Serological diagnosis and host range studies of important viral diseases of a few cucurbitaceous crops in Maharashtra,India

Around 39 well characterised viruses affect cucurbits crops in developing countries and their viral diversity may be the consequence for genetic and ecological diversity of their hosts. Indeed, cucurbits are grown in variety of climatic, environmental and agricultural conditions, and this may provide more or less favourable conditions for the specific viruses or their hosts. The presence of various viral diseases caused by different viruses in Maharashtra was studied from the infected samples collected from cucurbits and melons during the survey conducted in 2009–2010 in different locations. The virus isolates collected from various cucurbitaceous crops were established and their host ranges were studied by sap transmission. The study revealed Cucumber Mosaic Virus (CMV), Zucchini yellow mosaic virus (ZYMV), Watermelon mosaic virus (WMV) and Cucumber green mottle mosaic virus infections predominately found in Nashik region, and Watermelon bud necrosis virus (WBNV), CMV, ZYMV, WMV and Watermelon silver mottle virus (WSMoV) infections in Aurangabad and Paithan regions. In Sangamner region, the crop was mostly affected by WBNV, ZYMV and WSMoV, and CMV was found only in Sillod region. The protocols for performing sap transmission tests in assay hosts were standardised for ZYMV, CMV and WBNV. Using direct antigen-coating enzyme-linked immunosorbent assay, of all the plant parts, young leaves were found to have high concentration of virus and suitable for virus detection in screening programmes. CMV and ZYMV was found to have high concentration of virus and suitable for virus detection in screening programmes.     

Identification and distribution of viruses infecting sweet potato in Kenya

Four hundred and forty-eight symptomatic and 638 asymptomatic samples were collected from sweet potato fields throughout Kenya and analysed serologically using antibodies to Sweet potato feathery mottle virus (SPFMV), Sweet potato chlorotic stunt virus (SPCSV), Sweet potato mild mottle virus (SPMMV), Cucumber mosaic virus (CMV), Sweet potato chlorotic fleck virus (SPCFV), Sweet potato latent virus (SwPLV), Sweet potato caulimo-like virus (SPCaLV), Sweet potato mild speckling virus (SPMSV) and C-6 virus in enzyme-linked immunosorbent assays (ELISA). Only SPFMV, SPMMV, SPCSV, and SPCFV were detected. Ninety-two percent and 25% of the symptomatic and asymptomatic plants respectively tested positive for at least one of these viruses. Virus-infected plants were collected from 89% of the fields. SPFMV was the most common and the most widespread, detected in 74% of the symptomatic plants and 86% of fields surveyed. SPCSV was also very common, being detected in 38% of the symptomatic plants and in 50% of the fields surveyed. SPMMV and SPCFV were detected in only 11% and 3% of the symptomatic plant samples respectively. Eight different combinations of these four viruses were found in individual plants. The combination SPFMV and SPCSV was the most common, observed in 22% of symptomatic plants. Virus combinations were rare in the asymptomatic plants tested. Incidence of virus infection was highest (18%) in Kisii district of Nyanza province and lowest (1%) in Kilifi and Malindi districts of Coast province.     

Viruses infecting sweet potato in Rwanda: occurrence and distribution

A survey of sweet potato virus diseases was conducted in the major sweet potato production areas in low, medium and high altitude zones of Rwanda. A total of 205 symptomatic and 103 asymptomatic samples were collected from 51 sweet potato fields and assayed for Sweet potato feathery mottle virus (SPFMV), Sweet potato chlorotic stunt virus (SPCSV), Sweet potato mild mottle virus (SPMMV), Sweet potato chlorotic fleck virus (SPCFV), Sweet potato latent virus (SwPLV), Sweet potato caulimo‐like virus (SPCaLV) and Cucumber mosaic virus (CMV) using nitrocellulose membrane enzyme‐linked immunosorbent assay. The viruses detected in the samples were SPFMV, SPMMV, SPCSV, SPCFV and SwPLV. Viruses were detected in 83% and 31% of the symptomatic and asymptomatic samples, respectively. SPFMV was detected in 49% of the samples. SPCSV, the second most common virus, was detected in 28% of samples collected from 73% of the fields. About 19% of the samples were tested positive for SPMMV. Thirteen combinations of multiple virus infections were detected in the samples. Viruses were detected in samples from all the fields surveyed, and the frequency of detection was greatest in samples from low altitude zones.     

