First record of tomato yellow leaf curl Sardinia virus (TYLCSV) on pepper in Italy.

During a survey in summer 2007, a disease of pepper (Capsicum annuum) under plastic tunnels was observed in Policoro (Matera), on the Ionic coast of Basilicata Region, with a disease incidence in some cases of more than 50%. Affected cultivars were Eppo and Almund (S Et G). The diseased plants exhibited light mosaic or mottling, leaf distortion, interveinal and marginal leaf chlorosis, upward curling of leaf margins of older leaves. The causal pathogen was suspected to be a begomovirus due to the large population of the whitefly Bemisia tabaci observed on the crop. Detection assays for Tomato yellow leaf curl Sardinia virus (TYLCSV) and Tomato yellow leaf curl virus (TYLCV) were used. In DAS-ELISA, positive results (178 plants resulted positive over 200 symptomatic plants assayed) were obtained using a "broad-spectrum" reagent combination (distributed by Bioreba AG) detecting TYLCV, TYLCSV, and other begamoviruses. A couple of synthetic oligonucleotides allowing the amplification of the whole coat protein (CP) gene of TYLCSV and TYLCV was used for PCR of ELISA positive samples in order to perform the molecular characterisation of the viral isolate responsible of the disease. RFLP analysis performed on the PCR product, 1008 bp long, showed the presence of only TYLCSV in the infected pepper plants. The same couple of primers allowed the detection of the virus also in symptomless pepper plants. To test whitefly transmission, adults of B. tabaci allowed to feed on naturally infected pepper plants were transferred on 10 healthy Eppo pepper seedlings (15 whiteflies/plant). Insects were killed 2 days later using an insecticide. Twenty days post exposition 10 plants/10 resulted positive in ELISA, and showed the same symptoms observed in natural infection. TYLCSV was not reported before on pepper in the surveyed area, but it was recorded with severe outbreaks on tomato, both in protected and in open field crops. This species was probably the primary source of infection from which subsequent diffusion by way of the vector B. tabaci followed on pepper. To our knowledge this is the first time that a natural infection of TYLCSV on pepper is recorded in Italy, with serious implications for the epidemiology of TYLCSV in our country.

Integrated management of TYLCV/TYLCSV on greenhouse hydroponic tomatoes in Southern Italy.

Tomato yellow leaf curl (TYLC) caused by Tomato yellow leaf curl virus (TYLCV) and Tomato yellow leaf curl Sardinia virus (TYLCSV), vectored by the whitefly Bemisia tabaci, is a major disease of tomato in Sardinia and Sicily, and is becoming a serious threat in Southern Italy too. TYLCSV was first reported in Calabria region in 1991, but apparently it was an occasional outbreak, and B. tabaci was not detected. Later, during the 2003-2004 winter, a serious epidemic was observed in protected tomato crops in Castrovillari, Cosenza province. TYLCV was first described in Sicily in 2003 and during 2004 in continental Italy. Both viruses were detected in winter 2005-2006 on the Basilicata Ionic coast, in the Metapontum area, both in protected and in open field tomato crops. Experiments were conducted in Calabria Region, Southern Italy, under controlled conditions in a group of greenhouses where several tomato crops were grown hydroponically to determine the separate and integrated effects of UV-reflective mulch (UVRM), Acibenzolar-S-methyl (Actigard) and the two insecticides Imidacloprid (ADMIRE 2F) and Thiamethoxam (ACTARA 25WG). Highly UV-reflective mulch covered plots were treated with Actigard and insecticides, both alone or in combination. TYLC disease incidence was determined from late August 2005 to late January 2006. The highly UVRM alone was effective in reducing disease incidence of about 28.6% at the end of October, and of 31.7% at the end of January. However, Actigard with UVRM significantly reduced TYLC disease incidence to 70% and 48.5%, in 2 months and 5 months after the first treatment, respectively. The insecticides with UVRM, resulted in a moderate reduction of disease incidence (22.5%) at the end of October. At the end of January a reduction in disease incidence due to insecticide applications was not significant. The use of Actigard combined with the insecticides on UVRM reduced the disease incidence (63.4% with Admire and 56.1% with Actara) at the end of January. Actigard alone or with insecticides on UVRM was effective in reducing disease incidences. Highly UVRM and Actigard were effective in reducing the primary spread of TYLCV/TYLCSV in greenhouse hydroponic tomatoes. Comparative analysis of their effects at different periods post-treatment suggests that multiple applications of Actigard may be necessary to reduce progress of this disease.

