Apple mosaic virus (ApMV) has been reported to naturally infect a number of hosts in the Rosaceae family including Rosa spp., Malus spp., and Rubus spp. etc., as well as several hosts such as Humulus spp. and Betula spp. in other plant families (2), but has not been reported to naturally infect Fragaria spp. although it has been grafted into Fragaria spp. (1). In an attempt to develop a detection method for strawberry leafroll as part of an overall strategy to develop diagnostics for all reported virus and virus-like diseases of strawberry, double-stranded RNA (dsRNA) was extracted and cloned as described elsewhere (3) from a single-leafroll-infected plant of Fragaria vesca ‘UC-5’ (CFRA no. 9026 maintained at the USDA-ARS National Clonal Germplasm Repository [NCGR] in Corvallis, OR). Newer leaves on the plant had a chlorotic peacock pattern with a few leaves exhibiting a downward leafroll. The dsRNA pattern suggested possible infection with a crinivirus and an ilarvirus. A database search (BLAST at the National Center for Biotechnology Information) showed that clones with sequence homology to two criniviruses (Strawberry pallidosis associated virus [SPaV] and Beet pseudo-yellows virus [BPYV] as well as ApMV) were obtained from this leafroll-infected plant. Sequences of the three viruses from this plant have been deposited in GenBank (Accession Nos. AY854050, AY854051, and AY854052 for ApMV, BPYV, and SPaV, respectively). The sequence obtained for the strawberry isolate of ApMV represents part of RNA 3 and covers the majority of the coat protein (CP) gene. The nucleotide sequence identity of the CFRA 9026 isolate with the CPs of other ApMV isolates range from 87 to 91% and the amino acid sequence identity ranges from 79 to 88% with similarities ranging from 84 to 94%. Two sets of primers were designed to amplify fragments of ApMV from strawberry (ApMV-str) and one set of degenerate primers to amplify ApMV from several hosts. With the ApMV-str specific primers, we were able to detect ApMV from strawberry but not from rose, whereas with the degenerate primers, we were able to detect ApMV in strawberry and rose. Amplicons obtained with all three sets of primers were sequenced and were identical to the sequence obtained during the cloning process. In addition, polyclonal antibodies were used to trap ApMV from infected strawberries onto polystyrene immunoassay plates (NUNC, Rochester, NY). After washing, reverse transcription (RT) reactions were carried out in these wells and used in polymerase chain reaction (PCR). The sequence of the amplicons obtained were ApMV-specific as determined by sequencing. At this time, only the leafroll infected plant from the collection at the NCGR in Corvallis, OR has been tested. To our knowledge, this is the first characterization of a virus associated with strawberry leafroll disease, and development of the RT-PCR test will allow for detection of this virus in field samples to determine its distribution and importance in strawberry production.
References: (1) R. H. Converse. Pages 69--70 in: Virus Diseases of Small Fruits, USDA-ARS Handb. 631, 1987. (2) R. W. Fulton, No. 38. in: Descriptions of Plant Viruses CMI/AAB. Kew, Surrey, England, 1972. (3) I. E. Tzanetakis et al. J. Virol. Methods 124:73, 2005.
