Development of an RT-PCR assay to detect genetically divergent wheat streak mosaic virus isolates for plant quarantine inspections in South Korea.

Wheat streak mosaic virus (WSMV), a member of the genus in the family , has been designated as a plant quarantine pathogen in South Korea. Several diagnostic methods can be applied to diagnose viral infections in plants, but polymerase chain reaction and enzyme-linked immunosorbent assay, which can identify viral species with speed and accuracy, are mainly used in Korean plant quarantine. Many variants of different viral species with highly divergent genomic sequences are constantly being discovered by high-throughput sequencing technology. This means that previously established primers may no longer be suitable for diagnostic use. In this study, we developed a reverse transcription polymerase chain reaction assay for detecting WSMV isolates/strains using all of the WSMV sequences available in NCBI GenBank. All 13 primer sets were able to produce amplicons of the expected sizes from WSMV-infected samples. To check whether nonspecific reactions occur, some closely related viruses (one tritimovirus and five potyviruses) and target imported plants (wheat, maize, oat, and proso millet) were tested. Consequently, four primer sets, which did not produce nonspecific bands, were finally selected among the 13 primer sets. Concentration-dependent amplification tests showed that the four primer sets are adequate for use in the diagnosis of WSMV in Korean plant quarantine.