scholarly journals First Report of Calibrachoa mottle virus Infecting Petunia

Plant Disease ◽  
2003 ◽  
Vol 87 (12) ◽  
pp. 1538-1538 ◽  
Author(s):  
H.-Y. Liu ◽  
J. L. Sears ◽  
M. Bandla ◽  
A. M. Harness ◽  
B. Kulemeka

Calibrachoa mottle virus (CbMV), a tentative carmovirus, was first isolated and reported by Liu et al. (1) from infected Calibrachoa plants. During the spring of 2003, petunia samples from Florida and California sent to testing services at Agdia, Inc (Elkhart IN) tested positive for CbMV by enzyme-linked immunosorbent assay (ELISA) and lateral flow immunoassay (ImmunoStrips). These samples also tested positive by carmovirus group-specific polymerase chain reaction (PCR) primers and by immunocapture PCR (2). RNA extracted from these samples with the RNeasy Plant Kit (Qiagen Inc., Valencia, CA) hybridized with a digoxigenin labeled probe derived from purified CbMV viral RNA. All plant samples that tested positive for CbMV were symptomless except one symptomatic sample that also tested positive for Tobacco mosaic virus. From samples that tested positive for CbMV only, mechanical inoculations were made to Chenopodium quinoa at a USDA-ARS greenhouse in Salinas, CA. Representative single, local lesions were used to inoculate additional C. quinoa plants. The resulting local lesions from these inoculations were freeze-dried and further used as virus inoculum (CbMV petunia). Similar inoculum was made with CbMV isolated from Calibrachoa plants (CbMV calibrachoa). Virus-free Petunia hybrida cultivars Surfinia ‘Baby Pink’ and Surfinia ‘Violet’ (Jackson and Perkins Inc., Somis, CA) were mechanically inoculated with CbMV petunia and CbMV calibrachoa. Four weeks postinoculation, all plants were tested using ELISA for the presence of CbMV. In greenhouse conditions, 14.3% of ‘Baby Pink’ plants were positive for CbMV petunia, whereas none were positive for CbMV calibrachoa. ‘Violet’ plants were 64.3 and 33.3% positive for CbMV petunia and CbMV calibrachoa, respectively. None of the positive plants expressed virus-like symptoms. Virus particles resembling those of CbMV were observed from infected petunia plants with transmission electron microscopy in leaf-dip preparations. To our knowledge, this is the first report of CbMV infecting petunia. Commercial reproduction of petunia plants and maintenance of genetic mother stock are usually by vegetative propagation. CbMV can be transmitted mechanically and is readily propagated along with its host. To produce healthy petunia plants, virus-free mother stock should be used, which requires regular screening of mother stock for CbMV. Reference: (1) H.-Y. Liu et al. Plant Dis. 87:167, 2003. (2) A. M. Harness et al. (Abstr.) Phytopathology 92:S34, 2002.

Plant Disease ◽  
2002 ◽  
Vol 86 (2) ◽  
pp. 186-186 ◽  
Author(s):  
J. Th. J. Verhoeven ◽  
T. M. Willemen ◽  
J. W. Roenhorst

