Diversity of Citrus Tristeza Virus Isolates Indicated by dsRNA Analysis

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Abstract
One major dsRNA of molecular weight (MW) 13.3 × 106 and two others (MW 1.9 × 106 and 0.8 × 106) were routinely detected by polyacrylamide gel electrophoresis in extracts from sweet orange {Citrus sinensis) or citron (Citrus medica) infected with each of 66 isolates of citrus tristeza virus (CTV). Several additional dsRNA were also commonly detected, usually as weakly stained bands in reproducible positions in gels, but some were very prominent, e.g., a dsRNA of MW 1.7 × 106 associated with a seedling yellows isolate (sy-1). No dsRNA was detected in equivalent extracts from noninoculated sweet orange and citron. End-labeled [32P] probes were made from purified full-length viral RNA or polyacrylamide gel-purified full-length dsRNA of a nonseedling yellows (nsy-1) and a seedling yellows (sy-1) isolate of CTV. Each of the four probes was able to hybridize to all major and most minor dsRNAs of both isolates in composite polyacrylamide/agrarose gels, including the 1.7 × 106 dsRNA specific to the seedling yellows isolate, and could readily detect CTV nucleic acid sequences in extracts from bark of infected sweet orange plants spotted onto nitrocellulose membranes. One dsRNA (MW 0.5 × 106) was very prominent in some isolates and much less so, or undetectable, in other isolates and 66 isolates have been screened for the presence of this dsRNA. There was a strong correlation between inability to detect the 0.5 × 106 dsRNA and the designation of an isolate as neither a seedling yellows type nor a stem pitting isolate of grapefruit; these properties were typical for isolates of CTV from southern California.