Presence Of Cucumber Mosaic Virus Research Articles

Cucumber mosaic virus and turnip mosaic virus occurrence in garlic mustard in Ukraine

Garlic mustard (Alliaria petiolata) is an herbaceous biennial plant native to Europe. In Ukraine, in addition to becoming a serious invader, garlic mustard can serve as a host to several viruses, which may affect agricultural crops. In view of this, the purpose of the study was to identify the virome of garlic mustard growing in Ukraine. Plant samples collected in Kyiv regions were tested for the presence of cucumber mosaic virus (CMV), turnip mosaic virus (TuMV), turnip yellow mosaic virus (TYMV), watermelon mosaic virus II (WMV), and turnip crinkle virus (TCV) by serological and/or molecular methods. According to the results found in the present study, symptomatic A. petiolata obtained in 2021 were infected with CMV (60%), TuMV (20%), or co-infected with CMV + TuMV (20%). TYMV, WMV II, and TCV were not detected in any of the collected samples. The cDNA fragments encoded the coat protein (CP) gene of CMV and TuMV were sequenced and named as CMV-Ap and TuMV-Ap, respectively. In phylogenetic analysis, the CMV-Ap closely resembled the German isolate MW582807 (Sarracenia sp.), with 99.8% nucleotide identity and belongs to subgroup II of CMV. In the phylogenetic tree, TuMV-Ap clustered with isolates AP017803, AP017764, AP017791, and JQ073722, and represented the highest identity (98.6%) to Iranian isolate IRNTRa9 (AP017803) from Rapistrum rugosum and Turkish isolate TUR49 (AP017872) from Raphanus raphanistrum. The sequences of CMV-Ap and TuMV-Ap were deposited in the GenBank under Accession Numbers MZ540213 and OM799323, respectively. The results obtained in the study indicate the important role of infected garlic mustard as alternative host and natural reservoir of CMV and TuMV from which these economically important viruses can spread to other wild and cultivated plants. This is the first molecular evidence of TuMV infection in A. petiolata from Ukraine.

Incidence and detection of cucurbit mosaic disease in Assam

The cucurbits belong to the family Cucurbitaceous (gourd family) under the order Cucurbit ales which include many economically important vegetable and fruit species. Among various factors, viruses are one of the important factors reducing in quantity as well as quality of cucurbits. These crops are severely affected by the cucurbit mosaic disease worldwide, which is the major viral disease of the cucurbits. Considering the importance of the disease, the present investigation was taken up for incidence, detection, host range study and characterization of viruses associated with cucurbit mosaic disease in Assam. Survey was conducted in 5 districts of Assam (Sonitpur, Biswanath, Jorhat, Sivasagar, Dibrugarh) during 2021-22. Symptomatology revealed presence of various mosaic symptoms in the surveyed areas. Among the five surveyed locations, the symptoms observed in Sonitpur district were severe, whereas Biswanath district were moderate and mild symptoms were observed in Jorhat, Sivasagar, and Dibrugarh districts. Vector incidence study showed that aphid is the predominant vector. Molecular assay confirmed the presence of Cucumber Mosaic Virus in the collected cucurbit samples of surveyed areas; with severe incidence of mosaic disease in Sonitpur (90.91%), moderate in Biswanath (66.67%), mild in Jorhat (14.29%), Sivasagar (11%) and Dibrugarh (25%) districts. Molecular characterization of Cucumber mosaic virus cucurbit isolate from Jorhat district of Assam, was done through partial sequencing and similarity analysis showed 90.94 to 94.74 per cent sequence homology with different worldwide isolates of Cucumber mosaic virus.

Біологічні властивості вірусу огіркової мозаїки овочевих культур

Goal. To carry out diagnostics and to establish ways of spreading cucumber mosaic virus on the territory of Ukraine. Methods. A range of methods was used, which included visual diagnostics, enzyme immunoassay, method in various modifications, electron microscopy method, and statistical data processing method. Results. Monitoring of crops in different climatic regions of Ukraine for visual symptoms was carried out. Enzyme immunoassay revealed that the symptoms on the plants were caused by cucumber mosaic virus. Using electron microscopy, the morphological properties of the virus were studied, and spherical viral particles with a size of 29 nm were detected. Commercial seeds of different varieties of cucumbers, zucchini, tomatoes, and vegetable peppers were tested for the possibility of their viral contamination. Commercial seeds were found to be contaminated with cucumber mosaic virus antigens. Conclusions. Plants of the Cucurbitaceae and Solanaceae families were selected according to their characteristic symptoms. The presence of cucumber mosaic virus in Ukraine has been proved using electron microscopy and enzyme immunoassay. Virus-infected plants were diagnosed in agrocnosis of Vinnytsia, Odesa, and Kherson regions. Various commercial seed varieties of Cucurbitaceae and Solanaceae were tested for viral contamination. It was found that 37% of them were infected with cucumber mosaic virus and therefore became a potentially dangerous reservoir for viral infection and, as a consequence, a serious cause of outdoor vegetable disease.