In 2000, a breeding company submitted a tomato (Lycopersicon esculentum) sample from Guatemala for diagnosis. The plants showed necrotic lesions on leaves surrounded by some chlorosis, necrotic streaks on stems, and large superficial necrotic lesions on fruits. By mechanical inoculation of plant sap to different plant species, symptoms appeared in Capsicum annuum ‘Westlandse Grote Zoete’, Lycopersicon esculentum ‘Money-maker’, Nicotiana benthamiana, N. bigelovii, N. glutinosa, N. hesperis-67A, N. occidentalis-P1, N. tabacum ‘White Burley’, and Physalis floridana. Systemically infected leaves from N. occidentalis-P1 were used for all further experiments. Leaf dip preparations were analyzed by transmission electron microscopy and revealed the presence of filamentous virus particles typical of a potyvirus. Double-antibody sandwich enzyme-linked immunosorbent assay tests for Potato virus A, V, and Y, Tobacco etch virus, and Wild potato mosaic virus were negative. An antiserum (PepMoV/DSMZ As 0186) to Pepper mottle virus (PepMoV), however, gave a positive reaction. To obtain further evidence for the presence of this virus, the nucleotide sequence of the complete 3′ nontranslated region (3NTR) and the 3′ terminal part of the coat protein gene (3CPG) was determined using the set of degenerate primers P9502/CPUP (1). The obtained nucleotide sequence (approximately 700 bp) was deposited in GenBank under Accession No. AF440801. It showed 93 to 94% 3NTR and 90 to 93% 3CPG homology with the three sequences of PepMoV from pepper already present in GenBank. The two viruses with the next closest nucleotide sequence homology were Potato virus V and Potato virus Y showing up to 80 and 75% homology with the 3CPG and up to 53 and 48% homology with the 3NTR, respectively. Based on these results, we concluded that the virus isolated from the symptomatic tomato plants was PepMoV. Because of the relatively low homologies with the pepper isolates of PepMoV, this tomato isolate might be a separate strain of the virus. To our knowledge, this is the first report of the occurrence of PepMoV in tomato. Reference: (1) R. A. A. van der Vlugt et al. Phytopathology 89:148, 1999.


Plant Disease ◽  
2001 ◽  
Vol 85 (12) ◽  
pp. 1290-1290 ◽  
Author(s):  
A. R. Golnaraghi ◽  
N. Shahraeen ◽  
R. Pourrahim ◽  
Sh. Ghorbani ◽  
Sh. Farzadfar

During the summers of 1999 and 2000, 3,110 soybean (Glycine max) leaf samples were randomly collected from soybean fields in the Ardebil, Goletan, Khuzestan, Lorestan, and Mazandaran provinces of Iran. Tomato spotted wilt virus (TSWV) was detected in leaf samples by TSWV-specific polyclonal antibody (As-0526 and As-0580, DSMZ, Braunschweig, Germany) in double-antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA). Mechanical inoculation of 26 plant species (10 plants per species) and cultivars with extracts of positive leaf samples produced necrotic local lesions in Beta vulgaris, Chenopodium quinoa, C. amaranticolor, Gomphrena globosa, Phaseolus vulgaris cv. Talash, Vicia faba, and Vigna unguiculata cv. Mashad; produced systemic chlorosis followed by necrosis in Datura stramonium, D. metel, Nicotiana rustica, N. tabacum cv. Samsun, N. glutinosa, N. bentamiana, and Glycine max cv. Hill; and produced chlorosis, stunting, and bud necrosis in Arachis hypogaea (peanut). Plants developing these symptoms following mechanical inoculation with extracts from original soybean leaves were positive in ELISA for TSWV. ELISA results indicate that the overall incidence of TSWV on soybean in the five provinces was 5.4%. TSWV has been reported in potato (2) and tomato (1) from Iran, but to our knowledge, this is the first report of the occurrence of TSWV on soybean in Iran. References: (1) K. Bananej et al. Iran. J. Plant Pathol. 34:30, 1998. (2) R. Pourrahim et al. Plant Dis. 85:442, 2001.


Plant Disease ◽  
2006 ◽  
Vol 90 (1) ◽  
pp. 112-112 ◽  
Author(s):  
N. Borodynko ◽  
B. Hasiów ◽  
H. Pospieszny