Transmission Studies with Cucumber Green Mottle Mosaic Virus

Cucumber green mottle mosaic virus may spread in bottlegourd under field conditions through soil contaminated with infected plant debris followed by contact. No seed transmission was noticed in bottlegourd (Lagenaria siceraria) or vegetable marrow (Cucurbita pepo) although pollen grains and cotyledons from infected bottlegourd flowers or seeds, respectively, contained negliginle amounts of virus. Cucumber leaf beerles (Raphidopalpa fevicollis) are probable vectors since their regurgitated fluid and excretes contained infective virus particles. No vector fungi were found in the soil around infected bottlegourd plants.     

Resistance to six viruses in the legume goat's rue {Galega orientalis Lam.)

Plants from 2nd to 6th year leys of the legume goat's rue (Galega orientalis Lam.) were tested for infection with bean yellow mosaic (BYMV), bean common mosaic (BCMV), alfalfa mosaic (AMV), broad bean stain (BBSV), red clover mottle (RCMV) and cucumber mosaic (CMV) viruses by enzyme-linked immunosorbent assay (ELISA), electron microscopy, and by sap-inoculation to various test plant species. No virus infections were observed in goat's rue in the field. Glasshouse-grown seedlings of goat's rue were inoculated with the above viruses. No virus was detected in the inoculated plants. The results suggest that goat's rue is extremely resistant to the above six viruses which are important in other forage legumes.     

Expressed sequence tag analysis of Antarctic hairgrass Deschampsia antarctica from King George Island, Antarctica

Deschampsia antarctica is the only monocot that thrives in the tough conditions of the Antarctic region. It is an invaluable resource for the identification of genes associated with tolerance to various environmental pressures. In order to identify genes that are differentially regulated between greenhouse-grown and Antarctic field-grown plants, we initiated a detailed gene expression analysis. Antarctic plants were collected and greenhouse plants served as controls. Two different cDNA libraries were constructed with these plants. A total of 2,112 cDNA clones was sequenced and grouped into 1,199 unigene clusters consisting of 243 consensus and 956 singleton sequences. Using similarity searches against several public databases, we constructed a functional classification of the ESTs into categories such as genes related to responses to stimuli, as well as photosynthesis and metabolism. Real-time PCR analysis of various stress responsive genes revealed different patterns of regulation in the different environments, suggesting that these genes are involved in responses to specific environmental factors.     

Disease intensity of some tomato viroses in Serbia

In this paper we present the data on the disease intensity of the tomato plants grown in glass and plastic-houses, and in the open field. The infection was caused by the following viruses: Tomato mosaic virus (ToMV), Tobacco mosaic virus (TMV), Tomato spotted wilt virus (TSWV), Alfalfa mosaic virus (AMV), Potato virus X (PVX), Potato virus Y (PVY), Tomato black ring virus (TBRV), Tomato ringspot virus (ToRSV), Tomato aspermy virus (TAV), and Cucumber mosaic virus (CMV). These viruses represented most frequent tomato pathogens in Serbia. According to the obtained results, it could be concluded that 92.94% of the tested tomato plants grown in glass and plastic-houses, and 89.82% grown in the open field were infected by one of the above viruses. Most of the plant samples were infected by two or more viruses. The most frequent viruses — tomato pathogens in Serbia were ToMV, PVY and TMV.     

Nucleotide sequence of Chinese rape mosaic virus (oilseed rape mosaic virus), a crucifer tobamovirus infectious on Arabidopsis thaliana

The complete nucleotide sequence of Chinese rape mosaic virus has been determined. The virus is a member of the tobamovirus genus of plant virus and is able to infect Arabidopsis thaliana (L.) Heynh systemically. The analysis of the sequence shows a gene array that seems to be characteristic of crucifer tobamoviruses and which is slightly different from the one most frequently found in tobamoviruses. Based on gene organization and on comparisons of sequence homologies between members of the tobamoviruses, a clustering of crucifer tobamoviruses is proposed that groups the presently known crucifer tobamovirus into two viruses with two strains each. A name change of Chinese rape mosaic virus to oilseed rape mosaic virus is proposed.Abbreviations 2-ME 2-mercaptoethanol - EDTA ethylenediaminetetraacetic acid - SDS sodium dodecyl sulfate - UTR untranslated region - MP movement protein - CP capsid protein - CRMV Chinese rape mosaic virus - TVCV turnip vein clearing virus - PaMMV paprika mild mottle virus - PMMV-I pepper mild mottle virus (Italian isolate) - PMMV-S pepper mild mottle virus (Spanish isolate) - ToMV tomato mosaic virus - TMV tobacco mosaic virus - TMGMV tobacco mild green mosaic virus - ORSV odontoglossum ringspot virus - SHMV sunn hemp mosaic virus - CGMMV cucumber green mottle mosaic virus - ORMV oilseed rape mosaic virus     