Prisustvo i molekularna karakterizacija virusa mozaika krastavca u usevu šafranike u Srbiji

Safflower (Carthamus tinctorius L.) is an important oilseed crop belonging to the family Asteraceae. A total of 46 safflower samples were collected from Srbobran locality (South Backa District) in Serbia in 2015 and analysed for the presence of cucumber mosaic virus (CMV), alfalfa mosaic virus (AMV), and lettuce mosaic virus (LMV), using commercial double-antibody sandwich (DAS)-ELISA kits. Both viruses, CMV and AMV, were detected serologically in the collected samples. None of the analysed samples was found to be positive for LMV. The presence of CMV was further confirmed by mechanical transmission to test the plants including Chenopodium quinoa, C. amaranticolor, Nicotiana glutinosa, and Datura stramonium as well as C. tinctorius, confirming the infectious nature of the disease. Molecular detection of CMV was performed by amplification of a 871 bp fragment in all the tested samples, using the specific primers CMVCPfwd/CMVCPrev that amplify the entire coat protein (CP) gene and part of 3'and 5'-UTRs of CMV RNA 3. The RT-PCR products derived from the isolates 290Saff and 294Saff were sequenced (MH577791 and MH577792, respectively) and compared with the CMV sequences available in GenBank. Phylogenetic analysis based on CP gene sequences showed clustering of the selected isolates into three subgroups: IA, IB and II. Serbian CMV isolates found in safflower belong to subgroup II. To our knowledge, this is the first report on CMV infection of safflower in Serbia, which has the potential to cause substantial damage to safflower production and pose a threat to other economic crops grown in Serbia.

Characterisation of a subgroup IB isolate of Cucumber mosaic virus from Xanthosoma sp. in sub-Saharan Africa

Three tannia (Xanthosoma sp.) plants showing mosaic, mottling and vein chlorosis symptoms were observed in Buikwe district, Uganda. Next generation sequencing (NGS) of RNA extracted from the three plants revealed the presence of Cucumber mosaic virus (CMV). The complete genome sequence of this isolate, designated CMV-Xa, was assembled and sequence analyses based on the three genomic RNAs revealed that CMV–Xa belongs to subgroup IB. This is the first report of CMV infecting Xanthosoma sp. and also the first CMV isolate from subgroup IB detected from sub-Saharan Africa.

English

Watermelon plants can be naturally infected by several viruses in single or mixed infections, of which the diagnosis is difficult and require specific techniques. This study aims to detect and to verify the presence of Cucumber mosaic virus (CMV), Papaya ringspot virus type W (PRSV-W), Watermelon mosaic virus (WMV), Zucchini lethal chlorosis virus (ZLCV), Zucchini yellow mosaic virus (ZYMV), and the main cucurbit viruses in Brazil using multiplex reverse transcriptase polymerase chain reaction (RT-PCR) assay. Oligonucleotides were designed towards the conserved regions of the virus genomes. In the duplex-PCR, it was possible to detect all the virus combinations, except ZLCV with ZYMV. The amplified product sizes were 644 bp (CMV), 535 bp (WMV-2), 398 bp (PRSV-W), 244 bp (ZLCV), and 214 bp (ZYMV). To test the efficacy of the methodology, we analyzed plants with virus symptoms from four municipalities in the state of Tocantins, located at Brazilian Cerrado. Mixed infections were detected in 80% of samples in all the municipalities. The multiplex RT-PCR assay can be used to detect and differentiate watermelon viruses that affect crops in the state of Tocantins.   Key words: Cucumovirus, multiplex RT-PCR, Orthotospovirus, potyvirus, watermelon.

