First Report of Papaya ringspot virus Infecting Zucchini Plants in Poland.

Abstract

Papaya ringspot virus (PRSV), a member of the aphid-transmitted genus Potyvirus, is the cause of a destructive disease and a major limiting factor for papaya and cucurbit cultivation worldwide. The virus occurs in China, France, Germany, India, Italy, Mexico, Taiwan, and the United States. Its P biotype is a devastating pathogen of papaya crops and its W biotype is a pathogen of cucurbits (4). In 2009, zucchini plants with leaf mosaic and marbled fruit were collected from the Kujawsko-Pomorskie Region of Poland. Samples came from the same region where Zucchini yellow mosaic virus (ZYMV) (3) and Watermelon mosaic virus (WMV) (1) have been found previously. Forty leaf and ten fruit samples of zucchini (Cucurbita pepo cv. giromontina) were tested by double-antibody sandwich (DAS)-ELISA with commercial antisera against WMV, ZYMV, and PRSV (DSMZ, Braunschweig, Germany). PRSV was found in two samples tested. Leaf extracts from infected plants were mechanically inoculated onto Carborundum-dusted leaves of the following indicator plants: Cucumis sativus, Chenopodium quinoa, Cucurbita pepo, Nicotiana benthamiana, and N. tabacum cv. Xanthi. After 2 weeks, symptoms of leaf chlorosis on cucumber and chlorotic lesions on zucchini were observed. Total RNA was extracted from infected leaves with a phenol-chloroform based extraction procedure. The presence of PRSV was confirmed by reverse transcription (RT)-PCR reaction using primers 04-02 and 04-04, which amplify the coat protein gene (2). Amplified DNA was gel purified with a Qiaex Kit (Qiagen, Valencia, CA) and cloned into pGEM-T easy (Promega, Madison, WI). Overlapping sequences were obtained using universal M13F and M13R primers. BioEdit software ( http://www.mbio.ncsu.edu/BioEdit/bioedit.html ) was used to assemble the nucleotide consensus sequence. The obtained sequence (861 bp encoding 287 amino acids) was deposited in the GenBank database under Accession No. GQ927328. The comparison with PRSV sequences retrieved from the GenBank database were carried out to investigate the genetic diversity between Polish PRSV isolates and establish their molecular relationships to the previously characterized PRSV isolates from different parts of the world. The sequences of PRSV Polish isolates obtained from two infected plant samples were identical. Comparisons revealed that the Polish isolate designated PRSV-BON shared the highest identity (97%) with three Australian isolates (U14739, U14740, and U14744). To our knowledge, this is the first report of PRSV infecting zucchini plants in Poland. The occurrence of subtropic viruses like PRSV in Poland indicated the introduction of new pathogens that likely affect cucurbit production in this country and beyond.