Diversity of Citrus Tristeza Virus Isolates Indicated by dsRNA Analysis

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.

DsRNA Cloning and Diagnosis of Beet Pseudo-Yellows Virus by PCR and Nucleic Acid Hybridization

“…However, plants infected with such viruses as well as containing a genomic-length dsRNA also con tain smaller dsRN As [21], suggesting a different expression strategy to BPYV. The subgroup C closteroviruses, while being more similar than subgroup B closteroviruses to BPYV in particle length, also produce multiple dsRNA in in fected plants [22]. Thus, at present, BPYV does not fall easily into any of the proposed closterovirus subgroups.…”

Populations of citrus tristeza virus contain smaller-than-full-length particles which encapsidate sub-genomic RNA molecules

“…Several dsRNAs, including a 20 kbp molecule corresponding to the replicative form (RF) and a number of smaller species of dsRNAs, were consistently found following electrophoresis of RNA extracts from CTVinfected plants (Dodds & Bar-Joseph, 1983 ;Dodds et al, 1987). Two small dsRNA segments with estimated sizes of 0"8 kbp and 1.7 kbp were isolated and translated in vitro into two polypeptides of 27 kDa and 23 kDa, respectively (Dulieu & Bar-Joseph, 1990).…”

Analysis of the dsRNAs of apple chlorotic leaf spot virus

“…As different isolates have been used in these studies, these conflicting results could indicate some level of variability between isolates of ACLSV. Differences between isolates of CTV involving the presence of specific dsRNA segments have already been demonstrated, and it was suggested that the dsRNA electrophoretic patterns could be used as markers to identify specific CTV isolates (Dodds et al, 1987).…”