Search

Plum pox virus (PPV) is a potyvirus widely spread in many species of the Prunus genus such as plum, apricot, peach, sweet cherry and others. This potyvirus causes great damage to stone fruit trees in Romania and other European countries as Hungary, Italy, Czech Republic, France, Spain, Greece, Turkey, and Slovak Republic. The Research Station for Fruit Tree Growing Baneasa in Bucharest has realized many studies on the epidemiology and spread of PPV and also on the disease symptomatology and detection possibilities. The control of sharka disease by sanitary selection measures requires corresponding detection techniques. The aim of this study was to determine the presence or absence of PPV in some apricot and peach varieties and hybrids in 1995-1997 by the enzyme-linked immunosorbent assay (ELISA) and to verify if some of our biological materials evaluated as symptom-free under field conditions for many years are also virus-free and can be considered healthy.

Reverse transcription-polymerase chain reaction (RT-PCR) technique and restriction fragment length polymorphism (RFLP) analysis were used to analyse six isolates of plum pox virus (PPV). Whole coat protein (CP) gene was amplified in four isolates using the unipoty-polyT primer pari. PPV-D was identified by RFLP analysis using SfuI and DraI enzymes in two of the isolates. Two isolates of PPV-M strain yielded RT-PCR products which could not be digested by the two enzymes. Other isolates were subjected to RT-PCR using P1-P2 primers. The specificity of the RT-PCR products was confirmed by AluI digestion, while RsaI digestion enabled strain differentiation. No mixed infection was found.

Plum pox virus (PPV) is a major threat to the expanding Mediterranean stone fruit industry. In order to control the plum pox disease it is of utmost importance to detect early PPV foci and to identify the PPV isolates involved. A survey was therefore carried out in Albania, Cyprus, Egypt, Greece, Italy and Turkey by a double-antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) with the following monoclonal antibodies (MAbs): 5B (universal), 4DG5 (PPV-D-specific), AL (PPV-M-specific), TUV and AC (PPV-C-specific), and EA24 (PPV-El Amar-specific). A hundred and seventy Mediterranean PPV isolates were tested for strain type. PPV-M was detected in Albania, Cyprus, Greece, Italy, and Turkey; PPV-D was detected in Albania and Italy, whereas samples with natural mixtures of both strains were found in a couple of orchards in Albania. Seven PPV isolates from apricots in two Egyptian localities were recognized only by MAb EA24. In conclusion, DAS-ELISA with a combination of the universal MAb5B and the MAbs specific to the four PPV serotypes currently known (M, D, C and El Amar) is an efficient tool for a simple, sensitive and routine detection of PPV and discrimination of its serotypes.

Leaf tissues of stone fruit trees (plum, apricot, peach and myrobalan) carrying symptoms of plum pox virus (PPV) infection and of peach GF 305 seedlings and Nicotiana benthamiana infected experimentally with PPV were assayed for PPV by polymerase chain reaction (PCR). The expected 243 bp PCR products were subjected to restriction fragment length polymorphism (RFLP) analysis with restriction endonucleases AluI and RsaI. All of the PCR products contained the AluI site. The RsaI restriction profiles of the PCR products demonstrated the prevalence of PPV-M subgroup in the tested samples from western Slovakia.

The reverse transcriptase-polymerase chain reaction (RT-PCR) was used for detection of prunus necrotic ringspot virus (PNRSV) in dormant peach and almond trees by the application of two different pairs of primers yielding a short and a long product, respectively. The relative amount of the short (200 base pair, bp) product was higher than the longer (785 bp) product. PNRSV was detected better in plant tissues with a low virus concentration (e.g. dormant trees) by amplification of the short PCR product, whereas the long product was product was produced at higher virus titers. Simultaneous amplification of both short and long products was demonstrated using a three-primer mixture in a single reaction tube. In this assay, amplification of either PCR product indicated the presence of PNRSV-specific sequences in the plant tissue examined, thus covering a wide range of virus concentrations in a single test. Dilution of the RNA extracted from infected plant material resulted in a steep decline in the amplification of both short and long PCR products. In contrast, serial dilutions of the intermediate cDNA template differentially affected the amplification patterns: the relative amount of the short product increased whereas that of the long product decreased. These results may explain the preferential amplification of the short PCR product observed in samples containing low virus concentrations.

The complete nucleotide sequence of the RNA 3 of hydrangea mosaic virus (HdMV) was determined. It consists of 2268 nucleotides and contains two open reading frames (ORF). ORF 1 encodes for a putative translation product of 293 amino acids which shared 64.9% identity with the 3a protein of tobacco streak virus (TSV). ORF 2 encodes for a putative translation product of 220 amino acids which shared 54.2% identity with the coat protein of TSV. The relationship between the proteins of HdMV and the corresponding proteins of ilarviruses other than TSV was more distant. No zinc-finger-like motif was found in the coat protein of HdMV but the N-terminus of the protein was rich in basic amino acids. Both terminal, non-coding regions of HdMV RNA 3 contained repeated sequences with corresponding homologous fragments in the RNA 3 of TSV. On the basis of the similarities between HdMV and TSV that we detected, we propose that HdMV be included in subgroup 1 of the genus Ilarvirus together with TSV.

Olive latent virus 2 (OLV2), a virus with particle shapes resembling those of alfalfa mosaic alfamovirus (AMV), has four major RNA species, two of which (RNA3 and RNA4) were completely sequenced. RNA3 was a bicistronic molecule containing two clear-cut ORFs, one of which (ORF1) coded for a 36.5 kDa polypeptide with conserved motifs of the '30K superfamily' movement proteins and the other (ORF2) encoded a 20 kDa polypeptide identified as the viral coat protein. RNA4, which was a little smaller than RNA3 (2078 nt versus 2438 nt), also differed from RNA3 in a few positions, but its in vitro translation did not produce any protein. By contrast, an additional RNA, 1042 nt in size with strong sequence homology with RNA3 and RNA4, was identified in RNA extracts from infected plants. This RNA, which may be a subgenomic form of RNA3 carrying the coat protein cistron, is apparently encapsidated to a very small extent, or not at all. Comparative computer-assisted analysis of virus-coded protein sequences disclosed homologies suggesting that OLV2 may belong to the family Bromoviridae, but as an entity separated from the currently known genera.