Development of a multiplex RT-PCR detection and identification system for Potato spindle tuber viroid and Tomato chlorotic dwarf viroid

Potato spindle tuber viroid (PSTVd) and Tomato chlorotic dwarf viroid (TCDVd) are two closely related Pospiviroids which cause economically important diseases on tomato (Solanum lycopersicum). Until now, however, there have been no molecular diagnostic methods available for discriminating between them except sequencing. We have developed a multiplex reverse transcription-polymerase chain reaction (RT-PCR) system that simultaneously detects and discriminates between both viroids in one reaction. Using this system, amplified cDNAs resulted in a 271 bp PCR product when PSTVd is detected as the template and 191 bp when TCDVd is detected. This multiplex RT-PCR system was used to accurately detect both viroids in field cultivated tomato and petunia (Petunia x hybrida) plants. This is the first finding of PSTVd in field grown tomatoes in Japan.