Biological properties,transmission, serological characterization and varietal susceptibility of an isolate of Papaya leaf curl virus affecting papaya crops in Eastern Uttar Pradesh,India

Papaya leaf curl disease (PLCD) was recorded with 5–35% incidence at six districts of Eastern Uttar Pradesh during survey. The characteristic symptoms observed were severe downward leaf curling, swelling of veins, twisting and reduction of petioles, inverted leaf bowls and stunted growth of the entire plant which bore only few small and distorted fruit. The virus isolate was identified as Papaya leaf curl virus (PaLCuV).The PaLCuV isolate was successfully transmitted by whitefly (Bemisia tabaci) but not by mechanical (sap) transmission on Carica papaya plants. Plants could be proved efficiently from infected to healthy C. papaya, Capsicum annuum, Lycopersicon esculentum, Nicotiana tabacum, Crotalaria juncea, Ageratum conyzoides, Zinnia elegans, Datura stramonium and Petunia hybrida. Symptomatic samples of these plants were tested with polyclonal antiserum of Tomato leaf curl New Delhi virus by DAC-ELISA test showed the positive relationship of the samples with geminivirus. On the basis of symptomatology, whitefly transmission, host range studies and serological relationship, the isolate was identified as whitefly transmitted geminivirus. To identify potential varietal resistances source to PaLCuV, five cultivars of C. papaya were tested against PaLCuV using whitefly insects to transmit the infection. Results revealed that two cultivars (Washington and Ranchi Dwarf) were found to be moderately resistant.     

Relative incidence,spatial distribution and genetic diversity of cucurbit viruses in eastern Spain

Viral diseases that could cause important economic losses often affect cucurbits, but only limited information on the incidence and spatial distribution of specific viruses is currently available. During the 2005 and 2006 growing seasons, systematic surveys were carried out in open field melon (Cucumis melo), squash and pumpkin (Cucurbita pepo), watermelon (Citrullus lanatus) and cucumber (Cucumis sativus) crops of the Spanish Community of Valencia (eastern Spain), where several counties have a long standing tradition of cucurbit cultivation and production. Surveyed fields were chosen with no previous information as to their sanitation status, and samples were taken from plants that showed virus‐like symptoms. Samples were analysed using molecular hybridisation to detect Beet pseudo‐yellows virus (BPYV), Cucurbit aphid‐borne yellows virus (CABYV), Cucumber mosaic virus (CMV), Cucumber vein yellowing virus (CVYV), Cucurbit yellow stunting disorder virus (CYSDV), Melon necrotic spot virus (MNSV), Papaya ring spot virus (PRSV), Watermelon mosaic virus (WMV) and Zucchini yellow mosaic virus (ZYMV). We collected 1767 samples from 122 independent field plots; out of these, approximately 94% of the samples were infected by at least one of these viruses. Percentages for the more frequently detected viruses were 35.8%, 27.0%, 16.5% and 7.2% for CABYV, WMV, PRSV and ZYMV, respectively, and significant deviations were found on the frequency distributions based on either the area or the host sampled. The number of multiple infections was high (average 36%), particularly for squash (more than 57%), with the most frequent combination being WMV + PRSV (12%) followed by WMV + CABYV (10%). Sequencing of WMV complementary DNA suggested that ‘emerging’ isolates have replaced the ‘classic’ ones, as described in southern regions of France, leading us to believe that cucurbit cultivation could be severely affected by these new, emerging isolates.     

Production of Polyclonal Antibodies Using Recombinant Coat Protein of Papaya ringspot virus and their Use in Immunodiagnosis

Production of polyclonal antibodies requires large amount of purified virus that can be avoided by the use of recombinant coat protein (CP). Recombinant CP of Papaya ringspot virus (PRSV) was thus used for the production of polyclonal antibodies as the virus purification from papaya tissues provides low virus yields. CP was expressed as a fusion protein (～72 kD) containing a fragment of E. coli maltose binding protein. Polyclonal antibodies from rabbits immunized with the fusion protein, successfully detected natural infection of PRSV in papaya and cucurbits samples collected from different locations at 1:4000 dilution in direct antigen-coated enzyme-linked immunosorbent assay.     

