Details of DPV and References

DPV NO: 408 December 2004

Family: Unallocated ssRNA+ viruses
Genus: Sobemovirus
Species: Southern bean mosaic virus | Acronym: SBMV

This is a revised version of DPV 274

Southern bean mosaic virus

Roger Hull Department of Disease and Stress Biology, John Innes Centre, Norwich Research Park, Colney, Norwich NR4 7UH, UK

Contents

Introduction

Described by Zaumeyer & Harter, 1942; 1943.

Selected synonyms
Bean mosaic virus 4 (Zaumeyer & Harter, 1942)
Southern bean mosaic virus 1 (Zaumeyer & Harter, 1943)
Marmor laesiofaciens (Zaumeyer & Harter, 1944)
Bean virus 4 (Zaumeyer & Harter, 1944)
Bean southern mosaic virus (Schmelzer & Pop, 1957)
Southern bean mosaic virus, bean strain

Type member of the genus Sobemovirus. A virus with isometric particles about 30 nm in diameter sedimenting as a single component. The genome consists of one species of positive-sense ssRNA. The virus has a very restricted host range, is transmitted by beetles (Chrysomelidae), by seed and by inoculation of sap. It occurs predominantly in warm temperate and tropical regions. This species has been separated from Southern cowpea mosaic virus (SCpMV) (Othman & Hull, 1995), which was previously known as Southern bean mosaic virus, cowpea strain.

Main Diseases

Causes mosaic and mottle diseases of economic importance in common bean (Phaseolus vulgaris), urd bean (V. mungo) and to a lesser extent in soybean (Glycine max). Good descriptions of disease symptoms in common bean are given by Zaumeyer & Harter (1943) and Yerkes & Patino (1960), and in urd bean by Singh & Singh (1975).

Geographical Distribution

Warm temperate and tropical regions of Africa (Givord, 1981; Shoyinka et al., 1979) and the Americas: USA, Mexico and Costa Rica (Gamez, 1977), Columbia (Morales & Castaño, 1985), and Nicaragua (Fuentes & Anderson, 1990). Reported also in Phaseolus vulgaris in France and Spain (Férault et al., 1969; Verhoeven et al., 2003) and Brazil (Cupertino et al., 1982), and in urd bean (Vigna mungo) in India (Singh & Singh, 1975).

Host Range and Symptomatology

The host range is very narrow; with the exception of Gomphrena globosa (Givord, 1981) only species of Leguminosae are susceptible. SBMV isolates rarely infect cowpea, and are distinguished from SCpMV, which rarely infects bean. Both viruses are readily transmissible by inoculation of sap.

Diagnostic species

Phaseolus vulgaris (common bean). The type and M (Mexican severe) strains induce local chlorotic spots or local lesions (Fig. 1) with or without systemic spread (Fig. 2) in Bountiful, Pinto, Blue Lake, Kentucky Wonder and other varieties (Grogan & Kimble, 1964). Systemic symptoms in common bean infected by these strains range from prominent mosaic with leaf distortion (especially in younger leaves) to mild mosaic, depending on variety.

P. lunatus (lima bean). Only small-seeded varieties are susceptible and show small necrotic lesions. Not reported to be susceptible to other strains.

Glycine max (soybean). Mild systemic mottle is induced by most strains, depending on variety.

Vigna radiata (mung bean). Isolates from urd bean induce systemic mosaic.

Gomphrena globosa. Systemic symptoms are induced by the Ivory Coast isolate (Givord, 1981). Not reported to be susceptible to other strains.

Propagation species

Systemically infected varieties of bean are commonly used for maintaining virus cultures and for propagating virus for purification.

Assay species

Local lesion varieties of bean such as Bountiful and Pinto (Grogan & Kimble, 1964) are suitable.

Strains

Bean (type) strain (Strain B) (Zaumeyer & Harter, 1943). Infects most common bean varieties systemically and induces local lesions in others; can infect a limited number of other legumes but not cowpea or other legumes susceptible to the cowpea strain.

Severe bean mosaic strain or Mexican strain (Strain M) (Yerkes & Patino, 1960; Grogan & Kimble, 1964). Causes more severe symptoms (e.g. local necrosis and systemic symptoms, often with necrosis) in common bean than the type strain; also infects cowpea.

