A comparison of molecular methods for the routine detection of viroids*

Abstract

Viroids, such as Chrysanthemum stunt viroid (CSVd) and Potato spindle tuber viroid (PSTVd), are important plant pathogens. However, because of their unique biological properties, viroids have proved, in the past, difficult to diagnose. The use of molecular methods has now changed this and this paper reports the comparison of three such methods (dot‐blot hybridization using DIG‐labelled cRNA probes, reverse transcription‐polymerase chain reaction (RT‐PCR) and TaqMan), which have been developed for routine detection of CSVd. Sensitivity comparisons show that the TaqMan assay is more sensitive than either RT‐PCR (100 times) and hybridization (1000 times). RT‐PCR and TaqMan assays have also been developed to detect PSTVd. In addition to the development of sensitive detection methods, considerable emphasis has been placed on making these assays amenable to mass‐scale detection through the use of internal controls and the development of a rapid, reliable probe capture extraction system.