Incidence of Viruses and Nematode Vectors in Lebanese Vineyards

Surveys for virus diseases and nematode vectors were conducted in 95 commercial vineyards of four different Lebanese districts (Bekaa valley, Mount Lebanon, North and South Lebanon). Out of 915 randomly collected grapevine samples tested by ELISA, 511 (55.8%) were infected by one or more viruses. Grapevine virus A (30.9%) and Grapevine leafroll‐associated virus 3 (23.7%) were the prevailing viruses, followed by Grapevine fleck virus (15.1%), Grapevine leafroll‐associated virus 1 (10.6%) and Grapevine leafroll‐associated virus 2 (8.7%). Arabis mosaic virus was not found whereas Grapevine fanleaf virus (GFLV) and Grapevine virus B were little represented. The most important Lebanese grapevine varieties, i.e. Maghdouchi, Tfeifihi and Beitamouni, had average infection rates between 70% and 87%, whereas varieties of foreign origin had a better sanitary status with the exception of cvs Cinsaut and Thompson (c. 83% infection). Grapevine rupestris stem pitting‐associated virus was detected in 79 of 90 (87.8%) samples tested by RT‐PCR and closteroviruses were recorded in seven of 70 (10%) vines tested. One of these viruses was identified as Grapevine leafroll‐associated virus 5 by ELISA and partial genome sequencing. No nepoviruses other than GFLV were detected in any of 90 samples tested using three different sets of degenerate primers. Xiphinema index was found in 23 of 89 soil samples collected from vineyards, and in three of 15 samples collected primarily under fig trees in fields where no grapevines were grown.     

Serological detection of yam viruses in farmers' fields in Ogun State,Nigeria

A field survey was conducted in eight local government areas (LGA) of Ogun state, Nigeria to assess the incidence of viral diseases of yams in the areas. Leaf samples were collected from 90 yam plants which were either symptomatic or asymptomatic. These were bulked into 45 during serological tests and the viruses indexed include yam mosaic virus (YMV); Dioscorea alata bacilliform virus (DaBV) and cucumber mosaic virus (CMV). DaBV was the most prevalent virus on the field with incidence of 48.9% (22/45) followed by YMV which occurred in 42.2% (19/45). CMV had the lowest percentage of incidence; 2.2% (1/45). Of all the LGAs visited, Abeokuta north and Abeokuta south had the highest incidence of YMV and DaBV, respectively. Mixed virus infections were also detected.     

The incidence and distribution of viruses infecting lettuce, cultivated Brassica and associated natural vegetation in Spain

A virus survey was conducted during the spring and autumn of 2001 and 2002 to determine the presence, prevalence and distribution in Spain of the viruses that are most commonly found infecting lettuce and Brassica worldwide. Crop plants showing virus symptoms from the principal lettuce and Brassica-growing regions of Spain, and some samples of the annual and perennial flora nearby, were tested by enzyme-linked immunosorbent assays using specific commercial antibodies against the following viruses: Alfalfa mosaic virus (AMV), Broad bean wilt virus 1 (BBWV-1), Beet western yellows virus (BWYV), Cauliflower mosaic virus (CaMV), Cucumber mosaic virus (CMV), Lettuce mosaic virus (LMV), Pea seed-borne mosaic virus (PSbMV), Turnip mosaic virus (TuMV) and Tomato spotted wilt virus (TSWV). Samples were also tested with a Potyvirus genus antibody. Virus incidence was much lower in spring than in autumn, especially in 2001. In spring 2002, CMV and LMV were the most prevalent viruses in lettuce, while CaMV was the most important virus present in Brassica crops grown in Navarra, followed by CMV and BWYV. In the autumn, the spectrum of viruses was different; potyviruses were widespread in lettuce grown in Madrid, but TSWV and BWYV were predominant in the Murcia region. The prevalent Potyvirus detected in lettuce fields was LMV, but none of the samples collected were positive for PSbMV or TuMV. In Brassica crops, TSWV was the most abundant in autumn-sown crops, especially in the Navarra region. All of the viruses present in lettuce and Brassica were also frequently detected in their associated natural vegetation at the same time, suggesting that they probably play an important role as virus reservoirs. Sonchus spp. were particularly common and were frequently infected with CMV, LMV and BWYV. Another common species, Chenopodium album, was often infected with TSWV and BWYV. Multiple infections were common, especially in non-crop plants, and the most common combination was BWYV and TSWV. The role of weeds in the epidemiology of viruses that infect lettuce and Brassica crops in Spain is discussed.     