Preliminary studies on sap-transmissible viruses of red clover (Trifolium pratense L.) in England and Wales

Red clover plants, collected from nine widely separated permanent pastures in England and Wales, were tested for sap-transmissible viruses. Viruses were identified by the symptoms they caused in test plants, by electron microscopy, and by serological tests. Of the 265 plants tested 14% were infected. Only pea mosaic virus was common and widespread; it was found in 8% of the plants, and in seven of the fields. Other viruses isolated were arabis mosaic, bean yellow mosaic, red clover mottle, and red clover vein mosaic; only red clover mottle virus produced diagnostic symptoms in red clover. No viruses were detected in seedlings grown from seed from eighty-nine commercial seed crops. Attempts to transmit red clover mottle virus by the Collembolan Sminthurus viridis L., which is common on red clover, failed.     

Immunodiagnosis of plant viruses by a virobacterial agglutination test

A new virobacterial agglutination (VBA) test for the immunodiagnosis of plant viruses is described. The test is based on the agglutination of Staphylococcus aureus cells by virus particles after treatment of the cells with homologous antiserum. The agglutination occurs within 1–5 min. The sensitivity of the test is 0·1-0·4 μg virus/ml and is not affected by the shape of the virus particle. The use of affinity purified antibodies for sensitisation of S. aureus cells increases the sensitivity of the reaction 50-fold and enables the detection of tobacco mosaic and cucumber green mottle mosaic viruses at a concentration of 2 ng/ml. The VBA test allows the estimation of potato viruses X, S, M and Y in the eyes and sprouts of infected tubers and in the leaves of infected plants. The diagnosis of carnation mottle virus in carnation plants and of mushroom viruses in mushroom (Agaricus bisporus) fruit-bodies and mycelium are also described.     

黄瓜花叶病毒卫星RNA人工突变体的构建及其侵染性测定

从1个369nt的黄瓜花叶病毒(Cucumbermosaicvirus,CMV)卫星RNA的cDNA出发,采用DNA改组技术构建人工突变体,经过体外转录,将其与不携带卫星RNA的黄瓜花叶病毒株进行假重组,鉴定突变体的生物活性,结果显示所获得的4个卫星RNA的点突变子MS1、MS5、MS6和MS11中,只有MS11仍然具有复制能力;而其他3个点突变子,尽管均只有1个位点的替换,却不能在辅助病毒作用下复制。序列比较分析发现MS11的突变位点位于卫星RNA变异区内,发生的突变与自发突变一致,其他3个突变子的突变位点发生在卫星RNA的高度保守区。而且通过侵染性试验证实突变子MS11与野生型Yi没有明显的差异。由此可推测卫星RNA序列中的高度保守区与卫星RNA的生物活性密切相关,个别碱基的突变会导致RNA二级结构的改变,进而引起其复制能力或稳定性的完全丧失。     

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.     

黄瓜绿斑驳花叶病毒辽宁分离物全基因组序列测定

以感病组织总RNA为模板,采用RT-PCR方法扩增并测定黄瓜绿斑驳花叶病毒(Cucumber green mottle mosaic virus,CGMMV)辽宁分离物(CGMMV-LN)的基因组全序列。CGMMV-LN基因组全长6 422 nt,5'非编码区(noncoding region,NCR)和3'NCR分别为59 nt和175 nt。CGMMV-LN编码的4个蛋白依次是186 kD和129kD的复制酶,29 kD的移动蛋白和17.4 kD的外壳蛋白。CGMMV-LN与其他4个CGMMV分离物基因组核苷酸序列同源性为97.6%～99.3%,与同属其他3种病毒基因组核苷酸序列同源性仅为61.7%～62.8%。基于186kD复制酶和外壳蛋白氨基酸序列的同源树显示:侵染葫芦科作物的烟草花叶病毒属病毒可分为2个亚组,亚组I包括所有CGMMV分离物,亚组II包括Kyuri绿斑驳花叶病毒(Kyuri green mottle mosaic virus,KGMMV)、黄瓜果实斑驳花叶病毒(Cucumber fruit mottle mosaic virus,CFMMV)和小西葫芦绿斑驳花叶病毒(Zucchini ...     

