A multiplex RT-PCR for simultaneous detection and identification of five viruses and two viroids infecting chrysanthemum

Pathogens causing significant economic losses in chrysanthemum include tomato aspermy virus (TAV), chrysanthemum virus B (CVB), cucumber mosaic virus (CMV), tobacco mosaic virus (TMV), potato virus Y (PVY), chrysanthemum stunt viroid (CSVd) and chrysanthemum chlorotic mottle viroid (CChMVd). A multiplex reverse transcription polymerase chain reaction (RT-PCR) method, using specific primer sets for each virus or viroid, was developed for simultaneous detection and differentiation of TAV, CVB, CMV, TMV, PVY, CChMVd, and CSVd. The RT-PCR method was validated by testing chrysanthemum samples collected from different regions of China. In this study, CVB, TAV, TMV, PVY, CSVd, CMV, and CChMVd were detected, respectively, in 24.7 %, 17.5 %, 4.4 %, 4.4 %, 2.9 %, 2.5 %, and 1.5 % of the samples tested. These results indicate that CVB and TAV (24.7 % and 17.5 %) are common, whereas CMV, TMV, CChMVd, CSVd, and PVY (all below 5 %) are less frequently encountered. This new multiplex RT-PCR method has potential to be used routinely in large-scale virus and viroid surveys.