Broad Bean True Mosaic Virus Research Articles

Advancing broad bean true mosaic virus detection using conventional RT-PCR and real-time RT-PCR with novel primer set design

Broad bean true mosaic virus (BBTMV) infects broad beans and peas, reducing yield. As BBTMV is transmitted through broad beans, many countries have implemented regulations to prevent the distribution of infected seeds. Currently, enzyme-linked immunosorbent assay (ELISA) is commonly used to detect BBTMV. While the PCR-based method is preferred for seed virus detection due to its sensitivity and speed. A BBTMV-specific PCR detection method has not yet been reported. A universal detection method currently exists that utilizes reverse transcription PCR (RT-PCR) for the Comovirus genus, to which BBTMV belongs. However, sequence analysis is required for species identification. To address this limitation, we developed and verified RT-PCR detection methods using newly designed BBTMV-specific primers. RT-PCR and real-time RT-PCR with these primers were approximately 5 × 105–106 times more sensitive than ELISA and 100–1000 times more sensitive than previously reported RT-PCR methods. Using RT-PCR and real-time RT-PCR employing these primers, we could detect BBTMV with same sensitivity when more than 3.0 × 105 copies were present per gram of broad bean seeds. Our newly developed detection methods can test for BBTMV with high sensitivity and speed.

Australian Cool-Season Pulse Seed-Borne Virus Research: 1. Alfalfa and Cucumber Mosaic Viruses and Less Important Viruses.

Here, we review the research undertaken since the 1950s in Australia's grain cropping regions on seed-borne virus diseases of cool-season pulses caused by alfalfa mosaic virus (AMV) and cucumber mosaic virus (CMV). We present brief background information about the continent's pulse industry, virus epidemiology, management principles and future threats to virus disease management. We then take a historical approach towards all past investigations with these two seed-borne pulse viruses in the principal cool-season pulse crops grown: chickpea, faba bean, field pea, lentil, narrow-leafed lupin and white lupin. With each pathosystem, the main focus is on its biology, epidemiology and management, placing particular emphasis on describing field and glasshouse experimentation that enabled the development of effective phytosanitary, cultural and host resistance control strategies. Past Australian cool-season pulse investigations with AMV and CMV in the less commonly grown species (vetches, narbon bean, fenugreek, yellow and pearl lupin, grass pea and other Lathyrus species) and those with the five less important seed-borne pulse viruses also present (broad bean stain virus, broad bean true mosaic virus, broad bean wilt virus, cowpea mild mottle virus and peanut mottle virus) are also summarized. The need for future research is emphasized, and recommendations are made regarding what is required.

Genome sequence and phylogenetic analysis of a novel comovirus from tabasco pepper (Capsicum frutescens).

A virus isolate from tabasco pepper (Capsicum frutescens) has been reported as a strain of the comovirus Andean potato mottle virus (APMoV). Using the replicative intermediate viral dsRNA, the pepper virus strain was sequenced by Illumina MiSeq. The viral genome was de novo assembled resulting in two RNAs with lengths of 6028 and 3646 nt. Nucleotide sequence analysis indicated that they corresponded to the RNA-1 and RNA-2 of a novel comovirus which we tentatively named pepper mild mosaic virus (PepMMV). Predictions of the open reading frame (ORF) of RNA-1 resulted in a single ORF of 5871 nt with five cistrons typical of comoviruses, cofactor proteinase, helicase, viral protein genome-linked, 3C-like proteinase (Pro), and RNA-dependent RNA polymerase (RdRP). Similarly, sequence analysis of RNA-2 resulted in a single ORF of 3009 nt with two cistrons typical of comoviruses: movement protein and coat protein (large coat protein and small coat proteins). In pairwise amino acid sequence alignments using the Pro-Pol protein, PepMMV shared the closest identities with broad bean true mosaic virus and cowpea mosaic virus, 56% and 53.9% respectively. In contrast, in alignments of the amino acid sequence of the coat protein (small and large coat proteins) PepMMV shared the closest identities to APMoV and red clover mottle virus, 54% and 40.9% respectively. A phylogenetic tree constructed using the conserved domains for the Pro-Pol from all members of the family Secoviridae confirmed the comovirus nature of the virus. Phylogenetic and sequence analyses supports proposing PepMMV as a new species of the genus Comovirus.