The incidence of viruses in wild Brassica nigra in Dorset (UK)

Using enzyme‐linked immunosorbent assays, the frequency of occurrence of six viruses was determined in Brassica nigra collected from five coastal sites in Dorset, spanning approximately 24 km. During 1998–2000, the viruses detected were: Turnip mosaic virus (genus Potyvirus) (TuMV), Turnip yellow mosaic virus (genus Tymovirus) (TYMV), Turnip crinkle virus (genus Carmovirus) (TCV), Turnip rosette virus (genus Sobemovirus) (TRoV), Beet western yellows virus (genus Polerovirus) (BWYV) and Cauliflower mosaic virus (genus Caulimovirus) (CaMV). Multiple infections were detected in some individuals (48/447). TuMV was detected infrequently over the three‐year period (5/597). A representative isolate of each virus was tested for its effects on glasshouse‐grown individuals from different half‐sib families of B. nigra from four of the sites. Whether inoculated manually or via aphids (Myzus persicae), TuMV caused a rapid (within 10 days) lethal systemic necrosis in the B. nigra seedlings except when they were near flowering at the time of inoculation. Each of the other viruses invaded systemically but were not lethal. Indeed, BWYV systemically invaded 13/19 glasshouse‐grown B. nigra seedlings but did not produce any visible symptoms. Otherwise, the isolates tested differed in their pathogenicity and in the symptoms they produced in infected B. nigra. With TYMV or TCV viral antigen concentration was closely linked to pathogenicity; for TRoV or CaMV, there was little or no difference in virus concentration between plants with and without symptoms. Substantial and reproducible differences were observed in sensitivity/susceptibility among B. nigra genotypes from different sites in Dorset challenged with the same virus isolate.     

Detection of Olive‐infecting Viruses in Tunisia

During a virus survey in autumn 2007 and spring 2008 of two Tunisian olive mother blocks, 175 olive samples were collected from 19 different cultivars and tested by RT‐PCR for the presence of Arabis mosaic virus (ArMV), Cherry leaf roll virus (CLRV), Cucumber mosaic virus (CMV), Olive latent ringspot virus (OLRSV), Olive latent virus 1 (OLV‐1), Olive latent virus 2 (OLV‐2), Olive leaf yellowing‐associated virus (OLYaV) and Strawberry latent ringspot virus (SLRSV), using specific sets of primers. The PCR‐negative samples were also subjected to dsRNA and mechanical transmission tests. PCR results indicated that c. 86% of the trees were infected with at least one virus, whereas visible bands were shown by 3 of 24 PCR‐negative samples in dsRNA analysis. OLYaV was the most prevalent virus (49.1%), followed by OLV‐1 (34.3%), CMV (25.7%), OLRSV (16.6%), CLRV (13.1%), SLRSV (7.4%) and OLV‐2 (6.9%), whereas ArMV was not detected. Very high infection rates were found in the two main oil cvs. Chemlali (84.6%) and Chétoui (86.9%).     

Detection and Molecular Characterization of Squash leaf curl virus (SLCV) in Jordan

Squash leaf curl virus (SLCV) was detected for the first time in Jordan using degenerated oligonucleotide primers. Two isolates of the virus, SLCV‐E and SLCV‐R, were detected using specific oligonucleotide primers in symptomatic Cucurbita pepo. SLCV was also found to occur naturally in Malva parviflora, which showed severe leaf curling, yellowing and stunting of the whole plants. The full‐length genomes of Squash leaf curl virus‐Malva (SLCV‐Malva) isolate were amplified using the bacteriophage Φ DNA polymerase enzyme. Nucleotide sequence analysis showed that SLCV‐Malva shared high nucleotide identity (98% and 97%) with SLCV‐EG and SLCV‐E from Egypt and USA, respectively. A survey using dot‐blot hybridization indicated that squash leaf curl disease occurred in all surveyed areas. The highest disease incidence (95%) was recorded in Dir Alla area, whereas disease incidence did not exceed 69% in squash samples collected from North Ghor.     

Molecular Characterization of a Distinct Begomovirus Infecting Euphorbia pulcherrima in China

Virus isolate G35 was obtained from Euphorbia pulcherrima showing leaf curl and vein thickening symptoms in Tianyang, Guangxi Province, China. The virus was transmitted by whiteflies to Nicotiana tabacum, Lycopersicon esculentum, Datura stramonium and E. pulcherrima. DNA‐A contains 2746 nucleotides, with two open reading frames (ORFs) in the virion‐sense DNA and four ORFs in the complementary‐sense DNA. When compared with the DNA‐A sequence of other begomoviruses, the total DNA‐A of isolate G35 was most closely related to that of Ageratum enation virus (79.9% sequence identity). However, the deduced coat protein of G35 is most like that of Pepper leaf curl virus from Bangladesh (94.9% amino acid sequence identity), and the AC1 of G35 is most like that of Cotton leaf curl Multan virus‐Okra (87.2% amino acid sequence identity). The molecular data showed that G35 is a distinct Begomovirus species, for which the name Euphorbia leaf curl virus (ELCV) is proposed.     

