Amino acids in Tobamovirus coat protein controlling pepper L(1a) gene-mediated resistance

In pepper plants (genus Capsicum), the resistance against Tobamovirus spp. is conferred by L gene alleles. The recently identified L variant L^1a can recognize coat proteins (CPs) of Tobacco mild green mosaic virus Japanese strain (TMGMV‐J) and Paprika mild mottle virus Japanese strain (PaMMV‐J), but not of Pepper mild mottle virus (PMMoV), as the elicitor to induce resistance at 24 °C. Interestingly, L^1a gene‐mediated resistance against TMGMV‐J, but not PaMMV‐J, is retained at 30 °C. This observation led us to speculate that L^1a can discriminate between CPs of TMGMV‐J and PaMMV‐J. In this study, we aimed to determine the region(s) in CP by which L^1a distinguishes TMGMV‐J from PaMMV‐J. By using chimeric CPs consisting of TMGMV‐J and PaMMV‐J, we found that the chimeric TMGMV‐J CP, whose residues in the β‐sheet domain were replaced by those of PaMMV‐J, lost its ability to induce L^1a gene‐mediated resistance at 30 °C. In contrast, the chimeric PaMMV‐J CP with the β‐sheet domain replaced by TMGMV‐J CP was able to induce L^1a gene‐mediated resistance at 30 °C. Furthermore, viral particles were not detected in the leaves inoculated with either chimeric virus. These observations indicated that the amino acids within the β‐sheet domain were involved in both the induction of L^1a gene‐mediated resistance and virion formation. Further analyses using chimeric CPs of TMGMV‐J and PMMoV indicated that amino acids within the β‐sheet domain alone were not sufficient for the induction of L^1a gene‐mediated resistance by TMGMV‐J CP. These results suggest that multiple regions in Tobamovirus CP are implicated in the induction of L^1a gene‐mediated resistance.     

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

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

Nucleotide sequence analysis of a cDNA clone encoding the 34K movement protein gene of odontoglossum ringspot virus,ORSV-Cy,the Korean isolate

The partial nucleotide sequence of the 3-terminal region of the Korean isolate of odontoglossum ringspot tobamovirus (ORSV-Cy) from cool-growing Cymbidium was determined. The sequence contained a full length open reading frame (ORF) coding for the viral cell-to-cell movement protein (MP). The ORF was located upstream of the coat protein gene and 105 nucleotides longer than that of tobacco mosaic virus (TMV). The ORF predicts a polypeptide chain of 303 amino acids with a molecular weight of 33573. The ORF contained a similar region of conserved sequence motif of tobamoviruses and putative assembly origin of the viral RNA was located at about 1,100 nucleotides away from the 3 end. The predicted amino acid sequence for the MP gene of ORSV-Cy is more closely related to pepper mild mottle virus (PMMV), TMV-vulgare and TMV-Rakkyo than to tobacco mild green mosaic virus (TMGMV), TMV-L, cowpea strain of TMV (SHMV), and cucumber green mottle mosaic virus (CGMMV).     

A century of tobamovirus evolution in an Australian population of Nicotiana glauca.

The evolution over the past century of two tobamoviruses infecting populations of the immigrant plant Nicotiana glauca in New South Wales (NSW), Australia, has been studied. This plant species probably entered Australia in the 1870s. Isolates of the viruses were obtained from N. glauca specimens deposited in the NSW Herbarium between 1899 and 1972, and others were obtained from living plants in 1985 and 1993. It was found that the NSW N. glauca population was infected with tobacco mosaic tobamovirus (TMV) and tobacco mild green mosaic tobamovirus (TMGMV) before 1950 but only with TMGMV after that date. Half the pre-1950 infections were mixtures of the two viruses, and one was a recombinant. Remarkably, sequence analyses showed no increase in the genetic diversity among the TMGMV isolates over the period. However, for TMV, the genetic diversity of synonymous (but not of nonsynonymous) differences between isolates varied and was correlated with their time of isolation. TMV accumulated to smaller concentrations than TMGMV in N. glauca plants, and in mixed experimental infections, the accumulation of TMV, but not of TMGMV, was around 1/10 that in single infections. However, no evidence was found of isolate-specific interaction between the viruses. We conclude that although TMV may have colonized N. glauca in NSW earlier or faster than TMGMV, the latter virus caused a decrease of the TMV population below a threshold at which deleterious mutations were eliminated. This phenomenon, called Muller's ratchet, or a "mutational meltdown," probably caused the disappearance of TMV from the niche.     

