Differentiation of cucumber mosaic virus isolates using the polymerase chain reaction

Rizos, Helen; Gunn, L. V.; Pares, R. D.; Gillings, Michael R.

doi:10.1099/0022-1317-73-8-2099

Cited by 75 publications

(52 citation statements)

References 12 publications

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“…Serological differentiation, PCR followed by restriction analysis (Rizos et al 1992), and IC-RT-PCR (Yu et al 2005) methods have been used for subgrouping of CMV isolates. The latter method is more applicable because of avoiding the extraction of viral or plant total RNA and can be easily performed in a single tube (Yu et al 2005).…”

Section: Discussionmentioning

confidence: 99%

“…The virus is mechanically transmissible by plant sap and spread by more than 80 aphid species in the non-persistent manner. The genome of CMV consists of three singlestranded positive-sense RNA species (Palukaitis & Garcia-Arenal 2003) The CMV strain variation has been studied by serological assay (Wilson & Halliwell 1985), dsRNA analysis , PCR-RFLP (Rizos et al 1992), and microarray methods (Deyong et al 2005); thereby, it has been separated into two main subgroups, subgroup I (S-I) and subgroup II (S-II), on the basis of serological relationships, peptide mapping of the coat protein (CP), nucleic acid hybridisation, and nucleotide sequence identity (Palukaitis et al 1992). Another division of subgroup I into IA and IB has been proposed based on sequence data, analysis of 5'-non-translated region of RNA3 of several strains, and phylogenetic analysis of CP (Roossinck et al 1999;Roossinck 2002).…”

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confidence: 99%

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Bioassay and phylogeny of five Iranian isolates of Cucumber mosaic virus from different hosts based on CP gene sequence

Eyvazi¹,

Dizadji²,

Rastgou³

et al. 2015

Plant Prot. Sci.

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Using double-antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA), Cucumber mosaic virus (CMV) was detected in 31 out of 132 symptomatic leaf samples collected from different hosts of Urmia province of Iran, during 2011-2012. In biological assays, five different host isolates caused severe mosaic in Nicotiana species and Capsicum annum without significant difference in severity. Based on phylogenetic analysis of coat protein nucleotide and deduced amino acid sequence, two isolates were clustered into subgroup IA, while other three isolates were grouped into IB subgroup of CMV. All Urmian isolates shared a common MspI, and no EcoRI and BsuRI restriction sites. In contrast to S-IA isolates, the second MspI site was found at 473-476 position of only S-IB isolates, which could be used to differentiate two S-IA and S-IB subgroups. Here, we report the first case of Abutilon theophrasti infection, as a new reservoir weed host for CMV in the world.

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Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

Bioassay and phylogeny of five Iranian isolates of Cucumber mosaic virus from different hosts based on CP gene sequence

Eyvazi¹,

Dizadji²,

Rastgou³

et al. 2015

Plant Prot. Sci.

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“…PCR was done using specific primer pair CMV (CPf/CPr) [5] and a *867 bp expected fragment was amplified from one sample (Pe1). Attempts to detect the ZYMV using a primer pair ZYMV-F(?…”

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confidence: 99%

Capsicum annum, a new host of watermelon mosaic virus

Hajizadeh

Mohammadi

2016

VirusDis.

