scholarly journals Identification of Soybean mosaic virus Strains by RT-PCR/RFLP Analysis of Cylindrical Inclusion Coding Region

Plant Disease ◽  
2004 ◽  
Vol 88 (6) ◽  
pp. 641-644 ◽  
Author(s):  
Yul-Ho Kim ◽  
Ok-Sun Kim ◽  
Jae-Hwan Roh ◽  
Jung-Kyung Moon ◽  
Soo-In Sohn ◽  
...  
Keyword(s):  
Mosaic Virus ◽  
Soybean Mosaic Virus ◽  
Rflp Analysis ◽  
Cylindrical Inclusion ◽  
Enzyme Linked Immunosorbent Assay ◽  
Strain Identification ◽  
Coding Region ◽  
Rt Pcr ◽  
Detection And Identification ◽  
Pcr Rflp

A reverse-transcriptase polymerase chain reaction/restriction fragment length polymorphism (RT-PCR/RFLP) was employed successfully for detection and identification of Soybean mosaic virus (SMV) strains. A primer pair amplifying a 1,385-bp fragment of the cylindrical inclusion (CI) coding region was used for RT-PCR and the RFLP profiles of the RT-PCR products were compared after restriction digestion with RsaI, EcoRI, or AccI restriction endonucleases. These enzymes were chosen based on the nucleotide sequences of SMV strains G2, G5, G5H, G7, and G7H in the CI coding region. These five strains, as well as seedborne SMV isolates from local soybean cultivars, could be differentiated by RT-PCR/RFLP analysis. The results correlated well with strain identification by symptom phenotypes produced on differential cultivars inoculated with strains and isolates. The sensitivity of RT-PCR enabled detection of SMV from plants with necrotic symptoms in which the number of virus particles was too low to be detected by enzyme-linked immunosorbent assay.

RT-PCR-RFLP analysis for evaluating cross protection by an attenuated isolate of Cucumber mosaic virus

2005 ◽  
Vol 71 (3) ◽  
pp. 243-246 ◽  
Author(s):  
Eiko Nakazono-Nagaoka ◽  
Masako Suzuki ◽  
Yoshitaka Kosaka ◽  
Tomohide Natsuaki
Keyword(s):  
Cucumber Mosaic Virus ◽  
Mosaic Virus ◽  
Rflp Analysis ◽  
Cross Protection ◽  
Rt Pcr ◽  
Pcr Rflp ◽  
Attenuated Isolate

scholarly journals Genetic Diversity Among Viruses Associated with Sugarcane Mosaic Disease in Tucumán, Argentina

2009 ◽  
Vol 99 (1) ◽  
pp. 38-49 ◽  
Author(s):  
M. F. Perera ◽  
M. P. Filippone ◽  
C. J. Ramallo ◽  
M. I. Cuenya ◽  
M. L. García ◽  
...  
Keyword(s):  
Genetic Diversity ◽  
Mosaic Virus ◽  
Fragment Length Polymorphism ◽  
Rflp Analysis ◽  
Mosaic Disease ◽  
Rt Pcr ◽  
Rflp Profile ◽  
Pcr Rflp ◽  
Polymerase Chain ◽  
Sugarcane Mosaic Disease

Sugarcane leaves with mosaic symptoms were collected in 2006–07 in Tucumán (Argentina) and analyzed by reverse-transcriptase polymerase chain reaction (RT-PCR) restriction fragment length polymorphism (RFLP) and sequencing of a fragment of the Sugarcane mosaic virus (SCMV) and Sorghum mosaic virus (SrMV) coat protein (CP) genes. SCMV was detected in 96.6% of samples, with 41% showing the RFLP profile consistent with strain E. The remaining samples produced eight different profiles that did not match other known strains. SCMV distribution seemed to be more related to sugarcane genotype than to geographical origin, and sequence analyses of CP genes showed a greater genetic diversity compared with other studies. SrMV was detected in 63.2% of samples and most of these were also infected by SCMV, indicating that, unlike other countries and other Argentinean provinces, where high levels of co-infection are infrequent, co-existence is common in Tucumán. RFLP analysis showed the presence of SrMV strains M (68%) and I (14%), while co-infection between M and H strains was present in 18% of samples. Other SCMV subgroup members and the Sugarcane streak mosaic virus (SCSMV) were not detected. Our results also showed that sequencing is currently the only reliable method to assess SCMV and SrMV genetic diversity, because RT-PCR-RFLP may not be sufficiently discriminating.