Beet necrotic yellow vein virus (BNYVV), the casual agent of rhizomania disease, was identified in sugar beet plants from several fields in the Wielkopolska Region of Poland (1). In greenhouse studies, sugar beets were grown in the soil from one of these fields to bait soilborne viruses. Of 200 sugar beet plants, three developed symptoms of vein clearing, vein banding, and mosaic. Crude sap from symptomatic plants was used for mechanical inoculation of various plants species. In Chenopodium quinoa, C. amaranticolor, and Tetragonia expansa only local lesions were observed. Electron microscope examination of negatively stained leaf-dip preparations from symptomatic sugar beet plants showed a mixture of rod-shape particles from 70 to 400 nm long. Using double-antibody sandwich enzyme-linked immunosorbent assay tests, two symptomatic sugar beet plants gave positive reactions with antiserum against BNYVV (Bio-Rad, Hercules, CA) and a third plant gave a positive reaction with antisera against BNYVV and Beet soilborne virus (BSBV). Total RNA was extracted from roots and leaves of the symptomatic plants and used in a multiplex reverse transcription-polymerase chain reaction (mRT-PCR) assay. Specific primers were designed to amplify a fragment of the RNA1 for BSBV and RNA2 for BNYVV and Beet virus Q (BVQ) (2). Two mRT-PCR products amplified with the primers specific to BNYVV and BSBV were obtained and sequenced. A 274-nt amplicon sequence (GenBank Accession No. DQ012156) had 98% nucleotide sequence identity with the German BNYVV isolate F75 (GenBank Accession No. AF19754) and a 376-nt amplicon sequence (GenBank Accession No. AY999690) had 98% nucleotide and 98% amino acid sequence identity with the German BSBV isolate (GenBank Accession No. Z97873). The Polish BSBV isolate had 88% nucleotide and 62% amino acid sequence identity with BVQ, another pomovirus (GenBank Accession No. AJ 223596 formerly known as serotype Wierthe of BSBV (2). In 2005, mRT-PCR was used on samples collected from two fields of the Wielkopolska Region. Of 15 tested sugar beet plants, 12 gave positive reactions with primers specific for BSBV and nine with primers specific to BNYVV. To our knowledge, this is first report of BSBV in Poland. In Europe, BSBV was previously reported in England, the Netherlands, Belgium, Sweden, Germany, France, and Finland (2,3). References: (1) M. Jezewska and J. Piszczek. Phytopathol. Polonica, 21:165, 2001. (2) A. Maunier et al. Appl. Environ. Microbiol. 69:2356, 2003. (3) C. M. Rush and G. B. Heidel. Plant Dis. 79:868, 1995.


Plant Disease ◽  
2001 ◽  
Vol 85 (12) ◽  
pp. 1286-1286 ◽  
Author(s):  
A. R. Golnaraghi ◽  
N. Shahraeen ◽  
R. Pourrahim ◽  
Sh. Ghorbani ◽  
Sh. Farzadfar

During the summer of 2000, severe stunting, mosaic, bud necrosis, and chlorosis symptoms were observed on peanut (Arachis hypogaea cv. Gilan) plants growing in fields in the Golestan Province of Iran. Leaf extracts of peanut plants were infective (mechanical inoculation) causing necrotic local lesions on Chenopodium quinoa, C. amaranticolor, Gomphrena globosa, Phaseolus vulgaris cv. Talash, Vicia faba, and Vigna unguiculata cv. Mashad; systemic chlorotic spots were followed by systemic necrosis in Datura stramonium, D. metel, and Nicotiana rustica; chlorotic and necrotic spots were followed by top necrosis in Glycine max. About 2 weeks after inoculation, the chlorosis followed by stunting and bud necrosis observed in the field were reproduced in A. hypogaea cv. Gilan. Tomato spotted wilt virus (TSWV) was detected in the original peanut plants and in plant species that developed symptoms after inoculation with extracts from peanut plants, when analyzed by double-antibody sandwich enzyme-linked immunosorbent assay using TSWV-specific antisera (polyclonal antibody As-0526 and As-0580, DSMZ, Braunschweig, Germany). TSWV is one of the most important viruses in the world (2) and has been reported on potato (3) and tomato (1) in Iran. To our knowledge, this is the first report of TSWV infection of peanut in Iran. References: (1) K. Bananej et al. Iran. J. Plant Pathol. 34:30, 1998. (2) R. A. Mumford et al. Ann. Appl. Biol. 128:159, 1996. (3) R. Pourrahim et al. Plant Dis. 85:442, 2001.


