BCMV-ukr: isolate of bean common mosaic virus revealed in Ukraine

SUMMARY

Bean common mosaic virus (BCMV) is distributed worldwide and causes a serious disease in bean reducing growth and crops yield. The aim of this study was to investigate the occurrence of BCMV and Bean common mosaic necrotic virus (BCMNV) in Ukraine, to characterise host range and reactions of indicator plants to mechanical inoculation with the isolate and to differentiate it by using Drijfhout’s differentials. Leaf samples were positive for BCMV infection in RT-PCR assay employing specific primers with amplification of a 340-bp product. Based on a biological test on bean differentials, the isolate was assigned to pathogroup VII despite the fact that strain differed markedly from the standard strains in symptoms producing on differential groups IV and V. Partial sequence data of the coat protein region show 100% identity with BCMV 125 sequences tested. To our knowledge, this is the first attempt to characterise the BCMV circulating in Ukraine.     

Relationships among strains of bean common mosaic virus and blackeye cowpea mosaic virus - members of the potyvirus group

In host-range studies, bean common mosaic virus strains (BCMV-NL1, -NL3 and -NY 15) usually induced distinct systemic symptoms in susceptible bean cultivars and latent infection in several Vigna genotypes (except NY15 which gave mosaic symptoms in the latter), while blackeye cowpea mosaic virus (B1CMV-W) caused distinct systemic symptoms in several Vigna genotypes and only weak systemic symptoms in a few bean genotypes only. Biologically, B1CMV-W was closest to BCMV-NY15 and less close to -NL1. When using antisera to the three BCMV strains and five strains of B1CMV (including a strain originally considered cowpea aphid-borne mosaic virus CAMV-Mor) in SDS-immunodiffusion and ELISA, BCMV-NL1 and -NY15 were found to be closely related to each other and to BICMV-Fla, -NR and -W, and less closely to BICMV-Ind and -Mor. Serological relationships of BCMV-NL1 and -NY15 to BCMV- NL3 were more distant, which is in line with the biological distinction of NL3 in causing temperature-independent necrosis in bean cultivars with the necrosis gene I. PAGE analysis of coat proteins revealed that the three strains of BCMV and B1 CMV-W have similar but non-identical molecular masses. Although molecular hybridisation may further elucidate quantitative relationships between potyvir-uses, variation within and among the potyviruses may continue to pose problems in their classification and identification.     

The visualization of local lesions caused by the common bean mosaic virus

A very simple and quick procedure for the visualization of local lesions caused by the common bean mosaic virus was developed. It consists in a treatment of bean leaves in 96 % èthanol 50 °C warm and in the subsequent staining of the leaves in Lugol solution and washing in 30 % ethanol.     

Immunocapture RT-PCR detection of Bean common mosaic virus and strain blackeye cowpea mosaic in common bean and black gram in India

The strains of Bean common mosaic virus (BCMV) and blackeye cowpea mosaic (BICM), genus Potyvirus, were detected from 25 common bean and 14 black gram seeds among 142 seed samples collected from different legume-growing regions of India. The samples were subjected to a growing-on test, an indicator plant test, an electron microscopic observations, an enzyme linked immunosorbent assay and an immunocapture RT-PCR. The incidence of the two tested viruses in common bean and black gram seed samples was 1–6% and 0.5–3.5%, respectively in growing-on test evaluations. Electron microscopic observations revealed filamentous virion particles from the leaves of plants showing characteristic virus disease symptoms in growing-on and host inoculation tests. The identity of the strains was confirmed by immunocapture RT-PCR, with a final amplification product of approximately 700 bp for BCMV and BCMV–BICM. The complete identity of the two viruses was further confirmed by nucleotide sequencing of the partial coat protein and 3′-UTR regions. The sequences of the four BCMV and BCMV–BICM isolates each consisted of 583–622 and 550–577 nucleotides. The present report confirms the widespread nature of these two serious potyviruses in the two most important legume crops in India.     

