Diagnosis of Potato Mop-Top Virus in Soil Using Bait Plant Bioassay and RT-PCR-microplate Hybridization

Abstract

Spraing (brown rings or arcs) of potato caused by Potato mop-top virus (PMTV) occurred in a field of Tokachi, Hokkaido, Japan in 2005. To monitor the expansion of spraing-affected areas, we developed a soil diagnostic method that consisted of a bioassay using tomato as bait plant to trap the vector of PMTV, Spongospora subterranea, the causal agent of powdery scab of potatoes, followed by reverse transcription-polymerase chain reaction-microplate hybridization (RT-PCR-MPH) to detect the virus from roots of bait plants. After incubation of tomato seedlings grown with their roots immersed in a soil suspension at 18°C for 9 days, total RNA extracted from bait roots was analyzed by RT-PCR-MPH using PMTV-specific primers and a digoxigenin (DIG)-labeled probe. Soil diagnosis using the present method in an infested area revealed 137 of 224 fields (61.2%) were infested by PMTV although tubers harvested from only one of these fields had spraing.