Plant Disease ◽  
2004 ◽  
Vol 88 (8) ◽  
pp. 909-909 ◽  
Author(s):  
B. Komorowska ◽  
M. Cieślińska

Cherry virus A (CVA), a member of the genus Capillovirus, has been reported in sweet cherry in Germany, Canada, and Great Britain. No data are available on the effects of CVA on fruit quality and yield of infected trees. Little cherry disease (LChD) occurs in most cherry growing areas of the world. Symptoms on sensitive cultivars include discolored fruit that remain small, pointed in shape, and tasteless. Three Closterovirus spp. associated with LChD have been described (Little cherry virus-1 [LChV-1], LChV-2, and LChV-3). Diseased local and commercial cultivars of sour cherry trees were found in a Prunus sp. germplasm collection and orchards in Poland during the 2003 growing season. The foliar symptoms included irregular, chlorotic mottling, distortion, and premature falling of leaves. Some of the diseased trees developed rosette as a result of decreased growth and shortened internodes. Severely infected branches exhibited dieback symptoms. Because the symptoms were suggestive of a possible virus infection, leaf samples were collected from 38 trees and assayed for Prune dwarf virus and Prunus necrotic ringspot virus using double antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA). RNA extracted from leaves was used in a reverse transcription-polymerase chain reaction (RT-PCR) with the One-Step RT-PCR with Platinum Taq (Invitrogen Life Technologies) and primer sets specific for CVA (1), LChV-1 (3), and LChV-2 (3). The RNA samples were also tested using RT-PCR for detection of Cherry mottle leaf virus (CMLV), Cherry necrotic rusty mottle virus (CNRMV), and Cherry green ring mottle virus (CGRMV) with specific primer sets (2). Amplification of a 397-bp coat protein gene product confirmed infection of 15 trees with CVA. A 419-bp fragment corresponding to the coat protein gene of LChV-1 was amplified from cv. Gisela rootstock and local cv. WVIII/1. To confirm RT-PCR results, CVA amplification products from local cv. WX/5 and LChV-1 from cvs. Gisela and WVIII/1 were cloned in bacterial vector pCR 2.1-TOPO and then sequenced. The sequences were analyzed with the Lasergene (DNASTAR, Madison, WI) computer program. The alignment indicated that the nucleotide sequence of cv. WX/5 was closely related to the published sequences of CVA (Genbank Accession No. NC_003689) and had an 89% homology to the corresponding region. The nucleotide sequence similarity between the 419-bp fragment obtained from cvs. Gisela and WVIII/1 was 87% and 91%, respectively, compared with the reference isolate of LChV-1 (Genbank Accession No. NC_001836). The sampled trees tested negative for LChV-2, CGRMV, CMLV, and CNRMV using RT-PCR. Some trees tested positive for PNRSV and PDV. To our knowledge, this is the first report of CVA and LChV-1 in Poland. References: (1) D. James and W. Jelkmann. Acta Hortic. 472:299, 1998. (2) M. E. Rott and W. Jelkmann. Eur. J. Plant Pathol. 107:411,2001. (3) M. E. Rott and W. Jelkmann. Phytopathology. 91:61, 2001.


Plant Disease ◽  
2005 ◽  
Vol 89 (7) ◽  
pp. 775-775 ◽  
Author(s):  
N. Shahraeen ◽  
T. Ghotbi ◽  
M. Salati ◽  
A. Sahandi

