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Title: First report of taro vein chlorosis virus infecting taro [Colocasia esculenta (L.) Schott] in the United States of America

item MELZER, MICHAEL - University Of Hawaii
item LONG, M H - University Of Hawaii
item AYIN, C - University Of Hawaii
item Li, Ruhui
item HU, JOHN - University Of Hawaii

Submitted to: Plant Disease
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 2/25/2014
Publication Date: 2/28/2014
Citation: Melzer, M., Long, M., Ayin, C., Li, R., Hu, J. 2014. First report of taro vein chlorosis virus infecting taro [Colocasia esculenta (L.) Schott] in the United States of America. Plant Disease.

Interpretive Summary:

Technical Abstract: In March 2013, taro plants [Colocasia esculenta (L.) Schott cv. ‘iliuaua’] with leaves displaying veinal chlorosis and necrosis were observed at a germplasm collection on the island of Molokai. These symptoms were similar to those of taro vein chlorosis, an important disease of taro found in several South Pacific nations, caused by taro vein chlorosis virus (TaVCV; family Rhabdoviridae, genus Nucleorhabdovirus). To investigate this possibility, RNA was isolated from both symptomatic and asymptomatic taro leaves using the NucleoSpin® RNA II extraction kit (Macherey-Nagel, Bethlehem, PA) according to the provided protocol, except that RLT Buffer (Qiagen Inc., Valencia, CA) was used as the initial extraction buffer. The RNAs were converted to cDNA using random primers and MMLV-RT reverse transcriptase (Promega, Madison, WI) followed by a PCR assay using primers TaVCV1 and TaVCV2 which target the RNA-dependent RNA polymerase (RdRp) gene of TaVCV (1). A ~200 bp product was amplified from symptomatic but not asymptomatic leaf tissue and was cloned using pGEM-T® Easy (Promega, Madison, WI). Three clones were selected and their sequences determined by dye-terminator sequencing. The 170-bp consensus sequence was found to be 78% identical to a Fijian isolate of TaVCV (GenBank AY674964) following primer sequence removal. When converted to amino acid sequence, this fragment was found to be 87% identical to the RdRp of TaVCV (AAV92087). To confirm the presence of TaVCV in symptomatic taro tissue, primer sets Pol2A1/Pol2A2 and Cap2A/Cap2B which target the RdRP and putative nucleocapsid genes of TaVCV, respectively, were used in PCR-based detection assays (1). Amplification products of the correct size were obtained for both primers sets, then cloned and sequenced as described above. After primer sequence removal, the Pol2A1/Pol2A2 product (952 bp; GenBank KF921085) and Cap2A/Cap2B product (1050 bp; GenBank KF921086) were found to be 79% and 84 % identical to TaVCV (GenBank AY674964), respectively. Samples from 328 taro plants with and without taro vein chlorosis symptoms were collected from 35 sites on five of the major Hawaiian Islands and assayed for TaVCV using the Pol2A1/Pol2A2 primer set as described above. The overall incidence of TaVCV in these samples was 21.6%, with positive samples detected from each island. Although a very strong association between symptoms and the presence of TaVCV was observed, a few asymptomatic plants were also positive, suggesting the assay was able to detect the virus before the onset of symptoms. TaVCV exhibits a great deal of genetic diversity in the South Pacific nations where it is found; up to 27% nucleotide divergence in regions of the RdRp gene have been reported (1). The high genetic divergence between the TaVCV isolate characterized in Hawaii and the TaVCV accession in GenBank supports this observation. The widespread distribution of TaVCV in Hawaii suggests it is not a recent introduction. However, the common practice of farmers sharing taro propagules has likely accelerated its spread. An arthorpod vector of TaVCV has yet to be identified, so it is unknown whether natural spread is also occurring in Hawaii. Taro has both economic and cultural importance to Hawaii. These findings, representing the first detection of TaVCV in Hawaii and the USA, illustrate the need to develop virus-free germplasm for local, national, and international distribution of this important staple crop.