) to collect allantoic fluid and chicken tissue samples for Newcastle disease virus (NDV) molecular detection was evaluated.Trizol RNA extraction and one-step reverse transcriptase-polymerase chain reaction (RT-PCR) were used.FTA cards allowed NDV identification from allantoic fluid with a titre of 10 5.8 median embryo lethal doses/ml.The inactivated virus remained stable on the cards for 15 days.NDV was detected from FTA imprints of the trachea, lung, caecal tonsil and cloacal faeces of experimentally infected birds.RT-PCR detection from FTA cards was confirmed by homologous frozen-tissue RT-PCR and virus isolation.Direct nucleotide sequence of the amplified F gene allowed prediction of NDV virulence.No virus isolation was possible from the FTA inactivated samples, indicating viral inactivation upon contact.The FTA cards are suitable for collecting and transporting NDV-positive samples, providing a reliable source of RNA for molecular characterization and a hazard-free sample.Virus.LaSota NDV strain vaccine (Merial Select, Inc., Gainesville, Georgia.USA) was propagated by inoculation into embryonating chicken eggs, as previously described (Senne, 1998).The AF was collected and tested using rapid plate haemagglutination of 5% chicken red blood cells (Alexander, 1998).Virus titration was performed as previously described (Villegas, 1998b).The titre obtained was 10 8.8 median egg lethal doses (ELD 50 )/ml.The AF stock virus was stored at (/808C until needed.