First Report of Citrus leaf blotch virus in New Zealand.
Despite a high incidence of Citrus tristeza virus (CTV) in citrus in New Zealand, viral diseases have had only a minor impact on the New Zealand citrus industry, largely because of the use of Poncirus trifoliata and hybrid rootstocks derived from this. In August of 2007, a PCR-based survey for seven citrus viruses was conducted on 104 commercial orchard trees that represented a range of Citrus scion species, as well as P. trifoliata and P. trifoliata × Citrus sinensis rootstock, grown from imported and local budwood or from seed in the case of rootstocks, from the citrus-growing regions of Kerikeri, Tauranga, and Gisborne. Total RNA was extracted from young, green bark and leaf tissue from each source. Using a primer pair amplifying a 425-bp region of the Citrus leaf blotch virus (CLBV) coat protein gene (sense: 5'-AGCCATAGTTGAACCATTCCTC-3' and antisense: 5'-GCAGATCATTCACCACATGC-3'), 26 (25%) of the plant samples yielded a DNA fragment of the size expected for CLBV, including 6 of 21 C. sinensis, 1 of 2 C. limon, 3 of 10 C. unshiu, and 5 of 7 C. paradisi scion samples and 5 of 9 P. trifoliata and 6 of 9 P. trifoliata × C. sinensis rootstock samples. Identification of CLBV (an unclassified member of the family Flexiviridae) was confirmed by amplification of a second region of the genome of 1,045 bp spanning the ORF2/ORF3 domains (sense: 5'-ATGAAAAGCCAGTTATGCACCA-3' and antisense: 5'-CTCAGCATTCCCAGGAATAACC-3'). A subset of the PCR products for the CP and ORF2/ORF3 fragments were sequenced for analysis (GenBank Accession nos. EU670243-EU670251 and EU862284-EU862288). Sequences from all CLBV isolates showed 97 to 99% nucleotide identity with the CLBV reference isolate SRA-153 (GenBank Accession No. AF318061) (1) in the coat protein fragment and 98 to 99% nucleotide identity for the partial ORF2/ORF3 fragment. The bud-union crease symptom reported to be caused by CLBV was observed only on one C. paradisi scion on P. trifoliata rootstock, while the remaining samples were either asymptomatic or symptoms were attributed to co-infection with CTV. Since CLBV-infected plants were found from all major growing regions, it is apparent that CLBV is widespread in New Zealand, although it is not known where in this country it may have originated. Reference: (1) M. C. Vives et al. Virology 287:225, 2000.