Potato virus Y CFH, a putative recombinant isolate from Capsicum chinense cv. Habanero.

Ornamental plants of Chili pepper, Capsicum chinense cv. Habanero, with symptoms of leaf mosaic, necrotic rings on fruits and necrotic stems were observed in June 2003 in a private garden in the province of Naples (Italy). Preliminary serological characterisation allowed the association of these symptoms with infections by Potato virus Y (PVY). The virus was isolated on Nicotiana tabacum cv. Xanthi and characterised by mechanical inoculation on herbaceous hosts and molecular characterisation of the P1 and the coat protein (CP) genes. Symptoms produced on indicator plants were generally consistent with those described for PVY. The identity of PVY was further confirmed by reaction with PVYN, PVYC and PVYO specific monoclonal antibodies: the isolate reacted only with the PVYC specific Mab. Immuno capture reverse transcription polymerase chain reaction (IC-RT-PCR) was performed on extracts of PVY-CFH infected N. tabacum cv. Xanthi plants, using two couples of primers specifically designed out of the P1 and the CP coding regions of the so far fully sequenced PVY isolates. PCR products were then cloned into pCRII-TOPO vector using TOPO-TA cloning kit (Invitrogen) and sequenced. Sequence analysis suggests that PVY-CFH originated from a recombination event involving a virus of the PVYO type and another parental virus, maybe resembling the PVYNP isolates, given the reasonably high similarity shared by PVY-CFH and, respectively, non potato PVY isolates in the CP coding region, PVYO isolates in the P1 coding region. Evidence for the existence of such a recombination comes, apart from similarity analysis, by the different locations of CFH within phylogenetic trees constructed from P1 and CP genomic regions.

Characterisation of Potato virus Y nnp strain inducing veinal necrosis in pepper: a naturally occurring recombinant strain of PVY.

The full-length genome of Potato virus Y (PVY) nnp strain, recovered from pepper showing veinal necrosis of leaves, was cloned and sequenced, finding an organisation typical for PVY species. It consists of 9699 nucleotides (nt) excluding the 3' terminal poly(A) tail and contains an open reading frame of 9186 nt, encoding the putative polyprotein of 3061 amino acids. In ELISA, the isolate reacted with a monoclonal antibody specific for PVY(C) but not with antibodies against PVY(N) or PVY(O). Sequence analysis strongly suggests that PVY-nnp originated from a recombination event involving a virus of the PVY(O) type and another parental virus, maybe resembling the PVY(NP) isolates, given the reasonably high similarity shared by PVY-nnp and Lye84.2 and Son41 isolates. The recombination event involved a breakpoint near the middle of the P1 gene, around position 603 of the viral genome. Proof for the existence of such a recombination comes from several lines of evidence, including similarity analysis, recombination analysis using six different methods and the different locations of nnp within phylogenetic trees constructed from genomic regions on either side of the identified recombination breakpoint.

Severe outbreaks of tomato yellow leaf curl Sardinia virus in Calabria, Southern Italy.

During the winter 2003--2004 a serious disease was observed in protected tomato crops in Castrovillari, Reggio Calabria province, Southern Italy. Symptoms consisted in marginal leaf yellowing, leaf curling, plant stunting, flower abortion. The disease was detected in a group of greenhouses (about 10ha) where several tomato cultivars were grown hydroponically. The highest incidence of infection (60-100%) was observed in tomatoes grafted on Beaufort DRS tomato rootstock. Since the symptoms were similar to those described for Tomato yellow leaf curl Sardinia virus (TYLCSV) and Tomato yellow leaf curl virus (TYLCV), detection assays for these viruses were used. In DAS-ELISA positive results were obtained with a abroad-spectrums reagent combination (distributed by Bioreba AG) detecting TYLCV, TYLCSV, and other begomoviruses. When DNA probes were used in tissue print assays, positive reactions were obtained for TYLCSV, but not for TYLCV. The two probes consisted of digoxigenin-labelled DNAs representing the coat protein gene of either TYLCSV or TYLCV. Attempts to isolate the viral agent by mechanical inoculation failed, except in few cases where Potato virus Y and Tobacco mosaic virus were identified following transmission from symptomatic plants to herbaceous indicatorpplants. By contrast, grafting onto tomato seedlings always successfully transmitted the disease. In the Castrovillari area TYLCSV was not reported before. The rootstocks that nurseries used for grafting were obtained from Sicily, where the disease is endemic and both TYLCSV and TYLCV are widespread. Probably the grafted plantlets represented the primary source of infection from which subsequent diffusion by way of the vector Bemisia tabaci followed. In fact the vector had previously been detected in both the glasshouse-grown and open field tomato crops in Calabria region. TYLCV was previously reported in a different area of Calabria in 1991, but apparently it was an occasional outbreak, and B. tabaci was not detected. Since in the Castrovillari area surveyed in the present study tomato is grown throughtout the year in protected crops, the whitefly vector of the virus is present, and some natural hosts of the virus are found, it is feared that TYLCSV may become endemic, as already happened in Sicily, Sardinia, and Spain several years ago. In Spain and Sicily TYLCV, together with TYLCSV, was reported as the causal agent of very severe tomato crop losses. Therefore the danger exists that also TYLCV will reach this area, furthermore complicating the management of tomato crops.