Interaction of Cucumber mosaic virus and Bean yellow mosaic virus in co-infected plants of bean and broad bean

After evaluation of the responses of bean and broad bean common cultivars against an isolate of Cucumber mosaic virus (CMV-K) and Bean yellow mosaic virus (BYMV-K), interaction of isolates was statistically studied on co-infected plants of bean cv. Bountiful and broad bean cv. Lahijan at two trials. Based on viral relative concentration determined by quantitative enzyme-linked immunosorbent assay, BYMV interacts synergistically with CMV in bean at 14 days post inoculation, while in co-infection with BYMV, CMV interacts antagonistically in both host plants at least in one of the two trials. This suggests that CMV/BYMV interaction is dependent on host species and developmental stage of plant. Co-infection like single infection with CMV in bean plants led to significantly decrease in plants’ height and fresh weight than BYMV singly infected and healthy plants, while viral infection of broad bean plants did not significantly affect growth parameters. Decline effect of viral infection (especially co-infection) on chlorophyll and carotenoids value of bean plants was greater than those of broad bean. Viral infection (singly or doubly) caused irregular changes in nutrient elements values of both hosts compared with healthy ones.     

Sequence analysis of a soil-borne wheat mosaic virus isolate from Italy shows that it is the same virus as European wheat mosaic virus and Soil-borne rye mosaic virus

The complete sequence of the two RNAs of a furovirus isolate from durum wheat in Italy was determined. Sequence comparisons and phylogenetic analysis were done to compare the Italian virus with Soil-borne wheat mosaic virus (SBWMV) from the USA and with furovirus sequences recently published as European wheat mosaic virus (EWMV), from wheat in France, and Soil-borne rye mosaic virus (SBRMV), from rye and wheat in Germany. Over the entire genome, the Italian isolate RNA1 and RNA2 had respectively 97.5% and 98.6% nucleotide identity with EWMV, 95.5% and 85.8% with SBRMV-G and 70.6% and 64.5% with SBWMV. The Italian isolate was therefore clearly distinct from SBWMV. The European isolates all appear to belong to the same virus and the name Soil-borne cereal mosaic virus may resolve earlier ambiguities.     

The comparative morphology and anatomy of Dioscorea sylvatica Eckl. from Natal and the Transvaal

The morphology of Dioscorea sylvatica from Natal and the hills in the Lydenburg district of the Transvaal is described, as well as the anatomy of the leaf and stem. Differences were noticed between the Natal and Lydenberg types which are inconsistent with Burkill's classification of the species. From the differences it is suggested that the Lydenberg type be designated as D. sylvatica subspecies lydenbergensis subsp. nov.     

Metal-free southern bean mosaic virus crystals.

Native southern bean mosaic virus contains a significant number of Mg2+ and Ca2+ ions. These can be removed by treatment with EDTA causing the virus to swell by 7% in radius at alkaline pH values. The swollen virions are susceptible to protease and nuclease digestion. They are likely to be an intermediate during assembly and disassembly. Crystals of the metal-free virus have been grown and were found to be approximately isomorphous with the orthorhombic type III southern bean mosaic virus crystals (Akimoto, T., Wagner, M.A., Johnson, J.E., and Rossmann, M.G. (1975) J. Ultrastruct. Res. 53, 306-318), although the cell dimensions are longer by 2%. Native rhombohedral type II crystals disintegrate on changing the pH or increasing the ionic strength of the mother liquor. Damage can be prevented by addition of ethylene glycol. At alkaline pH values, these crystals also show a 2% increase in their cell dimensions as well as a significant alteration in their diffraction patterns. In the type II and III crystals, the viruses pack with only their 5-fold axes in contact. Thus, the difference of the apparent swelling in solution and in the crystals may be one of differential swelling over the virus surface.     

A Prague isolate of wisteria vein mosaic virus

An isolate of wisteria vein mosaic virus (WVMV) obtained fromWisteria sinensis in Prague resembled in its properties WVMV isolates described in Italy and Holland.Nicotiana megalosiphon is reported as a new host of WVMV. Other known host plants showed reactions similar to those described formerly. The incubation period extended in some hosts up to two or four weeks. Pea plants showed symptoms within five to seven days. Species ofApium, Brassica,Datura and others were not susceptible. TIP of Prague isolate of WVMV was 61 °C, at a dilution 1: 5000 47% of plants were infected, and 48 h > LIV > 28 h. Modal particle length was 743 nm. Massive granular inclusions were seen in pea epidermal cells, usually adhering to nuclei that did not show alterations. No serological relation to turnip mosaic virus could be established.     