scholarly journals Effect of Aphid Behavior on Efficiency of Transmission of Soybean mosaic virus by the Soybean-Colonizing Aphid, Aphis glycines

Plant Disease ◽  
2002 ◽  
Vol 86 (11) ◽  
pp. 1260-1264 ◽  
Author(s):  
R. Y. Wang ◽  
S. A. Ghabrial
Keyword(s):  
Mosaic Virus ◽  
Transmission Rate ◽  
Soybean Mosaic Virus ◽  
Aphis Glycines ◽  
Coding Region ◽  
Rt Pcr ◽  
Soybean Aphids ◽  
Efficient Vector ◽  
Tobacco Vein Mottling Virus ◽  
Soybean Leaves

The soybean aphid (Aphis glycines) was a poor vector (0.83% transmission) when the aphids were allowed overnight acquisition feed on Soybean mosaic virus (SMV)-infected soybean leaves. However, A. glycines was shown to be a very efficient vector (34.72% transmission) when individual aphids were allowed a 1-min acquisition probe on the same infected leaves used for the feeding treatment. Similar results were obtained with Myzus persicae and tobacco in transmission experiments of the potyviruses Tobacco etch virus (feeding: 1.36%; probing: 45.5%) and Tobacco vein mottling virus (feeding: 2.0%; probing: 47.5%). A reverse transcription-polymerase chain reaction (RT-PCR) assay was developed to detect SMV in single soybean aphids using a pair of primers designed to amplify a 469-bp PCR fragment in the coding region of SMV coat protein. In contrast to the low transmission rate obtained with the soybean aphids that acquired virus through overnight feeding, RT-PCR detected SMV in 100% of these aphids. Interestingly, the rate of SMV detection by RT-PCR in aphids that were allowed a 1-min acquisition probe (31.67%) coincided with percent transmission (34.72%). The practical application of RT-PCR in detecting nonpersistently transmitted viruses and its implications for virus epidemiology are discussed.

scholarly journals Aphis glycines as a Vector of Persistently and Nonpersistently Transmitted Viruses and Potential Risks for Soybean and Other Crops

Plant Disease ◽  
2006 ◽  
Vol 90 (7) ◽  
pp. 920-926 ◽  
Author(s):  
R. Y. Wang ◽  
A. Kritzman ◽  
D. E. Hershman ◽  
S. A. Ghabrial
Keyword(s):  
Mosaic Virus ◽  
Soybean Mosaic Virus ◽  
The United States ◽  
Plant Viruses ◽  
Soybean Aphid ◽  
Aphis Glycines ◽  
Yellow Mosaic Virus ◽  
Coding Region ◽  
Rt Pcr ◽  
First Report

The recently introduced soybean aphid (Aphis glycines), which is widespread in the soybean-growing regions in the United States, is the only aphid able to develop large colonies on soybean. Although its potential as a vector of plant viruses is recognized, reports on virus transmission efficiency by this aphid species are limited. In the present study, we examined the ability of A. glycines to transmit several economically important viruses. The results showed that A. glycines transmitted the potyviruses Bean yellow mosaic virus (BYMV) and Soybean mosaic virus from soybean to soybean more efficiently than Myzus persicae. However, M. persicae transmitted the alfamovirus Alfalfa mosaic virus and the potyviruses Tobacco etch virus (TEV) and Tobacco vein mottling virus (TVMV) from tobacco to tobacco more efficiently than A. glycines. This is the first report to demonstrate that the soybean aphid can vector TEV and TVMV, two economically important tobacco viruses. This also is the first report to document successful transmission of BYMV by A. glycines. All attempts to transmit the nepovirus Tobacco ringspot virus by A. glycines were unsuccessful, regardless of the length of the acquisition and inoculation feeding periods. Although the luteovirus Soybean dwarf virus (SbDV) was widely distributed in red and white clover in Kentucky, it was not detected in soybean. All transmission experiments of SbDV by A. glycines were unsuccessful. A reverse-transcription polymerase chain reaction (RT-PCR) assay was developed to detect SbDV in single aphids using a pair of primers designed to amplify a 372-bp PCR fragment in the coding region of SbDV coat protein. Although A. glycines was not a vector of SbDV, the virus was detected in 100% of tested aphids by RT-PCR after a 24- to 48-h virus acquisition access feeding. The practical applications of RT-PCR in detecting persistently transmitted viruses are discussed.