First Report of Cucumber mosaic virus Infecting Siam Tulip (Curcuma alismatifolia) in Florida

Siam tulip (Curcuma alismatifolia) has become an important crop for breeding new varieties owing to its high economic value as a tropical ornamental. In September 2017, C. alismatifolia potted plants, originally from Alachua, FL, were submitted to the Florida Department of Agriculture and Consumer Services, Division of Plant Industry, Gainesville, FL, for identification. The plants exhibited petal and bract necrosis. Initial serological testing for the presence of cucumber mosaic virus (CMV) using ELISA yielded positive results. To confirm pathogen identification, a two-step quantitative reverse transcription PCR, using primers targeting CMV-specific coat protein (amplicon of ∼450 bp), was performed. The PCR product, GenBank accession MG652768, had 99% nucleotide identity with CMV isolates in GenBank. Although all the tissues tested positive for CMV, it is unknown whether the necrotic symptoms were solely owing to CMV infection. CMV is known to infect a wide variety of plants in the ginger family, but to our knowledge, this is the first report of CMV in C. alismatifolia.

Occurrence of plant viruses on pepper cultivated in open fields in R. Macedonia and partial characterization of cucumber mosaic virus isolates

Screening of pepper plants cultivated in open fields for the presence of cucumber mosaic virus (CMV), alfalfa mosaic virus (AMV) and potato virus Y (PVY) in the Republic of Macedonia was performed by DAS-ELISA from 2012 to 2014. During the tested period, the predominant infection was by CMV, ranging from 34 to 61% of the tested samples, followed in 2012 by AMV and in 2013 and 2014 by PVY. Molecular detection and identification of CMV on selected pepper leaf samples was done by Reverse Transcription – Polymerase Chain Reaction (RT-PCR) and sequencing of the coat protein (CP) gene. Genetic relationships of the 18 CMV isolates generated in this study were compared with isolates from other parts of the world. Phylogenetic analysis was conducted, based on the partial sequences of the CP gene. In the constructed phylogenetic tree, all of the 18 Macedonian CMV isolates were clustered in the IA subgroup.

FIRST REPORT OF CUCUMBER MOSAIC VIRUS IN ANEMONE sp. IN GREECE

Virus-like symptoms, such as severe stunting and mosaic, were observed in a greenhouse-grown Anemone sp. (cv. Jerusalem), in Pella (northern Greece) in January 2014. Foliar samples were collected from 20 symptomatic and five symptomless plants and subjected to three initial tests: (i) DAS-ELISA to check for the presence of Cucumber mosaic virus (CMV), (ii) RT-PCR using generic primers for the detection of potyviruses (Pappu et al., 1993) and (iii) PCR for phytoplasmas (Gundersen and Lee, 1996). CMV was detected in all symptomatic plants but i n none of the asymptomatic ones, whereas PCR-based tests for potyviruses or phytoplasmas resulted negative. In order to verify the results of serological tests, a nested RT-PCR was performed on total RNA extracts from all 25 samples. Degenerate primers CMVup624a (5′- ATGGACAAATCTGRATC-3′) and CMVdo1244a (5′- TGRTGCTCRAYGTCKACATGA-3′), used in the first round of PCR were followed by the set CMVup624b (5′- GGACAAATCTGRATCTCCCAATGC-3′) and CMVdo1244b (5′- TGCTCRAYGTCRACATGAAG-3′), designed to amplify a 622 nt- long fragment of the viral coat protein gene. A mplicons of t h e expected size were obtained from all the symptomatic samples but not from the asymptomatic ones. Sequencing of two randomly chosen PCR products (GenBank accession Nos LN810056, LN810057) showed 99% nucleotide identity with CMV isolates from various hosts (KC763473, EU191027). Phylogenetic analyses showed that studied CMV isolates from Anemone sp. belong to sub-Group II. Finally, CMV was mechanically transmitted from the original symptomatic plants onto Cucurbita pepo and Nicotiana benthamiana, and its presence was confirmed by DAS-ELISA. To our knowledge this is the first report of CMV in Anemone sp. in Greece.