Soybean (Glycine max (L.) Merr.) has been increasing in importance and acreage for the past 5 years in Iran and is now planted on approximately 108,000 ha. Previous surveys in Iran of viruses infecting soybean failed to identify Bean pod mottle virus (BPMV), but the incidence of other common viruses of soybean in the field has been reported (1). During October 2004, symptoms characteristic of those caused by BPMV including mosaic, puckering of trifoliate leaves, and delayed maturity of stems or green stems were observed in soybean fields in the Takhti Mahaleh, Versen, and Hashemabad areas located in the Gorgan Province. Sporadic incidence of plants infected with BPMV has been usually of minor importance to growers. Symptoms were often overlooked or considered to be physiological disorders. A visual assessment was made to determine incidence of green stem in the commonly grown soybean cv. Sahar. Forty soybean plants showing symptoms of crinkling, mottling, green stem, and retaining green leaves were sampled by collecting one trifoliate leaf near the top of the plant. All samples were tested in parallel for BPMV using double-antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA). BPMV was detected in 40% of the samples. Seven of the samples shown to be infected with BPMV using DAS-ELISA were mechanically (2) transferred to soybean seedlings in the greenhouse. These plants developed systemic mottle symptoms typical of those caused by BPMV and tested positive for BPMV using DAS-ELISA. The distribution of BPMV within soybean-growing regions, exploration of potential virus reservoirs, and economic impact of this virus have yet to be determined. There is no published report on the presence of potential BPMV vectors including the bean leaf beetle (Cerotoma trifurcata) from soybean fields in Iran. To our knowledge, this is the first report of BPMV in Iran. References: (1) A. R. Golnaraghi et al. Plant Dis.88:1069, 2004. (2) R. Louie et al. Plant Dis.84:1133, 2000.


Plant Disease ◽  
2001 ◽  
Vol 85 (11) ◽  
pp. 1210-1210 ◽  
Author(s):  
J. Aramburu

During spring 2001, plants of different tomato (Lycopersicon esculentum) cultivars grown in several commercial fields in the eastern Catalonia Region of Spain had fruit with brown patches and young leaves with rings and a bright necrotic mosaic that progressed to stem necrosis of the apex, which might die and later develop new symptomless shoots. The symptoms were similar to those of Cucumber mosaic virus (CMV) and Tomato spotted wilt virus (TSWV). Sap of tomato sample R1 (in buffered saline [0.02 M sodium phosphate, 0.15 M NaCl at pH 7.2, containing 0.2% 2-mercaptoethanol]) was infective to Cucumis sativus (local necrosis), tomato cv. Marmande (systemic infection consisting of chlorotic local lesions and necrotic mosaic), Nicotiana clevelandii and N. benthamiana (chlorosis and rosetting), and Chenopodium quinoa (chlorotic local lesions, systemic mottle, and leaf distortion). The sap was not infective to N. glutinosa, N. tabacum cv. Xanthi, Datura stramonium, or Gomphrena globosa. The host range data indicated that the infective agent in sample R1 could be Parietaria mottle virus (PMoV) (1). Symptomatic plants inoculated in a greenhouse with the R1 isolate and symptomatic from tomato plants from the field were analyzed by indirect enzyme-linked immunosorbent assay (ELISA) and had minimum ELISA values at least 10-fold higher than healthy controls, using a polyclonal antiserum (provided by P. Roggero) of a tomato strain of PMoV denoted tomato virus 1 (2). The R1 isolate of PMoV was negative in ELISA when analyzed with commercial antisera to TSWV, CMV, Tomato mosaic virus, Tomato bushy stunt virus, Potato Y virus, Tobacco etch virus, Pelargonium zonate spot virus, and Tobacco streak virus. References: (1) P. Caciagli et al. Plant Pathol. 38:577, 1989. (2) P. Roggero et al. J. Plant Pathol. 82:159, 2000.


Plant Disease ◽  
1997 ◽  
Vol 81 (8) ◽  
pp. 959-959 ◽  
Author(s):  
A. S. Kline ◽  
E. J. Anderson