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The occurrence of Watermelon mosaic virus (WMV) in sweet pepper (Capsicum annuum L.) in Kurdistan province, Iran was confirmed by reverse transcriptase-polymerase chain reaction (RT-PCR) and partial characterization of coat protein. To the best of our knowledge, this is the first report of WMV infecting C. annuum, adding a new host to list of more than 170 species infected by this virus.Sweet pepper (Capsicum annuum L.) is an edible plant belongs to the family Solanaceae that is a valuable vegetable in Iran. Aphid transmitted viruses, Pepper mottle virus (PeMV), Cucumber mosaic virus (CMV), Zucchini yellow mosaic virus (ZYMV) and Potato virus Y (PVY) are known to affect sweet pepper. However, watermelon mosaic virus (WMV), a aphid transmitted virus that occurs worldwide is not known to infect sweet pepper. WMV has a wide host range and is able to infect cucurbits, some legumes, orchids and many weeds [2]. In the summer 2014, peppers with virus-like symptoms including mosaic, leafdistortion, mottling, stunting and yellowing were observed in the Kurdistan province of Iran. Twelve symptomatic pepper samples were collected and tested for the presence of three viruses including CMV, WMV and ZYMV.Total RNA was extracted from symptomatic leaves as described by [4] and subjected to random primed cDNA synthesis using the HyperScript TM kit (GeneAll, Seoul, Korea) according to the manufacturer's protocol. PCR was done using specific primer pair CMV (CPf/CPr) [5] and a *867 bp expected fragment was amplified from one sample (Pe1). Attempts to detect the ZYMV using a primer pair ZYMV-F(?)/ZYMV-R(-) [6] was unsuccessful. Whereas, a *657 bp expected amplification for WMV with another primer pair WMV-F(?)/WMV-R(-) [6] was detected from three samples (Pe1, Pe8 and Pe18). No DNA fragment was earned from non-symptomatic plant (Fig. 1b). WMV was detected of samples with leaf-distortion and mosaic (Fig. 1a). Nine samples did not result in amplification of any DNA fragment and probably these plants were infected with the other viruses, which incite similar symptoms. Since the WMV was not previously reported from Capsicum annuum in worldwide, to confirm the presence of WMV, PCR product from one isolate of WMV (Pe18) was cloned using pTG19 Vector (SinaClon, Iran) and sequenced. The initial BLAST between new isolate Pe18 (GenBank accession no. KP751208) and other Iranian isolates showed the range of nucleotide sequence identity was between 93 and 98 % and amino acid identity range was between 97 and 99 %. Nucleotide sequence identity between our isolate with non-Iranian isolates retrieved from GenBank ranged between 92 and 98 %. In addition, amino acid identity range was between 96 and 99 %.Previous studies have shown that WMV isolates are phylogenetically classified into three main genogroups called classical (Cl), group-2 (G-2) and emerging group (Em) with four subclades, Em1, Em2, Em3 and Em4 [2].Electronic supplementary material The online version of this article

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“…Many strains of CMV described from different parts of the world have been put into two subgroups, I and II, based on nucleic acid hybridization (Owen and Palukaitis, 1988), gene sequences (Owen et al, 1990;Szilassy et al, 1999), restriction fragment length polymorphism (RFLP) (Rizos et al, 1992), and serology (Hu et al, 1995). The diverse strains of CMV are divided into two main groups, I and II, according to their genomic sequences.…”

Section: Introductionmentioning

confidence: 99%

Partial characterization and development of sensitive and reliablediagnostic for the detection of cucumber mosaic virus

Khan¹

2015

Turk J Agric For

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A survey on Cucumber mosaic virus (CMV) was conducted on 21 banana orchards in three bananas producing states: Karnataka, Maharashtra, and Uttar Pradesh. Sampling was done during winter and autumn and all samples were initially examined for the presence of CMV using DAS-ELISA. Out of 300 tested samples, the presence of CMV was observed in 13. The lowest rate of infection was determined in Uttar Pradesh followed by Maharashtra, while high infections were observed in samples from Karnataka. Three CMV infected samples represented in each state were amplified through RT-PCR using the movement protein (MP) gene specific primers and cloned in the pGEMT essay vector. The RT-PCR products of amplified samples were sequenced. Gene sequences of CMV-KAR, CMV-MR, and CMV-UP isolates under study coding for the movement protein revealed 99.76%-99.88% and 99.28%-99.64% sequence identity at the nucleotide and amino-acid level. These isolates were compared with 29 CMV isolates reported from various plants around the world, which formed three distinct subclades: -IA, -IB, and -II. These isolates were most closely related to subgroup-I isolates, sharing up to 95.81% and 96.84% sequence identity at nucleotide and amino acid, respectively. However, these isolates shared 81.79%-81.90% and 82.80%-83.15% nucleotide and amino acid identity with subgroup II isolates. The isolates under study clustered with the CMV subgroup-IB as confirmed by phylogenetic analysis. Further, we standardized the minimum limit of RNA transcript for the sensitivity of RT-PCR with respect to the coat protein (CP) gene and movement protein (MP) gene. The results of RT-PCR showed that the CP gene was more sensitive than the MP gene for the detection of CMV as it amplified the gene product up to 0.02 ng RNA. The CP gene based RT-PCR assay may be a more sensitive, reliable, and convenient molecular tool for detection of the CMV pathogen, and can be used in quarantine, eradication, and tissue culture certification programs.

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Differentiation of cucumber mosaic virus isolates using the polymerase chain reaction

Cited by 75 publications

References 12 publications

Bioassay and phylogeny of five Iranian isolates of Cucumber mosaic virus from different hosts based on CP gene sequence

Bioassay and phylogeny of five Iranian isolates of Cucumber mosaic virus from different hosts based on CP gene sequence

Capsicum annum, a new host of watermelon mosaic virus

Partial characterization and development of sensitive and reliablediagnostic for the detection of cucumber mosaic virus

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