The presence of cucumber mosaic virus in pot marigold (Calendula officinalis L.) in Serbia

During 2014 a total of 67 pot marigold samples from five different localities in the Province in Vojvodina were collected and analysed for the presence of Cucumber mosaic virus (CMV) and Impatiens necrotic spot virus (INSV) using commercial double-antibody sandwich (DAS)-ELISA kits. CMV was detected serologically in all inspected localities in 67.16% collected samples. None of the analysed samples was positive for INSV. The virus was successfully mechanically transmitted to test plants including Chenopodium amaranticolor, C. quinoa, Datura stramonium, Nicotiana tabacum 'Samsun' and N. glutinosa, as well as pot marigold seedlings, confirming the infectious nature of the disease. The presence of CMV in pot marigold plants was further verified by RT-PCR and sequencing, using the specific primers CMV CPfwd/CMVCPrev that amplify coat protein (CP) gene. Phylogenetic analysis based on the CP gene sequences showed clustering of the selected isolates into three subgroups, IA, IB and II, and Serbian CMV isolates from pot marigold belong to subgroup II.

First Report of Cucumber mosaic virus in Tulipa sp. in Serbia.

Tulips (Tulipa sp. L.), popular spring-blooming perennials in the Liliaceae family, are one of the most important ornamental bulbous plants, which have been cultivated for cut flower, potted plant, garden plant, and for landscaping. In May 2013, during a survey to determine the presence of Cucumber mosaic virus (CMV, Cucumovirus, Bromoviridae) on ornamentals in Serbia, virus-like symptoms, including the presence of bright streaks, stripe and distortion of leaves, and reduced growth and flower size, were observed in an open field tulip production in the Krnjaca locality (a district of Belgrade, Serbia). Disease incidence was estimated at 20%. Symptomatic tulip plants were collected and tested for the presence of CMV by double-antibody sandwich (DAS)-ELISA using commercial diagnostic kit (Bioreba, AG, Reinach, Switzerland). Commercial positive and negative controls were included in each ELISA. Of the six tulip plants tested, all were positive for CMV. In bioassay, five plants of each Chenopodium quinoa, Nicotiana tabacum 'Samsun,' and N. glutinosa were mechanically inoculated with sap from selected ELISA-positive sample (79-13) using 0.01 M phosphate buffer (pH 7). Chlorotic local lesions on C. quinoa, and severe mosaic and leaf malformations on N. tabacum 'Samsun' and N. glutinosa, were observed 5 and 14 days post-inoculation, respectively. All mechanically inoculated plants were positive for CMV in DAS-ELISA testing. For further confirmation of CMV presence in tulip, total RNAs from all ELISA-positive symptomatic tulip plants were extracted with the RNeasy Plant Mini Kit (Qiagen, Hilden, Germany). Reverse transcription (RT)-PCR was performed with the One-Step RT-PCR Kit (Qiagen) using specific primer pair CMVCPfwd and CMVCPrev (1), which flank conserved fragment of the RNA3 including the entire coat protein (CP) gene and part of 3'- and 5'-UTRs. Total RNAs obtained from the Serbian watermelon CMV isolate (GenBank Accession No. JX280942) and healthy tulip leaves served as the positive and negative controls, respectively. The RT-PCR products of 871 bp were obtained from all six samples that were serologically positive to CMV, as well as from the positive control. No amplicon was recorded in the healthy control. The amplified product which derived from isolate 79-13 was purified (QIAquick PCR Purification Kit, Qiagen), directly sequenced in both directions using the same primer pair as in RT-PCR, deposited in GenBank (KJ854451), and analyzed by MEGA5 software (4). Sequence comparison of the complete CP gene (657 nt) revealed that the Serbian isolate 79-13 shared the highest nucleotide identity of 99.2% (99% amino acid identity) with CMV isolates from Japan (AB006813) and the United States (S70105). To our knowledge, this is the first report on the occurrence of CMV causing mosaic on Tulipa sp. in Serbia. Taking into account vegetative reproduction of tulips and the large scale of international trade with tulip seeding material, as well as wide host range of CMV including a variety of ornamentals (2,3), this is a very important discovery representing a serious threat for the floriculture industry in Serbia.

First Report of Bean common mosaic virus Infecting Azuki Bean (Vigna angularis) in China.