Cowpea aphid-borne mosaic potyvirus (CABMV) is one of several seed-borne viruses known to limit cowpea (Vigna unguiculata (L.) Walp. subsp. unguiculata) production in Africa, Europe, and Asia, but CABMV has not been reported on commercially grown cowpeas in the United States (1). However, a sesame (Sesamum indicum L.)-infecting isolate of CABMV was recently characterized from plants growing near cowpea introduction plots in Georgia (2). In February 1997, we received samples of three seed lots of cowpea cv. Chinese Red that had been harvested in southern Texas during 1996. Approximately 28% of the plants grown from these seed lots expressed strong mosaic symptoms on primary and trifoliate leaves. Viruslike symptoms were reproduced following mechanical transmission to plants of Chinese Red cowpea, Nicotiana benthamiana, and soybean (Glycine max L.) cv. Lee. When Coronet and Pinkeye Purple Hull-BVR cowpeas were inoculated with sap extracts from symptomatic Chinese Red plants, chlorotic lesions developed on inoculated leaves, but only Coronet plants supported symptomless systemic infections. Similarly inoculated plants of Chenopodium quinoa (L.) and common bean (Phaseolus vulgaris L.) cvs. Pinto and Black Valentine developed localized chlorotic lesions. In Ouchterlony gel diffusion assays, extracts from symptomatic cowpea plants did not react with antisera to blackeye cowpea mosaic potyvirus (BlCMV), cucumber mosaic cucu-movirus (CMV), southern bean mosaic sobemovirus, cowpea mosaic comovirus, cowpea severe mosaic comovirus, or cowpea chlorotic mottle bromovirus. In the indirect enzyme-linked immunosorbent assay, sap extracts from symptomatic plants reacted with antiserum to CABMV, giving OD values at A405 of 0.10 to 0.25, and reacted weakly with antiserum to BlCMV, with OD values at A405 less than 0.035. Extracts from healthy control plants gave OD values at A405 less than 0.010. No positive reactions were obtained with antisera to bean yellow mosaic potyvirus, peanut mottle potyvirus, soybean mosaic potyvirus, or CMV. To our knowledge, this is the first report of CABMV in commercially grown cowpea from the U.S. References: (1) A. G. Gillaspie et al. Plant Dis. 79:388, 1995. (2) H. R. Pappu et al. Arch. Virol. 142:1, 1997.


Plant Disease ◽  
2006 ◽  
Vol 90 (6) ◽  
pp. 828-828 ◽  
Author(s):  
J. J. Weiland ◽  
R. L. Larson ◽  
T. P. Freeman ◽  
M. C. Edwards

In October of 2005, sugar beet (Beta vulgaris L.) plants exhibiting symptoms of rhizomania caused by Beet necrotic yellow vein virus (BNYVV) (3) were observed in a production field near Greeley, CO. The roots of seven plants exhibiting moderate to severe symptoms characteristic of this disease were tested using double-antibody sandwich enzyme-linked immunosorbent assay with anti-BNYVV antiserum from rabbits. Of these, only two roots exhibiting the mildest symptoms tested positive for BNYVV (all roots tested negative for the presence of the related Beet soilborne mosaic virus (BSBMV). ‘Hairy’ lateral roots characteristic of the disease were combined from the remaining five roots, ground in phosphate buffer, and the supernatant from the suspension was mechanically applied to leaves of Chenopodium quinoa in an effort to isolate an infectious agent. Five days postinoculation (dpi), yellow lesions with necrotic centers were visible on inoculated leaves, well in advance of those typically observed for BNYVV or BSBMV. Lesions exhibiting a similar rate of development on C. quinoa subsequently were induced from extracts of root vascular tissue prepared from four of seven additional beet roots tested from this location. Transfer of the infection from the C. quinoa lesions to 32 healthy C. quinoa and 10 sugar beet plants (hybrid ACH9369; American Crystal Sugar Co., Moorhead MN) resulted in 100% infection. Inoculated leaves of C. quinoa exhibited a high density of necrotic local lesions within 3 dpi, whereas inoculated leaves of sugar beet exhibited pinpoint, necrotic to diffuse, chlorotic local lesions evident by 5 dpi. Electron microscopic examination of fixed, ultra-thin sections of symptomatic C. quinoa leaf tissue revealed aggregates of virus-like particles of icosahedral symmetry within the cell cytoplasm. Following a virus minipreparation procedure, nucleic acid extracted from the partially purified virus was found to be single-stranded RNA by ribonuclease digestion and alone was infectious when inoculated to C. quinoa leaves. The apparently monopartite RNA genome was 3.5 kb long and a candidate for the single coat protein (CP) had a mass of ˜25 kDa. The sole reference set found in the literature for a virus naturally occurring on sugar beet with similar characteristics was that for Beet black scorch virus (BBSV), a virus recently accepted by the ICTV into the genus Necrovirus within the family Tombusviridae (2). Prior to this communication, BBSV has only been reported in China where it was first documented affecting sugar beet in the late 1980s (1). Using the published sequence of BBSV (Genbank Accession No. AY626780), DNA primers directed to the 3′ half of the BBSV genome were used in reverse transcription-polymerase chain reaction to produce an amplicon from the unknown virus. Sequencing the amplicon revealed 88.8% nucleotide sequence identity to the BBSV CP gene and 97% amino acid sequence identity to the predicted CP gene product. Combined, the nucleotide sequence and physical characteristics confirm the presence of BBSV in a U.S. sugarbeet field for the first time. To our knowledge, this is the first report of the occurrence of BBSV outside of China. References: (1) Y. Cao et al. Arch. Virol. 147:2431, 2002. (2) C. M. Fauquet et al. Eighth Report of the International Committee on the Taxonomy of Viruses. Academic Press, New York, 2005. (3) C. M. Rush. Ann. Rev. Phytopathol. 41:567, 2003.