Resistance-breaking strains have been isolated from systemically infected bean cultivars resistant to the type strain (Valverde & Fulton, 1982; Lee & Anderson, 1998). Variants of the type strain differing in electrophoretic mobility and buoyant density have also been described (Magdoff-Fairchild, 1967).

Transmission by Vectors

Transmissible by leaf beetles (Chrysomelidae), probably in a non-circulative manner (Walters, 1969; Fulton et al., 1975) although the virus may be found in the vector's haemolymph (Wang et al., 1992). In North America, transmitted by Ceratoma trifurcata and Epilachna varivestis (Fulton et al., 1975) and Diabrotica undecimpunctata howardii (Wang et al., 1994). The length of time that vectors remain viruliferous depends on the length of acquisition time and also on the length of time spent not feeding after acquisition (Wang et al., 1994). Adult Mexican bean beetles, Epilachna varivestis, preferred to feed on SBMV-infected bean plants rather than on non-infected plants (Musser et al., 2003).

The virus can apparently also be transmitted in the absence of vectors: Teakle (1986) reported that the virus can be acquired from soil in which a previously infected plant had grown.

Transmission through Seed

Seed-borne in bean (1-5%; Zaumeyer & Harter, 1943); evidence of embryo transmission in bean is equivocal (McDonald & Hamilton, 1972; Uyemoto & Grogan, 1977). Transmitted to bean seedlings if germinating seeds are in contact with infective extracts (Hamilton, 1978) or are planted in soil near roots of infected plants (Teakle & Morris, 1981). There is no evidence of pollen transmission to seed or to the pollinated plant but the pollen exine is contaminated (Hamilton et al., 1977).

Serology

Highly immunogenic; microprecipitin titres of 1/2048-1/4096 are commonly found. Immunodiffusion in gels, including the use of antisera to sodium dodecyl sulphate-treated virus (Purcifull et al., 1981), immunosorbent electron microscopy and ELISA are applicable for detecting virus in tissue extracts.

Relationships

The physical and chemical properties of SBMV particles are similar to those of other members of the Sobemovirus genus such as Turnip rosette virus, Blueberry shoestring virus, Cocksfoot mottle virus, Rice yellow mottle virus, Sowbane mosaic virus (Matthews, 1982), Lucerne transient streak virus (Forster & Jones, 1980) and SCpMV (Grogan & Kimble, 1964). SBMV is not serologically related to other members, except for a moderately distant relationship to SCpMV and a possible distant relationship of the Ivory Coast isolate to Sowbane mosaic virus (Givord, 1981). All strains of SBMV are serologically related to each other (Grogan & Kimble, 1964; R. I. Hamilton, unpublished results).

Stability in Sap

In bean sap: infectivity dilution end-point, 10-5-10-8; thermal inactivation point (10 min), 90-95 °C; longevity (18-22 °C), 20-165 days (Zaumeyer & Harter, 1942; Yerkes & Patino, 1960; Givord, 1981).

Purification

Easily purified in high yield (1 mg/g infected fresh leaf tissue); yields obtained from tissue frozen at -10 °C for longer than 1 month are much lower.

Method 1. (Ghabrial et al., 1967, as modified for SCpMV by Johnson et al., 1974). Extract the infected tissue 3 weeks after inoculation in 0.5 M sodium citrate (pH 7.5) containing 0.1% 2-mercaptoethanol and clarify the extract by low speed centrifugation. Adjust the supernatant fluid to pH 4.5 with 10% acetic acid and after 1 h centrifuge the material at low speed. Adjust the supernatant fluid to pH 6 with 10% NaOH and precipitate the virus by adding polyethylene glycol, mol. wt 6000 (PEG), to 8% (w/v) and NaCl to 0.1% (w/v). Give the preparation four to five cycles of differential centrifugation and use 0.05 M phosphate (pH 7.0) as a suspending medium. Adjust the preparation to pH 4.5 between the second and third cycles.

Method 2. (Tremaine et al., 1981). Homogenize freshly harvested infected leaves in 0.2 M sodium acetate buffer (pH 5.0) containing 0.02 M sodium diethyldithiocarbamate and 0.1% 2-mercaptoethanol. Adjust the extract to pH 4.8 with 10% acetic acid, and leave at 5 °C for 4 h. Clarify the extract by low speed centrifugation and precipitate the virus with PEG at 8% (w/v). Suspend the pellet from low speed centrifugation in 0.1 M sodium acetate buffer (pH 5.0) and give the preparation two cycles of differential centrifugation. High speed pellets turn white and become insoluble when suspended at pH 5.0 but are very soluble when suspended in 0.1 M sodium phosphate buffer (pH 7.0). Particles suspended at pH 7.0 are stable and soluble after dialysis or adjustment to pH 5.0.

