Etiology and host range of a Closterovirus associated with plum bark necrosis-stem pitting disease

Diseased plum (Prunus salicina) cv. Black Beaut trees developed stem gumming, severe bark necrosis, and stem pitting symptoms on the woody cylinder of the main trunk and scaffold branches. The sucker shoots of the peach (Prunus persica) cv. Nemaguard understocks exhibited oak-leaf patterns, but lacked the wood or bark markings. Other susceptible hosts included almond (Prunus dulcis), sweet (Prunus avium) and Japanese flowering (Prunus serrulata) cherries, and several plum (Prunus salicina) and prune (Prunus domestica) varieties. A purified preparation containing high molecular size dsRNAs was obtained initially from diseased cherry (P. avium x Prunus pseudocerasus) cv. Colt tissues. Healthy preparations were devoid of similar sized dsRNAs. Reverse transcriptase-polymerase chain reaction (RT-PCR) assays with degenerate oligonucleotide Closterovirus primers, designed from the HSP70 gene, were used to amplify two DNA fragments of 0.67 and 0.56 kbp. The larger cDNA product was cloned, sequenced (AF195501), and compared with other viral sequences. Depending on the number of nucleotides used in the comparisons, identities ranged from 77% for Grapevine leafroll associated virus to 3 to 44% for Little cherry virus-1. Specific primers from the 0.67 kbp cDNA sequence were designed and used in subsequent RT-PCR assays. The associated 0.67 kbp HSP70 amplicon of Plum bark necrosis-stem pitting associated virus was detected in all graft-inoculated Prunus species and varieties except prune (P. domestica var, French Improved).