Use of one step rt-pcr for detection of Asparagus virus 1

Two viruses are responsible for most of the decline in asparagus crops (Asparagus officinalis L.) in the world: Asparagus virus 1 (AV-1) and Asparagus virus 2 (AV-2). AV-1 is a member of genus Potyvirus and has so far been identified in plant and tissue-culture material of asparagus using biological assays on indicator plants, electron microscopy and, rarely, by serodiagnosis. In this study, degenerate potyvirus primers were used in RT-PCR to amplify a cDNA product from an asparagus potyvirus, previously identified as AV-1 on the basis of host range results. RT-PCR products were sequenced, and the nucleotide sequences obtained were aligned with those of other members of the genus Potyvirus to devise primer pairs specific for AV-1. The use of one primer pair in a one-step RT-PCR procedure was found to yield an amplicon with RNA from an AV-1 infected plant, but not with RNA from plants infected with Turnip mosaic virus (TuMV), Pea seed-borne mosaic virus (PSbMV), or Lettuce mosaic virus (LMV). The method was sensitive and reproducible for the detection of AV-1 in leaves and spears of asparagus samples collected from fields and from tissue-culture plantlets.