Detection of new viruses in alfalfa,weeds and cultivated plants growing adjacent to alfalfa fields in Saudi Arabia

A total of 1368 symptomatic plant samples showing different virus-like symptoms such as mottling, chlorosis, mosaic, yellow mosaic, vein clearing and stunting were collected from alfalfa, weed and cultivated plant species growing in vicinity of alfalfa fields in five principal regions of alfalfa production in Saudi Arabia. DAS-ELISA test indicated occurrence of 11 different viruses in these samples, 10 of which were detected for the first time in Saudi Arabia. Eighty percent of the alfalfa samples and 97.5% of the weed and cultivated plants samples were found to be infected with one or more of these viruses. Nine weed plant species were found to harbor these viruses namely, Sonchus oleraceus, Chenopodium spp., Hibiscus spp., Cichorium intybus, Convolvulus arvensis, Malva parviflora, Rubus fruticosus, Hippuris vulgaris, and Flaveria trinervia. These viruses were also detected in seven cultivated crop plants growing adjacent to the alfalfa fields including Vigna unguiculata, Solanum tuberosum, Solanum melongena, Phaseolus vulgaris, Cucurbita maxima, Capsicum annuum, and Vicia faba. The newly reported viruses together with their respective percent of detection in alfalfa, and in both weeds and cultivated crop plant species together were as follows: Bean leaf roll virus (BLRV) {12.5 and 4.5%}, Lucerne transient streak virus (LTSV) {2.9 and 3.5%}, Bean yellow mosaic virus (BYMV) {1.4 and 4.5%}, Bean common mosaic virus (BCMV) {1.2 and 4.5%}, Red clover vein mosaic virus (RCVMV) {1.2 and 4%}, White clover mosaic virus (WCIMV) {1.0 and 5%}, Cucumber mosaic virus (CMV) {0.8 and 3%}, Pea streak virus (PeSV) {0.4 and 4.5%} and Tobacco streak virus (TSV) {0.3 and 2.5%}. Alfalfa mosaic virus (AMV), the previously reported virus in alfalfa, had the highest percentage of detection in alfalfa accounting for 58.4% and 62.8% in the weeds and cultivated plants. Peanut stunt virus (PSV) was also detected for the first time in Saudi Arabia with a 66.7% of infection in 90 alfalfa samples collected from the surveyed regions during the last visit that tested negative to all the previously detected viruses.     

Taxonomic patterns in the host ranges of viruses among grasses, and suggestions on generic sampling for host-range studies

Analyses of published host-range data for certain viruses reveal correlations with taxonomic groupings of grasses. Barley yellow dwarf virus (BYDV), cocksfoot mottle and phleum mottle viruses are found to have infected greater proportions of the festucoid grasses than of the non-festucoids to which they were inoculated. By contrast, all strains of sugarcane mosaic virus (SCMV) and of the closely related maize dwarf mosaic virus (MDMV) infected more non-festucoids than festucoids. In addition, infected plants from grass groups containing higher concentrations of genera susceptible to BYDV, SCMV and MDMV usually show clear symptoms, whereas infected plants from less susceptible groups are frequently symptomless. Some viruses, such as barley stripe mosaic, brome mosaic, cocksfoot streak and ryegrass mosaic, show no apparent preferences for particular grass groups. Samples of grasses employed in host-range studies are usually strongly biased towards festucoids. It is suggested that viruses ought to be adequately tested against genera from all the major groups, and a classified list of grass genera suitable for host-range studies is provided.     

Characterization of antifreeze activity in Antarctic plants

Deschampsia antarctica and Colobanthus quitensis are the only vascular plants to have colonized the Maritime Antarctic, which is characterized by its permanently low temperature and frequent summer frosts. To understand how the plants survive freezing temperatures year-round, antifreeze activity was assayed in apoplastic extracts obtained from both non-acclimated and cold-acclimated Antarctic plants. By observing the shape of ice crystals grown in dilution series of the extracts, it was found that D. antarctica had antifreeze activity, but C. quitensis did not. D. antarctica exhibited antifreeze activity in the non-acclimated state and this activity increased after cold acclimation. The antifreeze activity in D. antarctica was labile to proteolysis and high temperature, active over a wide pH range, and associated with molecules greater than 10 kDa in molecular weight. These results show that D. antarctica produces antifreeze proteins that are secreted into the apoplast. When examined by SDS-PAGE, the apoplastic extracts from cold-acclimated D. antarctica exhibited 13 polypeptides. It is concluded that D. antarctica accumulates AFPs as part of its mechanism of freezing tolerance. Moreover, this is the first plant in which antifreeze activity has been observed to be constitutive.