The complete nucleotide sequence of Plum pox virus isolates from sweet (PPV-SwC) and sour (PPV-SoC) cherry and their taxonomic relationships within the species.

Plum pox virus (PPV) sweet (SwC) and sour (SoC) cherry isolates were the first PPV isolates to be recovered from natural infection in sweet and sour cherry plants, respectively. Their complete nucleotide sequences have been determined finding a deduced genome organisation typical for PPV species. Both genomes are 9795 nucleotides long, excluding the 3' terminal poly(A) tail, and contain an open reading frame of 9432 nt, encoding a polyprotein of 3143 amino acids. The nucleotide and predicted amino acid sequences of PPV-SwC and SoC have been pairwise compared with available sequences of different PPV strains. Although a very high similarity exists between the whole genomes and polyproteins of the two cherry isolates, high levels of divergence have been calculated with sequences of PPV-M, D and EA isolates. In particular, the most considerable divergence has been found in part of 5' non coding region, in regions encoding P1, P3 + 6K1, 6K2 and NIa-VPg proteins as well as in the N-terminal domain of the coat protein. Phylogenetic analysis have been undertaken in order to establish the taxonomic localisation of SwC and SoC isolates within PPV species, showing that they are always clustered together and separated from the rest of PPV strains, being clearly the most distant.

Postprocessing of images by filtering the unmasked coding noise.

This paper presents a methodology for the restoration of the visual quality of still images affected by coding noise. This quality restoration is achieved only by considering the additive coding noise and is therefore limited to an adaptive postprocessing filtering. It is based on a model of the human visual system that considers the relationship between visual stimuli and their visibility. This phenomenon known as masking is used as a criterion for the locally adaptive filtering design. An image transformation that yields visual stimuli tuned to the frequency and orientation according to the perceptual model is proposed. It allows a local measure of the masking of each perceptual stimulus considering the contrast between signal and estimated noise. This measure is obtained by analytic filtering. Processing schemes are presented with applications to the discrete cosine transform (DCT) and subband coded images. One proposed solution considers the characteristics of DCT coding noise for the estimation of the noise. Another solution is based on a "blind" neural estimation of the noise characteristics. Experimental results of the proposed approaches show significant improvements of the visual quality, which validates our perceptual model and filtering.

Production of strain specific antibodies against a synthetic polypeptide corresponding to the N-terminal region of the plum pox potyvirus coat protein.

Comparison of the predicted coat protein amino acid sequence of the 'sweet cherry' strain of plum pox potyvirus (PPV-SwC) with the corresponding regions of several other PPV strains indicated that the main differences are in the N-terminal region. Polyclonal antibodies were produced against a synthetic peptide corresponding to the 1-14 sequence of the N-terminal region of PPV-SwC coat protein. They specifically detected PPV-SwC in different immunochemical tests.

Characterization of the Sweet Cherry Isolate of Plum Pox Potyvirus.

An isolate of plum pox potyvirus from sweet cherry (PPV-SwC) in southern Italy was investigated. The isolate was mechanically or graft transmissible to different Prunus and Nicotiana spp. but not to Chenopodium spp. It was transmitted also by Aphis fabae and Myzus persicae in a nonpersistent manner. Restriction fragment length polymorphism analysis indicated and nucleotide sequencing confirmed that the isolate lacks AluI and RsaI sites in the C-terminal region of the coat protein (CP) gene. Western immunoblot results showed that the PPV-SwC CP has an electrophoretic mobility similar to that of strain PPV-D and faster than that of strain PPV-M. Double-antibody sandwich indirect enzyme-linked immunosorbent assay of the CP showed that PPV-SwC, although reacting with universal monoclonal antibodies to PPV, failed to react with antibodies specific to strains M and D. Results indicate that PPV-SwC is different from conventional strains of PPV but closely related to the sour cherry isolate of PPV from Moldova.