The RNA genome of grapevine leafroll-associated closterovirus-3 (GLRaV-3) was cloned as a cDNA generated from GLRaV-3-specific dsRNA, and a partial genome sequence of 13154 nucleotides (nt) including the 3' terminus was determined. The sequenced portion contained 13 open reading frames (ORFs) potentially encoding, in the 5'-3' direction, proteins of > 77 kDa (ORF1a; helicase, HEL), 61 kDa (ORF1b; RNA-dependent RNA polymerase, RdRp), 6 kDa (ORF2), 5 kDa (ORF3, small transmembrane protein), 59 kDa (ORF4; heat shock protein 70, HSP70), 55 kDa (ORF5), 35 kDa (ORF6; coat protein, CP), 53 kDa (ORF7; diverged coat protein, CPd), 21 kDa (ORF8), 20 kDa (ORF9), 20 kDa (ORF10), 4 kDa (ORF11), 7 kDa (ORF12), and an untranslated region of 277 nt. ORF1b is probably expressed via a +1 ribosomal frameshift mechanism, most similar to that of lettuce infectious yellows virus (LIYV). Phylogenetic analysis using various gene sequences (HEL, RdRp, HSP70 and CP) clearly demonstrated that GLRaV-3, a mealybug-transmissible closterovirus, is positioned independently from aphid-transmissible monopartite closteroviruses (beet yellows, citrus tristeza and beet yellows stunt) and whitefly-transmissible bipartite closterovirus (lettuce infectious yellows, LIYV). However, another alleged mealybug-transmissible closterovirus, little cherry virus, was shown to be more closely related to the whitefly-transmissible LIYV than to GLRaV-3.


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