First Report of Cucumber mosaic virus Infecting Chinese Mallow in China

The virus in naturally infected, stunted Chinese mallow plants and mosaic leaves was identified as Cucumber mosaic virus (CMV). Six symptomatic plants and one symptomless plant were collected in Chongqing, China. DAS‐ELISA suggested CMV was likely associated with the diseased Chinese mallow. Double‐stranded RNA was extracted from the samples, analysed by RT‐PCR, and the coding sequences of their coat proteins (CPs) were sequenced. The results further confirmed CMV was the pathogen causing Chinese mallow stunted, mosaic disease. The isolate was named CMV‐DXC. The full sequence of CMV‐DXC CP was determined, and it had the highest nucleotide identity (99.4%) of those of CMV‐lily, CMV‐WSJ and CMV‐Hnt, respectively. Phylogenetic analysis shows that CMV‐DXC belongs to CMV subgroup II. To our knowledge, this is the first report of CMV infecting Chinese mallow in China.     

Identification and Characterization of a Potyvirus Isolated from Siratro Plants

The present work describes the identification and characterization of a potyvirus isolated from siratro (Macroptilium atropurpureum Urb.) in the north‐west region of the State of São Paulo, Brazil. The virus was transmitted by mechanical inoculation. Its host range was restricted mainly to members of the Fabaceae. A cDNA fragment of about 930 bp was amplified by RT/PCR, cloned and sequenced. The fragment, which included the coat protein gene, had amino acid identity percentages between 88 and 98% with isolates of Bean common mosaic virus (BCMV). Phylogenetic analysis grouped the siratro potyvirus and BCMV isolates in 99% of the replicates, including Azuki mosaic virus, Dendrobium mosaic virus, Blackeye cowpea mosaic virus and Peanut stripe virus, which have been classified as BCMV strains. This is the first citation on the presence of BCMV in siratro plants in Brazil.     

Biological and Molecular Characterization of Alfalfa Mosaic Virus Infecting Trumpet Creeper (Campsis radicans) in Iran

Samples of trumpet creeper (Campsis radicans) leaves showing mottling and mosaic were collected from plants growing in a private garden in Tehran province, Iran, in 2012. Symptomatic leaf samples were tested for Alfalfa mosaic virus (AMV), Cucumber mosaic virus (CMV) and Peanut stunt virus (PSV) infection in enzyme‐linked immunosorbent assay (ELISA), using specific antibodies. None of the samples were positive for CMV and PSV; however, all reacted positively with that of AMV antiserum. In biological assay, systemic infection was found on Datura stramonium, Nicotiana tabacum cvs., White Burley, and Xanthi, 21 days postinoculation (DPI), while necrotic local lesions were obtained following inoculation of Phaseolus vulgaris and Vigna unguiculata within three to four DPI. Using a pair of primers specific for AMV, a DNA fragment of 880 bp was RT‐PCR‐amplified. Analysis of the sequences revealed the presence of 657 nucleotides of AMV complete coat protein (CP) gene (translating 218 amino acid residues). Phylogenetic analysis using neighbour‐joining (NJ) method clustered AMV isolates into two main types and the IRN‐Tru (GenBank Accession No. JX865593 ) isolate fell into type I. Pairwise nucleotide distances also confirmed two main types with the highest and lowest similarities for type I and II, respectively. The association of AMV with mosaic disease of C. radicans represents the first record from the world.     