scholarly journals Identification, Characterization, and Molecular Detection of Alfalfa mosaic virus in Potato

2006 ◽  
Vol 96 (11) ◽  
pp. 1237-1242 ◽  
Author(s):  
H. Xu ◽  
J. Nie
Keyword(s):  
Mosaic Virus ◽  
Gene Sequence ◽  
Alfalfa Mosaic Virus ◽  
Enzyme Linked Immunosorbent Assay ◽  
Polymorphism Analysis ◽  
Rt Pcr ◽  
Potato Leaf ◽  
Simple Method ◽  
Cp Gene ◽  
Nucleotide Sequence Alignment

Alfalfa mosaic virus (AMV) was detected in potato fields in several provinces in Canada and characterized by bioassay, enzyme-linked immunosorbent assay, and reverse-transcription polymerase chain reaction (RT-PCR). The identity of eight Canadian potato AMV isolates was confirmed by sequence analysis of their coat protein (CP) gene. Sequence and phylogenetic analysis indicated that these eight AMV potato isolates fell into one strain group, whereas a slight difference between Ca175 and the other Canadian AMV isolates was revealed. The Canadian AMV isolates, except Ca175, clustered together among other strains based on alignment of the CP gene sequence. To detect the virus, a pair of primers, AMV-F and AMV-R, specific to the AMV CP gene, was designed based on the nucleotide sequence alignment of known AMV strains. Evaluations showed that RT-PCR using this primer set was specific and sensitive for detecting AMV in potato leaf and tuber samples. AMV RNAs were easily detected in composite samples of 400 to 800 potato leaves or 200 to 400 tubers. Restriction analysis of PCR amplicons with SacI was a simple method for the confirmation of PCR tests. Thus, RT-PCR followed by restriction fragment length polymorphism analysis may be a useful approach for screening potato samples on a large scale for the presence of AMV.

scholarly journals Role of Soybean mosaic virus–Encoded Proteins in Seed and Aphid Transmission in Soybean

2013 ◽  
Vol 103 (9) ◽  
pp. 941-948 ◽  
Author(s):  
Sushma Jossey ◽  
Houston A. Hobbs ◽  
Leslie L. Domier
Keyword(s):  
Mosaic Virus ◽  
Seed Quality ◽  
Soybean Mosaic Virus ◽  
Seed Transmission ◽  
Aphid Transmission ◽  
Plant Introduction ◽  
Sequence Motif ◽  
Coding Region ◽  
Encoded Proteins ◽  
Seed Transmissibility

Soybean mosaic virus (SMV) is seed and aphid transmitted and can cause significant reductions in yield and seed quality in soybean (Glycine max). The roles in seed and aphid transmission of selected SMV-encoded proteins were investigated by constructing mutants in and chimeric recombinants between SMV 413 (efficiently aphid and seed transmitted) and an isolate of SMV G2 (not aphid or seed transmitted). As previously reported, the DAG amino acid sequence motif near the amino terminus of the coat protein (CP) was the major determinant in differences in aphid transmissibility of the two SMV isolates, and helper component proteinase (HC-Pro) played a secondary role. Seed transmission of SMV was influenced by P1, HC-Pro, and CP. Replacement of the P1 coding region of SMV 413 with that of SMV G2 significantly enhanced seed transmissibility of SMV 413. Substitution in SMV 413 of the two amino acids that varied in the CPs of the two isolates with those from SMV G2, G to D in the DAG motif and Q to P near the carboxyl terminus, significantly reduced seed transmission. The Q-to-P substitution in SMV 413 also abolished virus-induced seed-coat mottling in plant introduction 68671. This is the first report associating P1, CP, and the DAG motif with seed transmission of a potyvirus and suggests that HC-Pro interactions with CP are important for multiple functions in the virus infection cycle.

scholarly journals Isolado do vírus do mosaico do pepino obtido de bananeira no Estado de São Paulo pertence ao subgrupo Ia