Role of Viral Diagnostics in Quarantine for Plant Genetic Resources and Preparedness

International exchange of plant genetic resources (PGR) and trade play an important role in the long-distance dissemination of destructive viruses/strains. The ICAR-NBPGR has been empowered for quarantine processing of imported PGR including transgenics meant for research purposes. Adopting a workable strategy of post-entry quarantine growing/inspection followed by use of combination of detection techniques viz., electron microscopy, ELISA and RT-PCR, 45 viruses of great economic and quarantine importance were intercepted in imported PGR in the last three decades (1985–2016). The interceptions include 19 viruses not yet reported from India viz., Barley stripe mosaic virus, Bean mild mosaic virus, Bean pod mottle virus, Broad bean mottle virus, Broad bean stain virus, Broad bean true mosaic virus, Cherry leaf roll virus, Cowpea mottle virus, Cowpea severe mosaic virus, Dioscorea latent virus, Garlic virus C, High plains virus, Maize chlorotic mottle virus, Pea enation mosaic virus, Peanut sunt virus, Pepino mosaic virus, Raspberry ringspot virus, Tomato ringspot virus and Wheat streak mosaic virus. Besides, 21 viruses not known to occur on particular host(s) in India were intercepted and 23 viruses were intercepted in germplasm imported from CGIAR centres. Fourteen viruses including nine viruses not reported from India have been intercepted in transgenic germplasm. Even though some of the intercepted viruses are not known to occur in India, their potential vectors exist and also congenial conditions to multiply, disseminate and spread destructive exotic viruses/strains. India also need to establish a strong network of interconnected accredited laboratories able to quickly diagnose new viruses/strains; enhance surveillance capacity and develop early warning systems. This makes it imperative that India should put in place a “National Plant Pests Diagnostic and Certification Network” to enhance preparedness.

Revolutionize in Protein Patterns of Different Vicia faba L. Cultivars Infected with Broad bean true mosaic virus under Salicylic Acid Treatments

Revolutionize in Protein Patterns of Different Vicia faba L. Cultivars Infected with Broad bean true mosaic virus under Salicylic Acid Treatments

Susceptibility of pea, horse bean and bean to viruses in dependence on the age of the inoculated plants

Three cultivars of pea did not differ in their susceptibility to Cucumber Mosaic Virus (CMV) notwithstanding the age of the inoculated plants. But their susceptibility to infection with Bean Yellow Mosaic Virus (BYMV) differed. Horse bean cultivars 'Nadwiślański' and 'Major' proved to be less susceptible to Broad Bean True Mosaic Virus (BBTMV) when older plants were-inoculated. Two bean cultivars 'Złota Saxa' and 'Earle' appeared to be susceptible to BBTMV only in the phase of developing primary leaves and the age-dependent resistance to infection increased faster in plants of the cv. 'Złota Saxa'. Both cultivars of bean showed also age-dependent resistance to infection by BYMV. All these viruses restricted growth and yield of plants. The decreases were greater when younger plants were inoculated. These dependences appeared most distinctly in pea cv. 'Sześciotygodniowy' infected with CMV and in two cultivars of bean infected with BYMV.

Complete genome sequence of broad bean true mosaic virus

The complete genome sequence of a severe isolate of broad bean true mosaic virus (genus Comovirus, subfamily Comovirinae, family Secoviridae) is presented. Comparison of the amino acid sequences of the capsid proteins and the polymerase showed striking differences to other comoviruses and highest similarities to legume-infecting comoviruses. Red clover mottle virus was recognized as the most similar virus with amino acid sequence identities ranging from 43 to 67% for individual genes.

Further studies on Pea seed-borne mosaic virus in cool-season crop legumes: responses to infection and seed quality defects

