CHERRY VIRUS-A - CDNA CLONING OF DSRNA, NUCLEOTIDE-SEQUENCE ANALYSIS AND SEROLOGY REVEAL A NEW PLANT CAPILLOVIRUS IN SWEET CHERRY

The nucleotide sequence (7383 nucleotides) of a newly identified member of the genus Capillovirus, cherry virus A (CVA), was obtained from cDNA clones. The cDNA was generated from dsRNA extracted from plant tissue infected with little cherry virus (LCV). Small amounts of LCV dsRNA served as template nucleic acid and enabled the construction of a library of which, unexpectedly, 7.5% of the recombinant plasmids were specific for CVA. The genome organization of CVA resembles that of apple stem grooving virus (ASGV), the type member of the genus Capillovirus and is composed of a 266 kDa polyprotein (ORF1), a 52 kDa ORF2 located within ORF1 and a poly(A) tail. The 266 kDa ORF1 contains all the elements of a replication-related protein and has high identity with 'Sindbis-like' viruses. The ORF encodes the coat protein (CP) in the C-terminal region. The 52 kDa ORF2 has high identities with the putative viral cell-to-cell movement proteins of capillo- and trichoviruses. The CP was identified in immunoblot analysis and estimated to have a molecular mass of 24 kDa. Antiserum was obtained by expression of antigens as fusion proteins in Escherichia coli. There is significant sequence identity between CVA CP and the corresponding proteins of other capillo- and trichoviruses. However, no serological cross-reaction was obtained in immunoblot analysis with ASGV, apple chlorotic leafspot trichovirus (ACLSV), apple stem pitting virus (ASPV) and cherry mottle leaf virus (CMLV) antisera. Flexuous filamentous CVA virions were identified in extracts of sweet cherry by immunosorbent electron microscopy (ISEM) and decorated with the antiserum to the fusion protein. CVA was identified in three cherry sources of different disease status by ISEM, immunoblot analysis and hybridization to dsRNA. CVA is not closely related to any of the currently described diseases in cherry but it has all the properties of a capillovirus. It is suggested that CVA should be classified as a new member of the genus Capillovirus.