Patchouli virus X, a new potexvirus from Pogostemon clabin

This work describes a potexvirus obtained from patchouli, Pogostemon clabin, collected in São Paulo, Brazil in 1992. The plants showed mosaic and were infected by a potyvirus and a potexvirus. The potexvirus had a host range limited to Amaranthaceae, Solanaceae and Labiatae and was named Patchouli virus X (PatVX). PatVX was not transmitted by scissors pruning, in tobacco seeds or by Myzus nicotinae. The virus was purified and a specific antiserum with a titre great than 1:512 000 in dot‐ELISA was produced. The virus was serologically related to Papaya mosaic virus, Potato virus X, Viola mottle virus, White clover mosaic virus and Lily virus X. It had a coat protein of 21 071 ± 1 010 Mr. as determined by SDS‐PAGE. Immunolabelling tests demonstrated that fibrillar masses in the cytoplasm contain the coat protein. The presence of a dsRNA was detected in PatVX infected plants.     

Effects of the <Emphasis Type="Italic">Papaya meleira virus</Emphasis> on papaya latex structure and composition

Spontaneous latex exudation is the main symptom of papaya sticky (meleira) disease caused by the Papaya meleira virus (PMeV), a double-stranded RNA (dsRNA) virus. This paper describes different effects of PMeV on papaya latex. Latex samples were subjected to different histochemical tests to evaluate their chemical composition. Additionally, the integrity of the latex particles was assessed by transmission and scanning electron microscopy analysis. Biochemical and micro- and macro-element measurements were performed. PMeV dsRNA extraction was performed to evaluate the interaction of the virus with the latex particles. Sticky diseased latex was positive for alkaloid biosynthesis and showed an accumulation of calcium oxalate crystals. PMeV also increased H₂O₂ synthesis within sticky diseased laticifers. The protein, sugar and water levels were altered, probably due to chemical changes. The morphology of the latex particles was further altered; PMeV particles seemed to be bound to the latex particles. The alkaloid and H₂O₂ biosynthesis in the papaya laticifers indicate a papaya defense response against PMeV. However, such efforts failed, as the virus affected the plant latex. The effects described here suggest some advantages of the infection process, including facilitating the movement of the virus within the papaya plant.     

First Report of Tomato infectious chlorosis virus on Tomato in Bulgaria

Virus‐like chlorotic symptoms were observed on tomato plants, cv. Velocity, grown in a greenhouse, region of Plovdiv. Samples collected from the leaves with interveinal yellowing and with initial interveinal chlorosis were tested for virus presence. Only the samples collected from the upper leaves with slight interveinal chlorosis were positive for Tomato infectious chlorosis virus (TICV) in indirect ELISA. Further, RT‐PCR analysis with specific primers for Tomato chlorosis virus (ToCV) heat shock protein 70, for TICV heat shock protein 70 and for TICV minor capsid protein was positive for TICV in all tested samples. No signals were obtained with primers for ToCV. Phylogenetic analysis showed that the Bulgarian sequence of Hsp70 and a sequence of Greek isolate clustered together having the highest resampling score. Regarding CPm, the Bulgarian isolate was more relevant to the French isolate. The obtained results from phylogenetic analysis supported the idea of a close relationship between the Bulgarian and Greek isolates.     

Molecular Characterization of a Strain of Squash Leaf Curl China Virus from the Philippines

The complete nucleotide sequence of infectious cloned DNA components (A and B) of the causal agent of squash leaf curl disease in the Philippines was determined. DNA‐A and DNA‐B comprise 2739 and 2705 nucleotides, respectively; the common region is 174 bases in length. Five ORFs were found in DNA‐A and two in DNA‐B. Partial dimeric clones containing DNA‐A and DNA‐B, constructed in a binary vector and transformed into Agrobacterium tumefaciens, induced systemic infection in agro‐inoculated pumpkin plants (Cucurbita moschata). The total DNA‐A sequence was most closely related to that of Squash leaf curl China virus (SLCCNV) (88% identity), although the existence of B component of SLCCNV has not been reported. The deduced coat protein was like that of SLCCNV (98% amino acid sequence identity) and the Philippines virus has low sequence identity to Squash leaf curl virus (SLCV) and Squash mild leaf curl virus (SMLCV) (63 and 64% total nucleotide sequence identities, respectively). From these results, we propose that the Philippines virus be designated Squash leaf curl China virus‐[Philippines] (SLCCNV‐[PH]).