Molecular phylogeny of geminivirus infecting wild plants in Japan

Few studies have been made on the molecular divergence of plant viruses. To remedy this deficiency, we examined the molecular divergence of the tobacco leaf curl geminivirus (TLCV). TLCV infects not only tobacco but alsoEupatorium andLonicera in the field and causes yellow vein disease. A total of 29 nucleotide sequences of the replication protein gene (ORF C1) of geminiviruses infecting wild plants ofE. makinoi, E. glehni andL. japonica collected from ten localities was determined. Highly divergent sequences were obtained not only among host plant populations but also within a host population. Phylogenetic analyses showed that the TLCVs infectingEupatorium andLonicera were clustered into three different clades, and were either paraphyletic or polyphyletic. This result is the first evidence demonstrating that wild populations of single plant species possess genetically diversified virus strains. Comparison with recently reported genetic variations of tobacco mild green mosaic tobamovirus (TMGMV) revealed three characteristics of TLCV evolution: (1) a higher nucleotide substitution rate, (2) more frequent migration among geographically isolated host populations, and (3) more frequent host changes to different plant families. While TMGMV is an RNA virus, TLCV has DNA genomes. In animal viruses, RNA viruses tend to evolve faster than DNA viruses. Our results indicated that this trend may not hold for plant viruses.     

Properties of the coat protein of the tobacco mosaic virus Kazakh isolate K1

A study was made of the coat protein (CP) of thermosensitive semidefective tobacco mosaic virus strain K1 (TMV-K1). In contrast to CP of other TMV strains, K1 CP showed high nonspecific aggregation and did not form normal two-layered cylindrical aggregates. In none of the conditions tested, K1 CP formed virions with cognate K1 RNAin vitro. The abnormal properties were attributed to substitution Lys53→Glu differentiating the K1 CP from those of other tobamoviruses. It is assumed that the high structural plasticity allows the tobamovirus virions to incorporate CP subunits even with unfavorable amino acid changes.     

The Natural Occurrence of Turnip Mosaic Potyvirus in Allium ampeloprasum

An isolate of turnip mosaic potyvirus (TuMV) was obtained from Allium ampeloprasum grown in commercial greenhouses in Israel. Symptoms on infected plants include systemic chlorosis and yellow stripes, accompanied by growth reduction. Leaves were distorted, often showing necrotic flecking. The virus was readily transmitted mechanically, and in a non-persistent manner by aphids, among Allium, Chenopodium. Gomphrena and some Nicotiana spp. Purified preparations contained numerous filamentous particles similar to those observed in crude extracts of infected leaves. Particles from crude plant extracts had a normal length of 806 nm. Cells of infected plants contained cylindrical cytoplasmic inclusions with pinwheel, scrolls and laminated aggregates which indicated the presence of a potyvirus of Edwardson's subgroup III. and which resemble those of turnip mosaic virus (TuMV), The virus reacted strongly with antiserum to typical isolates of TuMV in immunoelectron microscopy and western blotting but not with antisera to several other potyviruses. Based on serological reactivity, electron microscopy, aphid transmission and cytopathology, the virus was identified as an isolate of TuMV.     

Nicotiana velutina mosaic virus: purification, properties and affinities with other rod-shaped viruses

Nicotiana velutina mosaic virus (NVMV), found in Australia, was transmitted by inoculation of sap to twenty species in the Solanaceae and Chenopodiaceae, and to Gomphrena globosa; its host range closely resembles that of potato mop-top virus (PMTV). Infectivity was abolished when sap was kept at room temperature between 1 and 4 days, or when heated for 10 min between 60 and 70 °C. NVMV was frequently transmitted through the seed of four Nicotiana spp. NVMV and PMTV were purified by a method that involved redissolving virus particles sedimented by low speed centrifugation of leaf extracts, followed by sedimentation through sucrose cushions. NVMV preparations contain rod-shaped particles about 18 nm wide and with a large range of lengths, the commonest being 125–150 nm. The particles have a helical structure with a pitch of 2–9 nm, break easily, and contain a single protein of apparent mol. wt. 21|400, slightly larger than that of PMTV (19 800). In serological tests assessed by electron microscopy, no relationship was detected between NVMV and PMTV, or barley stripe mosaic, beet necrotic yellow vein, soil-borne wheat mosaic, tobacco mosaic or tobacco rattle viruses. However, antiserum to soil-borne wheat mosaic virus reacted quite strongly with PMTV and weakly with tobacco mosaic virus. NVMV is considered to be a distinct member of the tobamovirus group; its frequent transmission through seed may be an adaptation to the arid environment where it was found. Its cryptogram is */*:*/*:E/E:S/*.     