Azuki bean (Vigna angularis Ohwi & Ohashi) is one of the traditional grain legumes in China. From 2010 to 2013, mosaic and crumpling symptoms on leaves and stunting, all typical symptoms of a viral disease, were observed on cultivars CWA030, CWA221, and JCA002 of azuki bean with incidence rates of 30 to 100% and yield losses of 50 to 95% in the three fields of Changping district, Beijing. To identify the possible viral pathogen(s), 21 symptomatic leaf samples from different cultivars were collected and total RNA was extracted from the samples and subjected to RT-PCR testing with degenerate primers targeting portions of the coding regions of Cucumovirus capsid protein (CP) (1) and Potyvirus NIb (2); these viruses had been reported in azuki bean. Fragments of 940 bp and 350 bp corresponding to Cucumovirus CP and Potyvirus NIb, respectively, were amplified from all the samples collected. Sequencing of the PCR products from nine samples, followed by BLAST analysis, confirmed the presence of Cucumber mosaic virus (CMV) and Bean common mosaic virus (BCMV). All the samples tested were also positive with direct antigen coating (DAC)-ELISA using specific antiserum to CMV or BCMV (Agdia, Elkhart, IN). The CMV CP gene (GenBank Accession No. KJ467817) shared 99% sequence identity with a China CMV isolate (DQ873558). To further characterize the BCMV strain found, fragments of 3,388 bp spanning BCMV NIa, NIb, CP and 3'UTR regions were amplified with another primer set, BCMV-F (5'-AGCAAGTCAATTTACAAGGGACTTC-3') and BCMV-R (5'-GGAACAACAAACATTGCCGTAGCTAC-3') from three samples, and three independent clones from each sample were sequenced. Sequence analysis revealed that this segment (KJ467816) shared 98% identity with the BCMV azuki bean strain (U60100). To the best of our knowledge, this is the first report of BCMV, together with CMV, naturally infecting azuki bean in China. Further attention should be paid to this emerging viral disease and measures should be taken to control the spread of BCMV.

First Report of Banana mild mosaic virus Infecting Plantain in Ivory Coast.

Plantain (Musa sp., genomic group AAB) is an important crop for millions of the world's poorest people. In Ivory Coast, it is the second most consumed food and an important source of income for farmers. Between 2010 and 2011, a survey for viruses infecting plantain (AAB) was conducted in 10 major plantain-growing regions located in eastern (Abengourou), middle-western (Bouaflé, Daloa, Issia, Oumé, Sinfra, Zuenoula), central (Yamoussoukro), and southern (Aboisso, Gagnoa) Ivory Coast. Leaf samples showing yellow streaks or mild chlorotic streaks were collected and dried on CaCl2 for storage. A representative sample from each location was selected and tested for the presence of Cucumber mosaic virus (CMV, genus Cucumovirus), Banana streak virus (BSV, genus Badnavirus), Banana mild mosaic virus (BanMMV, family Flexiviridae), and Banana bract mosaic virus (BBrMV, genus Potyvirus). Immunocapture (IC)-PCR was used for the detection of BSV while reverse transcription (RT)-PCR was used for the detection of CMV, BanMMV, and BBrMV. The following primers sets were used: BSV cl1 and BSV cl2 (1), CMV 3' and CMV 5' (3), BanMMV BanCP1 and BanCP2 (4), BBrMV Bract N2 and Bract NR (2). BanMMV was detected as mixed infections with BSV in the 10 tested samples, one of which also contained CMV. To confirm the identity of the amplification products from the BanMMV primers, one cDNA fragment was directly sequenced in the forward direction (Macrogen Inc., Seoul, South Korea). BLAST search in GenBank revealed that the partial coat protein (CP) sequence of the Ivorian isolate shared 80 to 88% nucleotides and 81 to 92% deduced amino acid similarities with BanMMV isolates. In contrast, partial CP sequence of the Ivorian isolate had less than 40% deduced amino acid sequence identity with other Flexiviridae CP sequence. The partial CP sequence of the Ivorian BanMMV isolate was deposited in GenBank under Accession No. JX014304. To further confirm the identification, all the samples were tested by plate trapped antigen (PTA)-ELISA with rabbit polyclonal antiserum specific to BanMMV (obtained from B. E. Lockhart, University of Minnesota, U.S.A.) and anti-rabbit IgG (Sigma-Aldrich, Belgium/A3687). The 10 samples reacted positive for BanMMV by ELISA. CMV and BSV have been reported in Ivory Coast, but to our knowledge, this is the first report of BanMMV in the country. The detection of BanMMV in association with BSV or CMV in mixed infection in 10 locations which are important plantain growing areas is a first step in the evaluation of the impact of virus diseases on plantain production in this country.