Field and glasshouse experiments (3 of each) were done during 2003–06 to determine the responses of a range of genotypes belonging to 13 species of cool-season crop legumes to infection with Pea seed-borne mosaic virus (PSbMV). Seed quality defects were determined and genotypes of some species were also tested for seed transmission of the virus. In field experiments, of 39 genotypes of field pea (Pisum sativum) evaluated, 15 were ranked as highly susceptible, 10 susceptible, 9 moderately resistant, and 5 resistant, while all 7 lupin species (Lupinus spp.) tested were resistant. In glasshouse sap and graft inoculations with PSbMV to genotypes not found infected in the field and 2 additional lupin species, no virus was detected in any of the 9 lupin species or in 5 field pea genotypes tested. Thus, the lupins all appeared to be non-hosts and the 5 field pea genotypes had resistance to the 2 PSbMV isolates used to inoculate them. All 14 genotypes of faba bean (Vicia faba) evaluated in the field were ranked highly susceptible, while 12 out of 16 lentil (Lens culinaris) genotypes were ranked as highly susceptible and 4 as susceptible. Chickpea (Cicer arietinum) genotypes were moderately resistant (50) or susceptible (7). Once infected, plant sensitivities (symptom severities) ranged from low in some field pea and most lentil genotypes to high in most faba bean genotypes. Chickpea genotypes all were ranked as moderately sensitive. Seed lots harvested from PSbMV-infected plants of field pea, faba bean, and chickpea all showed severe seed quality defects, but lentil was usually less affected. The predominant seed symptoms were necrotic rings and line markings on the seed coat, malformation, reduced size, and splitting. Kabuli chickpea types also showed darkening of the seed coat. Seed transmission of PSbMV was detected in faba bean (0.2%) and field pea (5–30%). When PSbMV infection foci were introduced into plots of lentil cv. Nugget, the virus spread to the lentil plants and decreased shoot dry weight by 23%, seed yield by 96%, and individual seed weight by 58%. Seed transmission of PSbMV (6%) was detected in seed from the infected lentil plants. In a survey for possible viral seed symptoms, all seed lots of kabuli chickpea (5) and field pea (70), and 10 of 18 of faba bean were affected, but none of the 23 of lentil. When seedlings from 16 faba bean and 7 field pea seed lots were tested for 3 viruses, neither Broad bean stain virus nor Broad bean true mosaic virus was detected, but PSbMV was found in 5 field pea seed lots at incidences of <1–14%. PSbMV was detected in commercial field pea seed stocks of cvv. Kaspa (33) and Parafield (12) at incidences of 0.5–47% and 0.3–30%, respectively. The implications of these findings in terms of genotype susceptiblility and sensitivity to PSbMV infection and their importance for the management of PSbMV in legume crops are discussed.

First record of Beet western yellows virus, Chickpea chlorotic dwarf virus and Faba bean necrotic yellows virus affecting faba bean (Vicia faba) crops in Iraq

A survey to identify virus diseases affecting faba bean(Vicia faba) in Iraq was conducted during April 2000.Around 4400 samples were collected from 36 faba beanfields distributed in five governorates (ten in Baghdad,nine in Al-Anbar, three in Diyala, five in At-Tamim andnine in Ninawa). These samples were tested for thepresence of 12 different viruses by tissue-blot immu-noassay (Lin et al., 1990), using a battery of polyclonaland monoclonal antibodies. Rabbit polyclonal antiseraspecific for Chickpea chlorotic dwarf virus (CCDV)(Horn et al., 1993); Broad bean true mosaic virus(BBTMV) (from H. Rohloff, BBA, Braunschweig,Germany); Broad bean wilt virus (BBWV); Pea enationmosaic virus (PEMV) (from IPO, Wageningen, theNetherlands); Alfalfa mosaic virus (AMV); Bean yellowmosaic virus (BYMV); Broad bean mottle virus(BBMV); Broad bean stain virus (BBSV); Cucumbermosaic virus (CMV); and Pea seed-borne mosaic virus(PSbMV) (from Virology Laboratory, ICARDA,Aleppo, Syria) were used, in addition to a monoclonalspecific for Faba bean necrotic yellows virus (FBNYV)(Franz et al., 1996) and a broad-spectrum legumeluteovirus monoclonal (5G4) (Katul, 1992). To identifyspecific luteoviruses, samples that tested positive usingmonoclonal 5G4 were tested against five furthermonoclonals: Beet western yellows virus (BWYV)(ATCC PVAS-647); Soybean dwarf virus (SbDV) (ATCCPVAS-650); Potato leaf roll virus (PLRV) (ATCC PVAS-649); and Bean leaf roll virus (BLRV) (4B10) and 6F9,which reacts with either BLRV or SbDV (Katul, 1992).BYMV was the most abundant virus in faba bean inIraq, identified in 68% of test samples. In 50% of thefields surveyed, BYMV was identified at extremely highlevels (80–100% infection). BWYV was detected in 135samples collected from 15 fields from all five governor-ates. Two fields in the Ninawa governorate were highlyinfected with BWYV (62 and 96%). CCDV wasdetected in eight samples collected from three fields(two in El-Anbar and one in Ninawa), and FBNYV wasdetected in seven samples collected from two fields inNinawa governorate.This is the first record of BWYV, CCDV and FBNYVaffecting faba bean in Iraq. Other viruses such as AMV,BBWVand CMV were detected in fewer than 1% of thesamples tested. BBMV, BBSV, BBTMV, PEMV andPSbMV were not detected in any of the samples.

