Comparisons of some properties of two laboratory variants of Raspberry bushy dwarf virus (RBDV) with those of three previously characterised RBDV isolates

The properties of two laboratory variants of Raspberry bushy dwarf virus (R BDV), genus Idaeovirus, were compared with those of their parental sources and with two naturally occurring variants. Isolate RB is a natural variant able to overcome the resistance to RBDV present in some red raspberry culti vars. Isolate M is a serological variant from black raspberry. Laboratory v ariant D1, was derived from the Scottish type isolate (D200) by continuous sub-culture in Chenopodium quinoa. Laboratory variant Can-S was derived fro m an isolate infecting Canby red raspberry in Canada (Can) after passage th rough Nicotiana benthamiana. All isolates reacted with a polyclonal antiser um to isolate D200 in agarose gel double-diffusion tests but, whereas isola tes D200, RB, Can and Can-S were serologically indistinguishable, the preci pitin lines formed by these isolates each spurred over those formed by isol ates D1 and M. All six isolates reacted strongly with the polyclonal antise rum in double antibody sandwich and plate-trapped antigen (PTA) forms of EL ISA and in Western blotting (WB) and when each of four monoclonal antibodie s (Mabs) to an unnamed red raspberry isolate from Canada was used to detect antigen trapped by the polyclonal antiserum. However, the virus isolates d iffered in their reactions to these four Mabs in PTA-ELISA and in WB. Isola tes RB, Can and Can-S behaved similarly in these tests as did isolates D200 and D1, but isolate M was distinct. In herbaceous test plants, variants D1 and Can-S were readily distinguished from their parental sources and from the other two isolates by producing either no symptoms (D1) or very severe symptoms (Can-S) in hosts. Unlike all other isolates studied world-wide, Ca n-S failed to infect C. quinoa systemically but induced severe necrotic loc al lesions in this and other hosts. Reverse transcription-polymerase chain reaction was used to amplify the gen e encoding the coat protein (CP) in RNA-2, and a region of the gene encodin g the polymerase in RNA-1. The nucleotide sequences of the CP genes of the six isolates were > 96% identical but isolate Can-S was the most distinctiv e. However, the similarity between Can-S and its parent isolate (Can) was n o greater than the similarity between Can-S and the other isolates, suggest ing that Can-S may not have arisen as the result of a mutation from isolate Can. Sequence comparisons of parts of the polymerase gene of isolates R15, D1, D200 and Can-S showed that they were 95-98% identical.