Characterisation of a satellite RNA of Cucumber mosaic virus that induces chlorosis in Capsicum annuum

The presence of Cucumber mosaic virus (CMV) satellite RNA dramatically changes symptoms on some hosts. A satellite RNA present in a strain of CMV (PepY-CMV) that induced chlorosis in pepper (Capsicum annuum) was shown to induce chlorosis in pepper in combination with another strain (Fny-CMV) that by itself induced a green mosaic symptom. The location of sequences within the PepY satellite RNA (PepY-satRNA) of CMV that conferred the ability to induce chlorosis on pepper plants were analyzed by exchanging sequence domains between cDNA clones of PepY-satRNA and an attenuated mosaic satellite RNA (Paf-satRNA), as well as site-directed mutagenesis of various clusters of the 22-nt sequence differences between the two satellite RNAs in the delimited central domain. The symptoms induced by site-directed mutants of PepY-satRNA and Paf-satRNA in the presence of Fny-CMV demonstrated an insertion within PepY-satRNA of 11 nt at positions 86-96 relative to Paf-satRNA determined the chlorosis-inducing phenotype. Within the chlorosis-inducing domain, deletion of nucleotides did not affect the satRNA replication but abolished the ability of PepY-satRNA to elicit chlorosis symptom. Conversely, a mutant satellite RNA derived from Paf-satRNA in which eleven nucleotides were inserted indicated that sequences of 11 nucleotides were found to be sufficient for chlorosis induction in pepper.

First Report of Cucumber mosaic virus and Turnip vein-clearing virus in Dichondra repens in France, Italy, and China.

During surveys of Dichondra repens (kidneyweed, family Convolvulaceae) turfs in public gardens of the Franco-Italian Riviera from 1993 to 2003, leaf mosaic and yellow ringspot symptoms have been observed in Antibes, Menton, Nice, and Vallauris (France) and San Remo and La Mortola (Italy). Isolates from these six locations and from two locations in China (Shanghai and Kunming) have revealed the presence of Cucumber mosaic virus (CMV) based on the behavior of a range of manually inoculated plants (1), the observation of 30 nm isometric particles in semipurified extracts of inoculated Nicotiana tabacum 'Xanthi' plants with the electron microscope, and positive reactions in double antibody sandwich (DAS)-ELISAs with specific polyclonal antibodies. All isolates were shown to belong to group II of CMV isolates (3) by double-immunodiffusion analysis. CMV was previously identified in D. repens in California in 1972 (4). Following isolation from local lesions on Vigna unguiculata and multiplication in 'Xanthi' tobacco plants, two of the isolates were used to inoculate seedlings of D. repens manually or by Aphis gossypii aphids. Two months later, all inoculated plants showed symptoms similar to those previously observed and were positive in DAS-ELISA. In 2000, a D. repens sample collected in Antibes showing similar symptoms as above, induced necrotic local lesions in inoculated 'Xanthi' plants in 48 h, followed by systemic mosaic symptoms typical of CMV, therefore revealing the presence of a second virus. That virus was separated from CMV in apical, noninoculated leaves of Chenopodium quinoa and then used to inoculate a range of test plants. It was infectious in most plants of the families Solanaceae (including Cyphomandra betacea) and Brassicaceae, together with in Chenopodium amaranticolor, C. quinoa, Claytonia perfoliata, Convolvulus spp. 'Belle de jour', Digitalis purpurea, Gomphrena globosa, Ocimum basilicum, Plantago lanceolata, and Valerianella olitoria. It induced asymptomatic systemic infections in D. repens. Numerous, rod-shaped, 300 nm long particles were observed in sap extracts of infected plants with the electron microscope, suggesting the presence of a tobamovirus. A set of primers polyvalent for tobamoviruses (2) allowed the amplification of a DNA product of approximately 800 bp through reverse transcription-PCR performed with total RNA extracts from inoculated 'Xanthi' plants. The DNA product was cloned and sequenced (GenBank Accession No. EU927306) revealing that the virus belonged to a tobamovirus lineage including Ribgrass mosaic virus and viruses infecting cruciferous plants (Turnip vein-clearing virus [TVCV] and Youcai mosaic virus) and was closest to TVCV (95% amino acid identity; GenBank Accession No. NC_001873). To our knowledge, this is the first report of TVCV in D. repens.

First Report of Cucumber mosaic virus in Viola hederacea in France and Italy.