Survey of Faba Bean ("Vicia faba" L.) Virus Diseases in Ethiopia

Surveys conducted in 1996 and 1997 to assess the status of virus diseases affecting faba bean in the major growing areas of Ethiopia indicated that leaf yellowing, rolling, necrosis and stunting were the most common disease symptoms. The highest visually-observed disease incidence in a field was 85%, recorded in the Wello region (north-eastern Ethiopia). When 3049 symptomatic samples collected from 211 fields from all over Ethiopia were tested by the tissue blot immunoassay (TBIA) for 14 viruses, 1592 samples (52.2%) were found to be infected with at least one virus. Faba bean necrotic yellows virus (FBNYV, genus Nanovirus) was the most frequent (63.2%), followed by luteoviruses (28.5%) [such as Beet western yellows virus (BWYV, genus Polerovirus, family Luteoviridae) and Bean leaf roll virus (BLRV, family Luteoviridae)] and Chickpea chlorotic dwarf virus (CCDV, genus Mastrevirus, family Geminiviridae) (3.1%). Mosaic/mottling symptoms were observed in some fields but incidence was always very low (<1%). The mosaic/mottling causing viruses identified were Broad bean stain virus (BBSV) and Broad bean true mosaic virus (BBTMV) (genus Comovirus, family Comoviridae), Pea seed-borne mosaic virus (PSbMV) and Bean yellow mosaic virus (BYMV) (genus Potyvirus, family Potyviridae). Testing of 3035 random samples collected in 1997 from 37 fields in the Shewa and Arsi regions revealed 1.6% luteovirus and 0.6% FBNYV infection, indicating that the incidence of these viruses on faba bean is very low in central Ethiopia. The survey suggested that virus diseases are currently of economic importance on faba bean in north-eastern Ethiopia and potentially important in the other areas, with FBNYV and luteoviruses being the most important.

Survey of faba bean (Vicia faba L.) for viruses in Morocco

A total of 52 faba-bean (Vicia faba L.) fields, located in the main growing areas in Morocco were surveyed for viruses. From 240 samples with symptoms suggestive of virus infection, the following viruses were detected using electron microscopy, serology, and biological indexing: Alfalfa mosaic virus (AMV), bean yellow mosaic virus (BYMV), broad bean mottle virus (BBMV), broad bean stain virus (BBSV), broad bean true mosaic virus (BBTMV), pea earlybrowning virus (PEBV), pea enation mosaic virus (PEMV), pea seed-borne mosaic virus (PSbMV), and a complex of luteoviruses including bean leafroll virus (BLRV). This is the first report of the occurrence of BBTMV, PEMV, PSbMV, and the luteoviruses (including BLRV) of faba bean in Morcco.

In vivo and In vitro Effects on the Electrophoretic Forms of Particles of Broad Bean True Mosaic Virus

SUMMARY Preparations of broad bean true mosaic virus (BBTMV) contained three centrifugal components: top, middle, and bottom. These had sedimentation coefficients of 59, 95, and 116S. Top component was only found when a reducing agent was used during all stages of purification and subsequent storage. When electrophoresed on acrylamide gels, particles of BBTMV migrated as two, and sometimes three, electrophoretic forms. In new infections a slower migrating form predominated; in older infections a faster migrating form predominated. The relative migration rates of the electrophoretic forms depended on the kind of infected leaf sampled, the environmental conditions during infection, the method of purification, and the period of storage after purification. The changes in mobility of the various forms are ascribed to limited proteolysis, which increased the negative charge of the virus particle by altering the charge of the coat protein. The coat protein is made up of two polypeptides, of mol. wt. 37500 and 20000 to 24500, respectively. The change in charge resulted from a decrease in size, and possibly a change in conformation, of the smaller polypeptide.

Vergleich zwischen Eigenschaften des Echten Ackerbohnenmosaik-Virus und des broad bean mottle-Virus

Broad bean true mosaic virus (EAMV) has been studied in comparison with broad bean mottle virus (BBMV), after both viruses had been purified by the same procedure. Spectrophotometric and/or chemical determinations revealed a nucleic acid content of 32% for EAMV and 23% for BBMV. Serologically, the two viruses are unrelated since in reciprocal testings they react with their homologous antisera only. The diameter of of the particles has been determined to 25 mμ for EAMV and 20 mμ for BBMV. Further differences include thermal inactivation point, seed transmission, host range, and symptomatology. There are no indications for a relationship among both viruses as suggested in the list of Common names (Review of Applied Mycology, Vol. 35, Suppl.).