DIFFERENTIATION OF EASTERN AND WESTERN STRAINS OF PEANUT STUNT CUCUMOVIRUS BASED ON SATELLITE RNA SUPPORT AND NUCLEOTIDE-SEQUENCE HOMOLOGY

Two western strains of peanut stunt cucumovirus (PSV), strains W and BV-15, were unable to support the replication of two distinct PSV satellite RNAs (satRNAs), or infectious transcripts from full-length satRNA cDNA clones, in either tobacco or several leguminous host species. Furthermore, these two strains did not support the replication of cucumber mosaic virus satRNAs in tobacco. All 10 eastern PSV strains tested, on the other hand, efficiently supported PSV satRNA replication. Strains PSV-W acid BV-15 can be further differentiated from the eastern strain PSV-ER by Northern hybridization using cloned cDNA probes to PSV-ER RNAs 1, 2 and 3. Whereas the PSV-ER RNA-specific probes did not hybridize to any PSV-W RNAs, the PSV-ER RNA 3 probe hybridized strongly to PSV-BV-15 RNAs 3 and 4, thus supporting the finding that strain BV-15 is serologically closely related to the eastern strains. The PSV-ER RNA 2-specific probe, but not the PSV-ER RNA 1-specific probe, hybridized to the respective RNA of PSV-BV-15. The results of Northern hybridization with strain PSV-BV-15 support the contention that strain BV-15 represents a reassortant between western and eastern strains. The lack of cross-hybridization between PSV-ER and PSV-W RNAs in Northern hybridization at high stringency is consistent with finding that the percentages of nucleotide identity between the RNAs of strains PSV-ER and PSV-W were 75, 73, 74, and 74%, respectively, for RNAs 1, 2, 3, and 4. A procedure based on reverse transcription and the polymerase chain reaction (RT-PCR) was developed that utilized primers specific for PSV-ER RNA 2. Restriction enzyme digestion of the RT-PCR products generated distinct restriction patterns that clearly differentiated western from eastern strains.