Viola hederacea Labill. (Australian violet or trailing violet), family Violaceae, is native to eastern Australia and used for its ornamental quality in humid conditions, especially in terrariums. Mosaic and chlorotic ringspots associated with a mild crinkling on leaves of V. hederacea were observed in gardens in southeast France and La Mortola, Italy in 2001 and 2003, respectively. These symptoms were different from the small chlorotic spots reported in the same species in Queensland, Australia, from which a rhabdovirus was isolated (1). In samples collected from both locations, presence of Cucumber mosaic virus (CMV) was suspected on the basis of the symptoms exhibited by a range of inoculated plants (2) and the observation of isometric particles of approximately 30 nm with the electron microscope in sap preparations. This was confirmed by positive reactions with CMV polyclonal antiserum (3) in double-antibody sandwich-ELISA. Because of the viscosity of the V. hederacea extracts, 0.01% (w/v) of pectolyase in phosphate buffer was used to grind the samples and the homogenate was kept at room temperature for 2 h. In double-immunodiffusion analysis, each isolate was shown to belong to the group II of CMV. Tomato strains D and To were used as positive controls for CMV groups I and II, respectively. The virus was transmitted by aphids (Myzus persicae) in a nonpersistent manner from infected Nicotiana tabacum cv. Xanthi-nc plants to healthy Xanthi-nc plants. To determine if CMV was responsible for the symptoms observed, the French and Italian isolates were multiplied in Xanthi-nc tobacco plants after isolation from local lesions on Vigna unguiculata and mechanically inoculated to 10 plants of V. hederacea. A systemic vein clearing developed on leaves 10 days after inoculation, followed by mosaic and crinkling. CMV was detected in each plant 3 weeks after inoculation. To our knowledge, this is the first report of CMV in V. hederacea in France and Italy.

Tobacco mosaic virus in Jalapeno Pepper in New York.

Jalapeno pepper plants (Capsicum annuum cv. Jaladuro) grown in Erie County, New York expressed chlorotic oak-leaf patterns along the primary vein of fully expanded leaves. Fruit had patterns of irregular ripening with a bumpy surface. Symptom expression was most obvious in August 2002, when leaf and fruit abscission occurred. Symptomatic fruit samples were tested by western blot analysis for the presence of Cucumber mosaic virus (CMV), Potato virus Y (PVY), Pepper mottle virus (PepMoV), Tobacco etch virus (TEV), and Tobacco mosaic virus (TMV). A positive reaction for TMV, but none of the other viruses, was observed. Symptomatic leaf samples were tested by Agdia, Inc. (Elkhart, IN) for Alfalfa mosaic virus, CMV, Impatiens necrotic spot virus, Pepper mild mottle virus, PepMoV, PVY, TEV, TMV, Tobacco ringspot virus, Tomato ringspot virus, and TSWV and for potyviruses using a group-specific test. The Agdia test confirmed that the pepper plants were infected with TMV. The pepper field where the original samples were collected was surveyed for TMV-infected plants. Fifty symptomatic plants expressing foliar and fruit symptoms similar to those originally tested, and 50 asymptomatic plants were sampled by collection of three leaves per plant and tested using enzyme-linked immunosorbent assay for the presence of TMV. All symptomatic plants and 18% of asymptomatic plants tested positive for TMV. To our knowledge, this is the first occurrence of TMV causing losses in commercially grown pepper in New York.

First Report of a Cucumber mosaic virus-Associated Satellite RNA in Lobelia erinus.

Lobelia (Lobelia erinus L.) is a common herbaceous annual used in flower beds and hanging baskets. The plant blooms from early to late summer. In the summer of 2000, Lobelia plants expressing virus-like symptoms were collected from a greenhouse-based production site in Ohio. Affected plants expressed a mild leaf mosaic and stunting. Viral-associated dsRNA was isolated from 7 g of symptomatic leaf tissue (1). Four dsRNAs were observed at 3.9, 3.0, 2.25, and 1.05 kb indicating the presence of a Cucumovirus. A fifth dsRNA at 0.75 kb also was observed, consistent with the presence of a satellite RNA. Enzyme-linked immunosorbent assay (ELISA) analysis (Agdia, Inc., Elkhart, IN) of symptomatic Lobelia tissue confirmed the presence of Cucumber mosaic virus (CMV). A (S)CARNA-5 (-) cDNA clone (American Type Culture Collection #45124) was labeled with digoxygenin (DIG) as per the manufacturer's instructions (Genius II DIG-DNA Labeling Kit, Boehringer Mannheim) and used as a diagnostic probe to detect this satellite RNA. Northern hybridization confirmed the identity of the satellite RNA (2). This is the first report of any satellite RNA associated with a virus infection in Lobelia and the first report of CMV in this host in Ohio.