2001 ◽  
Vol 26 (1) ◽  
pp. 53-59 ◽  
Author(s):  
MARCELO EIRAS ◽  
ADDOLORATA COLARICCIO ◽  
ALEXANDRE L.R. CHAVES
Keyword(s):  
Mosaic Virus ◽  
Vigna Unguiculata ◽  
Sao Paulo ◽  
São Paulo ◽  
Rt Pcr ◽  
Nicotiana Glutinosa ◽  
Musa Sp ◽  
De Se ◽  
Pcr Rflp ◽  
Das Elisa

Em 1996, foi feita a caracterização parcial de um isolado do vírus do mosaico do pepino (Cucumis mosaic virus, CMV) obtido de bananeira (Musa sp.) proveniente do município de Miracatu, SP. Com o objetivo de se determinar o subgrupo do isolado de CMV, recorreu-se às técnicas de ELISA, RT-PCR, RFLP e seqüenciamento de fragmentos de RNA genômico. Amostras de folhas infetadas, desidratadas com cloreto de cálcio e armazenadas à -20 °C desde 1994 na viroteca do Laboratório de Fitovirologia e Fisiopatologia, foram inoculadas em plantas de Nicotiana glutinosa. Dez dias após a inoculação, folhas apresentando mosaico foram utilizadas para DAS-ELISA e extração de RNAs totais. Em ELISA, houve reação apenas contra o anti-soro específico para CMV subgrupo I. Através de RT-PCR com primers desenhados para anelar em regiões conservadas da porção terminal 3' do gene da capa protéica, foi amplificado um fragmento de DNA com 486 pares de bases. O produto obtido via RT-PCR foi submetido à digestão com as enzimas EcoRI, HindIII, BamHI e MspI, obtendo-se um padrão de restrição esperado para o subgrupo I. Estes resultados foram confirmados através do seqüenciamento do produto de PCR, o qual apresentou homologia de 96% a 98% com os isolados do CMV pertencentes ao subgrupo I. Pelos sintomas observados na hospedeira diferencial Vigna unguiculata, o isolado foi confirmado como sendo do subgrupo Ia.

scholarly journals Allamanda Mosaic Caused by Cucumber mosaic virus in Taiwan

Plant Disease ◽  
2005 ◽  
Vol 89 (5) ◽  
pp. 529-529 ◽  
Author(s):  
Y. K. Chen ◽  
C. C. Yang ◽  
H. T. Hsu
Keyword(s):  
Cucumber Mosaic Virus ◽  
Mosaic Virus ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Chenopodium Quinoa ◽  
Rabbit Antiserum ◽  
Nucleotide Sequence Analysis ◽  
Enzyme Linked Immunosorbent Assay ◽  
Rt Pcr ◽  
Indicator Plants

Allamanda (Allamanda cathartica L., family Apocynaceae) is native to Brazil and is a popular perennial shrub or vine ornamental in Taiwan. Plants showing severe mosaic, rugosity, and leaf distortion symptoms on leaves are common in commercial nurseries and private gardens. Examination of crude sap prepared from symptomatic leaves using an electron microscope revealed the presence of spherical virus particles with a diameter of approximately 28 nm. The virus was mechanically transmitted to indicator plants and induced symptoms similar to those incited by Cucumber mosaic virus (CMV). The virus caused local lesions on inoculated leaves of Chenopodium quinoa and C. amaranticolor and systemic mosaic in Cucumis sativus, Lycopersicon esculentum, Nicotiana benthamiana, N. glutinosa, N. rustica, and N. tabacum. On N. tabacum, necrotic ringspots developed on inoculated leaves followed by systemic mosaic. Tests of leaf sap extracted from naturally infected allamanda and inoculated indicator plants using enzyme-linked immunosorbent assay were positive to rabbit antiserum prepared to CMV. Viral coat protein on transblots of sodium dodecyl sulfate-polyacrylamide gel electrophoresis reacted with CMV subgroup I specific monoclonal antibodies (2). With primers specific to the 3′-half of RNA 3 (1), amplicons of an expected size (1,115 bp) were obtained in reverse transcription-polymerase chain reaction (RT-PCR) using total RNA extracted from infected allamanda and N. benthamiana. The amplified fragment (EMBL Accession No. AJ871492) was cloned and sequenced. It encompasses the 3′ part of the intergenic region of RNA 3 (158 nt), CP ORF (657 nt), and 3′ NTR (300 nt) showing 91.8–98.9% and 71.4–72.8% identities to those of CMV in subgroups I and II, respectively. Results of MspI-digested restriction fragment length polymorphism patterns of the RT-PCR fragment and the nucleotide sequence analysis indicate that the CMV isolate from allamanda belongs to subgroup IB, which is predominant on the island. To our knowledge, CMV is the only reported virus that infects allamanda and was first detected in Brazil (3), and this is the first report of CMV infection in allamanda plants occurring in Taiwan. References: (1) Y. K. Chen et al. Arch. Virol. 146:1631, 2001. (2) H. T. Hsu et al. Phytopathology 90:615, 2000. (3) E. W. Kitajima. Acta. Hortic. 234:451, 1988.