The insect transmission of an isolate of okra mosaic virus occurring in Nigeria

Three species of chrysomelid beetles (Podagrica sjostedti, P. uniforma and Syagrus calcaratus) transmitted a Nigerian isolate of okra mosaic virus (OMV) to okra (Abelmoschus esculentus) and green gram (Vigna aureus), two (P. uniforma and S. calcaratus) also to cotton (Gossypium hirsutum) and one (P. uniforma) also to cowpea. Using one viruliferous beetle per test plant, P. sjostedti, P. uniforma and S. calcaratus infected 63, 68 and 34% of test plants respectively. Virus was retained by these vectors for up to 6 days. Whiteflies (Bemisia tabaci) also transmitted OMV, although much less efficiently than the beetle vectors.     

Distinction of strains of bean common mosaic virus and blackeye cowpea mosaic virus using antibodies to N- and C- or N-terminal peptide domains of coat proteins

Earlier attempts to discriminate serologically strains NL1, NL3 and NY15 of bean common mosaic virus (BCMV) and strain W of blackeye cowpea mosaic virus (B1CMV) had been unsuccessful. Antibodies directed towards N- and C-, or N-terminal peptide regions of the coat proteins of the above strains enabled the distinction between B1CMV-W, BCMV-NY15 and BCMV-NL3 in electroblot immunoassay and in ELISA. The distinction was better with antibodies directed towards N-termini than with those to N- and C-termini. Strain NL1 of BCMV cross-reacted with both B1CMV-W and BCMV-NY15, but not with BCMV-NL3. Taxonomic implications of these findings are discussed.     

Identification of soybean mosaic, southern bean mosaic and tobacco ringspot viruses from soybean in the People's Republic of China

Samples of soybean plants with virus-like symptoms were collected from several locations in the People's Republic of China in 1981. These samples were used to prepare inocula for mechanical inoculation to soybean. Twenty-one virus cultures were obtained, the identities of which were determined by serology, symptomatology and host range. Sixteen cultures contained only soybean mosaic virus, four of which were more pathogenic than any previously studied; one culture contained only tobacco ringspot virus, another only southern bean mosaic virus, and three other cultures mixed infections of soybean mosaic and southern bean mosaic viruses. This is the first report of the occurrence of tobacco ringspot virus and southern bean mosaic virus in soybean in the People's Republic of China.     

Application of in silico bulked segregant analysis for rapid development of markers linked to Bean common mosaic virus resistance in common bean

Background

Common bean was one of the first crops that benefited from the development and utilization of molecular marker-assisted selection (MAS) for major disease resistance genes. Efficiency of MAS for breeding common bean is still hampered, however, due to the dominance, linkage phase, and loose linkage of previously developed markers. Here we applied in silico bulked segregant analysis (BSA) to the BeanCAP diversity panel, composed of over 500 lines and genotyped with the BARCBEAN_3 6K SNP BeadChip, to develop codominant and tightly linked markers to the I gene controlling resistance to Bean common mosaic virus (BCMV).

Results

We physically mapped the genomic region underlying the I gene. This locus, in the distal arm of chromosome Pv02, contains seven putative NBS-LRR-type disease resistance genes. Two contrasting bulks, containing BCMV host differentials and ten BeanCAP lines with known disease reaction to BCMV, were subjected to in silico BSA for targeting the I gene and flanking sequences. Two distinct haplotypes, containing a cluster of six single nucleotide polymorphisms (SNP), were associated with resistance or susceptibility to BCMV. One-hundred and twenty-two lines, including 115 of the BeanCAP panel, were screened for BCMV resistance in the greenhouse, and all of the resistant or susceptible plants displayed distinct SNP haplotypes as those found in the two bulks. The resistant/susceptible haplotypes were validated in 98 recombinant inbred lines segregating for BCMV resistance. The closest SNP (~25-32 kb) to the distal NBS-LRR gene model for the I gene locus was targeted for conversion to codominant KASP (Kompetitive Allele Specific PCR) and CAPS (Cleaved Amplified Polymorphic Sequence) markers. Both marker systems accurately predicted the disease reaction to BCMV conferred by the I gene in all screened lines of this study.

Conclusions

We demonstrated the utility of the in silico BSA approach using genetically diverse germplasm, genotyped with a high-density SNP chip array, to discover SNP variation at a specific targeted genomic region. In common bean, many disease resistance genes are mapped and their physical genomic position can now be determined, thus the application of this approach will facilitate further development of codominant and tightly linked markers for use in MAS.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-903) contains supplementary material, which is available to authorized users.