First Report of Cucumber Mosaic Virus in Eryngium amethystinum, Canna spp., and Aquilegia hybrids in Ohio.

Eryngium amethystinum (amethyst sea holly) is a herbaceous plant commonly grown as an ornamental perennial in U.S.D.A. hardiness zones 3 to 8. The plant thrives in dry areas with infertile soils and the flowers are often used in dried floral arrangements. Canna spp. (Canna), soft perennials (U.S.D.A. zone 9 and above), are becoming popular flowering plants because of their bright flowers and spectacular foliage. There are a variety of species that fall under the heading Canna spp., of which the most popular are C. glauca, C. indica, C. edulis, and C. iridiflora. Hybrids of Aquilegia (garden columbine), a hardy perennial (U.S.D.A. zones 3 to 9), flower in late spring through early summer. The genus is made up of a wide variety of cultivars. E. amethystinum exhibiting severe mosaic, yellowing, and stunting, along with Canna plants exhibiting severe stunting, chlorotic and distorted foliage, and mosaic, and garden columbine plants exhibiting stunting, leaf curl, chlorosis, and mosaic, collected from commercial plantings throughout the central Ohio area, were analyzed for the presence of virus infection with viral-associated, double-stranded RNA (dsRNA) analysis. dsRNA analysis resulted in a banding profile typical of that seen with members of the cucumovirus family of plant viruses. Plants positive for cucumovurus-like dsRNA were tested with a direct antibody sandwich enzyme-linked immunosorbent assay (ELISA). ELISA results confirmed the presence of cucumber mosaic virus (CMV) in all symptomatic plants tested. No evidence of dsRNA or CMV was found in any of the asymptomatic plants tested. Because all of these hosts are common in the perennial garden, they could serve as a reservoir host of CMV for other plants in the garden. This is the first report of CMV in E. amethystinum, Canna spp., and Aquilegia hybrids in Ohio.

Differential Effects of Satellite RNA on the Accumulation of Cucumber Mosaic Virus RNAs and Their Encoded Proteins in Tobacco vs Zucchini Squash with Two Strains of CMV Helper Virus

The presence of cucumber mosaic virus (CMV) satellite RNA usually reduces the yield of accumulated helper virus, although more so in solanaceous than in cucurbit hosts. The accumulation of viral RNA and viral-encoded proteins of two strains of CMV (Fny- and Sny-) known to differ in their ability to support satellite RNA in zucchini squash was examined in squash and tobacco to determine the effect of satellite RNA on the accumulation of viral-associated components. In the absence of satellite RNA, Fny- and Sny-CMV showed similar levels of accumulation of RNA at 7 days postinoculation (p.i.), but by 14 days p.i, the Fny-CMV RNAs accumulated to lower levels than did both strains at 7 days p.i., in either host. The levels of accumulated Sny-CMV-encoded proteins were higher than those encoded by Fny-CMV in tobacco, but not squash plants, at 7 days p.i. At 14 days p.i., for Fny-CMV vs Sny-CMV, there were differences in the levels of accumulation of most CMV-encoded proteins in both hosts, more exacerbated in tobacco vs squash. The effect of satellite RNA was to intensify these differences; that is, by 7 days p.i., satellite RNA reduced the accumulation of Fny-CMV RNAs 1 and 2 and their encoded proteins in both tobacco and squash but had little or no effect on the accumulation of Sny-CMV RNAs or encoded proteins. By 14 days p.i., the levels of accumulation of all Fny-CMV RNAs and encoded proteins were severely reduced in both hosts, and the levels of accumulation of Sny-CMV RNAs 1 and 2 and their encoded proteins were also reduced in tobacco, but not squash, Sny-CMV did not support satellite RNA accumulation in squash plants or protoplasts. Satellite RNA did not appear to have a direct effect on the movement of either CMV strain. Rather, accumulation studies in tobacco protoplasts indicated that the difference in response of Fny-CMV vs Sny-CMV to satellite RNA in tobacco was due to the extent to which satellite RNA affected the levels of RNA 1, and to a lesser extent RNA 2, and their encoded proteins, 1a and 2a, both components of the CMV replicase.