First Detection of Grapevine leafroll-associated virus 4 in Chilean Grapevines

Grapevine leafroll is one of the most widespread and economically relevant viral diseases of grapevines. At least nine distinct Grapevine leafroll-associated viruses (GLRaVs), all members of the Closteroviridae family, have been associated with this disease in grapevine. Grapevine leafroll-associated virus 4 (GLRaV-4), currently classified as a Closteroviridae member under the Ampelovirus genus, was initially described in California. To determine if GLRaV-4 was present in Chilean grapevines, in addition to the previously reported GLRaV-1, -2, -3, -7, and -9 (1,2), 35 dormant cane samples from 12 different cultivars were collected from different regions of Chile and screened by reverse transcription-PCR. Two of the 35 samples (both cv. Thompson Seedless) collected from the III and VI regions of Chile were found to be infected with GLRaV-4 using two different pairs of GLRaV-4 specific primers. The first pair of primers, HSPV-F: 5'- ACA TTC TCC ACC TTG TGC TTT T -3' and HSPC-R: 5'- CAT ACA AGC GAG TGC AAT TAC -3' (3), was used to amplify a 321-bp fragment corresponding to a partial region of the HSP70h gene. The sequence (GenBank Accession Nos. EU746618 and EU746619) from both positive samples shared 98.4% nucleotide identity and approximately 99% identity with the corresponding fragment of a Californian GLRaV-4 isolate (GenBank Accession No. AF039553). Since there are no commercial antibodies available for GLRaV-4 detection, a second pair of primers, LR4CPINT-F: 5'- GAG AGT GAC AAG CAC CAG GTG C -3' and LR4CPFIN-R: 5'- TCA CCT CCT GTT GCC CA -3' (4), that amplified a 492-bp fragment of the coat protein gene was also used. The sequences of the 492-bp fragment from both Chilean samples (GenBank Accession Nos. EU746620 and EU746621) shared 99.6% nucleotide identity with one another and had 96.5% identity with an Israeli GLRaV-4 isolate (GenBank Accession No. AM176759). To our knowledge, this is the first report of GLRaV-4 in Chile. Further studies will help to establish the effects and incidence of this virus in Chilean grapevines.