Plant Disease ◽  
2010 ◽  
Vol 94 (1) ◽  
pp. 125-125 ◽  
Author(s):  
S. T. Koike ◽  
T. Tian ◽  
H.-Y. Liu

In 2009 in coastal California (Santa Barbara County), commercially grown spinach (Spinacia oleracea) in two nearby fields exhibited symptoms of a previously unrecognized virus-like disease. Symptoms consisted of general chlorosis and bright yellow blotches and spots. Necrotic spots were also associated with the disease. In affected fields, disease occurred in limited, irregularly shaped patches that ranged from one to several meters in diameter. Symptomatic plants were unmarketable and these small patches of spinach were not harvested. With a transmission electron microscope, rigid, rod-shaped particles with a clear central canal were observed from plant sap of the symptomatic spinach. Analysis by a double-antibody sandwich-ELISA assay (Agdia Inc., Elkhart, IN) for Tobacco rattle virus (TRV) showed that the symptomatic plants were positive. Symptomatic spinach from the field was used for mechanical transmission to Chenopodium quinoa, C. murale, C. capitatum, spinach, and sugar beet (Beta vulgaris). All inoculated plants showed chlorotic local lesions and sugar beet showed chlorotic local lesions with rings. To further confirm the presence of TRV, reverse transcription (RT)-PCR was conducted. Total RNA was extracted from the mechanically inoculated symptomatic spinach plants using an RNeasy Plant Kit (Qiagen Inc., Valencia, CA) and used as a template in RT-PCR with forward (5′-TACATCACATCTGCCTGC-3′) and reverse (5′-CTTCATTCACACAACCCTTG-3′) primers specific to the movement protein gene from the spinach isolate of TRV (GenBank Accession No. AJ007294). Amplicons of the expected size (approximately 562 bp) were obtained. The RT-PCR products were sequenced (GenBank Accession No. GU002156) and compared with TRV sequences in GenBank to confirm the identity of the products. Sequences obtained had 96% nucleotide identity and 97% amino acid identity with TRV sequences available under the GenBank Accession Nos. FJ357571 and AJ007294. On the basis of the data from electron microscopy and serological and molecular analyses, the virus was identified as TRV. Soil samples collected from one of the fields were assayed for nematodes; however, Paratrichodorus or Trichodorus species were not recovered. To our knowledge, this is the first report of TRV in spinach in California. TRV has also been reported in spinach in England (1) and Germany (2). References: (1) A. Kurppa et al. Ann. Appl. Biol. 98:243, 1981. (2) K. Schmidt and R. Koenig. Arch. Virol. 144:503, 1999.


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