No abstract

The plum pox virus (PPV) and prunus necrotic ringspot virus (PNRSV) cause serious disease problems in stone-fruit trees. In this work, the possibility of obtaining plant material free from these viruses through thermotherapy and meristem-tip culture from infected nectarine shoots (Prunus persica var. nectarina Max, cv. 'Arm King') was studied. In addition, the detection of these viruses in in vitro cultures and young acclimatized plantlets with double antibody sandwich-enzyme-linked immunosorbent assay (DAS-ELISA) and multiplex reverse transcriptase-polymerase chain reaction (RT-PCR) was studied. Meristem-tip explants (0.8-1.3 mm) derived from sprouted buds of winter wood and spring shoots from field grown plants had a 2-5% regeneration response. However, application of thermotherapy to potted nectarine trees (3 weeks at a maximum temperature of 35 degrees C) facilitated excision of longer meristem tips (1.3-2.0 mm) that resulted in a significantly higher regeneration response (38%) in woody plant medium (WPM) without plant growth regulators. Such explants formed multiple shoots with the addition of 8 microM benzylaminopurine and 0.8 microM indoleacetic acid. When they were tested for the presence of PPV and PNRSV, 86% and 81% were found to be virus-free as detected by DAS-ELISA and multiplex RT-PCR, respectively. Individual shoots excised from virus-free cultures readily rooted in vitro (half-strength WPM plus 2 microM indolebutyric acid) and grew to plantlets. The combination of an efficient protocol for virus elimination and the establishment of highly sensitive diagnostics resulted in the production of nectarine plants free from PPV and PNRSV.

In a survey during 2000–2002 to determine the identity and prevalence of viruses affecting cucurbit crops in Cyprus, 2993 samples of cucumber, zucchini, melon and watermelon were collected from the five major cucurbit‐growing areas in Cyprus. Zucchini yellow mosaic virus (ZYMV), Papaya ringspot virus type W (PRSV‐W), Watermelon mosaic virus (WMV), Cucurbit aphid‐borne yellows virus (CABYV), Cucumber mosaic virus (CMV) and Squash mosaic virus (SqMV) were detected by enzyme‐linked immunosorbent assay (ELISA), and Cucurbit yellow stunting disorder virus (CYSDV), Beet pseudo‐yellows virus (BPYV) and Cucumber vein yellowing virus (CVYV) by reverse transcription polymerase chain reaction (RT–PCR). ZYMV was the most prevalent virus of cucurbits in Cyprus with an overall incidence of 45%. PRSV‐W, CABYV and WMV were detected in 20.8%, 20.8% and 7.8% of the samples tested, respectively. CYSDV was detected in most greenhouse cucumber samples with yellowing symptoms (88.1%), whereas BPYV and CVYV were found in only 2.4% and 9.5%, respectively, of samples. CMV and SqMV were not detected in any cucurbitaceous crop during this survey.

A survey of the incidence of yellowing viruses in Greek glasshouse (and occasional field) cucumber and melon crops was carried out during 2000 -03. In most cases disease incidence ranged from 50 to 80%. Simplex RT-PCR was used for the detection of Beet pseudo-yellows virus (BPYV) and Cucurbit yellow stunting disorder virus (CYSDV), and DAS-ELISA for the detection of Cucurbit aphid-borne yellows virus (CABYV). The results showed that BPYV was the predominant virus in cucumber and melon crops, whereas CYSDV, reported for first time in Greece, was isolated only in three regions of southern Greece: Rhodes, Crete and Arkadia. CABYV was detected only in three cucumber glasshouses in Pella (Macedonia). A simplified triplex RT-PCR method using a simple sample-preparation protocol was developed to allow rapid, sensitive and simultaneous detection of the three viruses. Sequence comparisons of the PCR products of BPYV and CYSDV revealed 98·7 and 100% amino acid identity, respectively, with previously reported sequences. The arable weed species Amaranthus retroflexus , Selosia cristata , Sonchus oleraceus and Sonchus sp. were identified as potential BPYV reservoirs.

A severe disease with combined symptomatology affecting only grafted watermelon plants appeared in several regions of northern and central Greece during the summers of 1999 and 2000. Disease symptoms included chlorotic mottling followed by pedicel necrosis at a later more mature fruit stage, and finally decomposition of fruit interior. Electron microscopy, double antibody sandwich enzyme-linked immunosorbent assay tests and reverse transcriptase-polymerase chain reaction assays identified Cucumber green mottle mosaic virus (CGMMV) as the causal agent. Our studies indicated that the CGMMV isolates from watermelon are closely related to the Israeli variant of CGMMV (CGMMV-Is). However, bioassays revealed that the Greek isolates elicit symptoms of higher severity and have a wider host range, compared with other described CGMMV isolates. Nucleotide sequence comparison of the movement protein gene of the Greek isolates showed a 97-99% identity with other CGMMV strains. This paper reports the presence of the virus in arable weeds such as Amaranthus blitoides, Amaranthus retroflexus, Heliotropium europaeum, Portulaca oleracea and Solanum nigrum.U. S.