scholarly journals Alternanthera mosaic virus Found in Scutellaria, Crossandra, and Portulaca spp. in Florida

Plant Disease ◽  
2006 ◽  
Vol 90 (6) ◽  
pp. 833-833 ◽  
Author(s):  
C. A. Baker ◽  
L. Breman ◽  
L. Jones
Keyword(s):  
Electron Microscopy ◽  
United States ◽  
Mosaic Virus ◽  
Plant Species ◽  
The United States ◽  
Enzyme Linked Immunosorbent Assay ◽  
Rt Pcr ◽  
Indicator Plants ◽  
Partial Identity ◽  
Pcr Products

In the fall of 1998, the Division of Plant Industry (DPI) received vegetative propagations of Scutellaria longifolia (skullcap) with symptoms of foliar mosaic, chlorotic/necrotic ringspots, and wavy line patterns from a nursery in Manatee County. Flexuous particles approximately 500 nm long were found with electron microscopy. The plants tested positive for Papaya mosaic virus (PaMV) in an enzyme-linked immunosorbent assay (ELISA) test with antiserum to PaMV (Agdia, Elkhart, IN). However, in immunodiffusion tests (antiserum from D. Purcifull, University of Florida), this virus gave a reaction of partial identity indicating it was related but not identical to PaMV (1). The original infected plants were kept in a greenhouse. In January 2005, a specimen of Crossandra infundibuliformis (firecracker plant) with mosaic symptoms was submitted to the DPI from a nursery in Alachua County. Inclusions found with light microscopy and particles found with electron microscopy indicated that this plant was infected with a potexvirus. This was confirmed by reverse transcription-polymerase chain reaction (RT-PCR) with primers designed to detect members of the virus family Potexviridae (3). These plants reacted positive to PaMV antiserum in ELISA and gave a reaction of partial identity to PaMV in immunodiffusion. A specimen of Portulaca grandiflora (moss rose) with distorted leaves found at a local retail store was also tested and gave the same results. Leaves from each of the three plant species were rubbed onto a set of indicator plants using Carborundum and potassium phosphate buffer. Total RNA was extracted from symptomatic indicator plants of Nicotiana benthamiana. RT-PCR (3) was performed, and PCR products were sequenced directly. Sequences of approximately 700 bp were obtained for all three plant species and showed 98% identity with each other. BLAST search results showed that these sequences were 93% identical to an Alternanthera mosaic virus (AltMV) sequence at the nucleotide level but only 76% identical to PaMV. The amino acid sequences were 98 and 82% identical to AltMV and PaMV, respectively. The PCR products of the virus from Scutellaria sp. were cloned, resequenced, and the sequence was entered into the GenBank (Accession No. DQ393785). The bioassay results matched those found for AltMV in Australia (2) and the northeastern United States (4), except that the Florida viruses infected Datura stramonium and Digitalis purpurea (foxglove). The virus associated with the symptoms of these three plants appears to be AltMV and not PaMV. AltMV has been found in ornamental plants in Australia, Italy, and the United States (Pennsylvania, Maryland, and now Florida). Since this virus is known to infect several plants asymptomatically and can be easily confused with PaMV serologically, it is likely that the distribution of this virus is much wider than is known at this time. References: (1) L. L. Breman. Plant Pathology Circular No. 396. Fla. Dept. Agric. Consum. Serv. DPI, 1999. (2) A. D. W. Geering and J. E. Thomas. Arch Virol 144:577, 1999. (3) A. Gibbs et al. J Virol Methods 74:67, 1998. (4) J. Hammond et al. Arch Virol. 151:477, 2006.

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