Viruses detected in Ullucus tuberosus (Basellaceae) from Peru and Bolivia

Ullucus tuberosus (Basellaceae) plants from 12 locations in the Andean highlands of Peru and Bolivia contained complexes of either three or four viruses. Specimens from six sites in Peru contained a potexvirus, a tobamovirus, a potyvirus and a comovirus, but those from another location lacked the potexvirus. All samples from five sites in Bolivia lacked the tobamovirus. The potexvirus (PMV/U) is a strain of papaya mosaic virus differing slightly from the type strain (PMV/T) in inducing milder symptoms in some common hosts and failing to infect a few other species. It symptomlessly infected U. tuberosus, and infected 15 of 29 species from seven of nine other families. PMV/U showed a close serological relationship to PMV/T and to boussingaultia mosaic virus and a distant relationship to commelina virus X, but it is apparently unrelated to any of ten other potexviruses. The tobamovirus (TMV/U) induced symptomless or inconspicuous infection in U. tuberosus, and infected 21 of 30 species from six of eight other families. It showed a very distant serological relationship to some strains of ribgrass mosaic, tobacco mosaic and tomato mosaic viruses, but failed to react with antisera to cucumber green mottle mosaic, frangipani mosaic, odontoglossum ringspot and sunn-hemp mosaic viruses. The potyvirus, tentatively designated ullucus mosaic virus (UMV), alone in U. tuberosus induced leaf symptoms indistinguishable from the chlorotic mottling and distortion found in naturally infected plants. UMV infected 12 of 20 species from four other families, and was transmitted in the non-persistent manner by Myzus persicae. It showed a distant serological relationship to only two (bidens mottle and alstroemeria mosaic) of 25 members or possible members of the potyvirus group tested. Some hosts and properties of the comovirus are described in an accompanying paper. None of the four viruses infected potato (Solanum tuberosum) and, with the possible exception of UMV, they differed from viruses reported previously to infect three other vegetatively propagated Andean crops (Oxalis tuberosa, Arracacia xanthorrhiza and Tropaeolum tuberosum).     

Disease distribution and characterisation of a new macluravirus associated with chirke disease of large cardamom

Large cardamom (Amomum subulatum), an important spice crop grown in eastern sub‐Himalayan mountains of India, is affected by a viral disease commonly known as ‘chirke’, which is characterised by light and dark green streaks on the leaf lamina. Although chirke has been known to affect large cardamom for over 50 years, its distribution in large cardamom growing regions and aetiology have remained unaddressed. In this study for the first time, distribution of chirke in the major large cardamom growing regions in India has been determined. North Sikkim and eastern region of Darjeeling hills were endemic region with average disease incidence of 19.2–35%, whereas, Mirik region of Darjeeling hills were free from the disease. Suckers, the commonly used planting material, were the major source for spread of the disease. The virus was sap transmissible to the popular large cardamom cultivars Golsey, Ramsey, Swaney and Varlangey and vectored by Rhopalosiphum maidis and Myzus persicae in a non‐persistent manner. Flexuous virus particles measuring 625–650 × 12.5 nm were observed consistently associated with the diseased samples. Polyclonal antiserum to the purified virus showed serological affinity with a macluravirus, cardamom mosaic virus (CdMV) associated with a similar disease known as katte disease of small cardamom (Elettaria cardamomum) occurring in southern India. The 3^′ terminal genome sequence (1776 nucleotides) of the virus was determined, which revealed a close sequence identity and phylogenetic relationships with the members of the genus Macluravirus. The deduced amino acid sequence of putative coat protein (CP) gene showed maximum similarity of 65.7% with the CdMV. Phylogenetic analysis based on CP and 3^′ UTR showed that the virus was closer to Alpinia mosaic virus, CdMV and Chinese yam necrotic mosaic virus subclade. The results suggest that the virus associated with the chirke disease of large cardamom is a new species under the genus Macluravirus in the family Potyviridae for which the name large cardamom chirke virus (LCCV) is proposed.