The virus can also be purified with the aid of organic solvents (Hull, 1977a). The type strain crystallizes on dialysis against distilled water (Miller & Price, 1946b) or in solutions of MgSO4 or (NH4)2SO4 (Price, 1946).

Properties of Particles

The virus particles are stable between pH 2.5 and 9.5 (Sehgal, 1980). The particles swell on treatment with EDTA under slightly alkaline conditions (Wells & Sisler, 1969) and dissociate into RNA and protein with the further addition of 1 M NaCl (Hull, 1977b). The particles are stabilized by divalent ion-dependent and pH-dependent protein-protein interactions and by RNA-protein interactions (Hull, 1977b). Stable virus particles can be reconstituted from type strain protein and either type strain RNA or Sowbane mosaic virus RNA (Tremaine & Ronald, 1977).

Sedimentation coefficient: Unswollen particles sediment as a single component with s20,w = 115 S at infinite dilution; swollen particles s20,w = 73 - 100 S (Wells & Sisler, 1969; Hsu et al., 1976; Hull, 1977b).

Particle weight (daltons): 6.6 × 106 (Miller & Price, 1946a); 6.1 × 106 (Lauffer et al., 1952); 6.5 × 106 (Yphantis, 1964).

Diffusion coefficient (D20,w × 10-7cm2/sec): 1.34 (Miller & Price, 1946a).

Partial specific volume: 0.696-0.700 (Miller & Price, 1946a; Lauffer et al., 1952).

Electrophoretic mobility: mobility varies with the strain (Hartmann & Lauffer, 1953; Magdoff-Fairchild, 1967; Tremaine & Wright, 1967).

Absorption coefficient A260 (0.1%, 1 cm): 5.85.

A260/A280: 1.6.

Buoyant density (g/cm3): 1.360 in CsCl; 1.402 in CsBr; 1.282 in metrizamide; two banding zones form in Cs2SO4 gradients, a single band in the heavy zone (1.32) and one to four bands in the light zone (1.28-1.305) (Hull, 1977a); 1.26 in sucrose (Lauffer et al., 1952).

Particle Structure

The particles are isometric, approximately 30 nm in diameter (Fig. 3) and have a T = 3 structure with 180 protein subunits. Negatively stained particles do not show distinct capsomeric structures in the electron microscope because the high protein density at the surface of the particle masks the differentiation of morphological features. Pentamer-hexamer clustering and an additional trimer clustering has been detected in the serologically related SCpMV by X-ray diffraction at 22.5 Å resolution (Johnson et al., 1976). Higher resolution X-ray diffraction studies at 2.8 Å, also on SCpMV (Abad-Zapatero et al., 1980), showed the protein is an eight-stranded, antiparallel beta barrel with the N-terminus forming an additional partially ordered arm in the interior of the virus particle. There are three conformations of the protein subunit (A, B, and C) within one icosahedral asymmetric unit. The exact three-dimensional positioning of residues 42 to 260 of the C subunits of SCpMV are known, including those residues in contact with the RNA, with adjacent subunits or with the exterior (Hermodson et al., 1982). The structure of the protein subunit of SBMV is very similar to that of Tomato bushy stunt virus (Harrison et al., 1978) and Satellite tobacco necrosis virus (Liljas et al., 1982).

Particle Composition

Nucleic acid:
Positive-sense ssRNA, comprising about 21% of the particle weight (Miller & Price, 1946a). RNA is best prepared from particles dissociated in SDS, EDTA and Tris buffer (pH 8) and extracted by adding two vol of a 1:1 vol/vol mixture of chloroform and phenol (Salerno-Rife et al., 1980). The genome is contained in a single ssRNA molecule of Mr 1.4 × 106 and sedimentation coefficient of 24.9 S, becoming 14.9 S after treatment with formaldehyde (Diener, 1965; Kaper & Waterworth, 1973). The sequence has been determined for the type strain (accession number L34672; Othman & Hull, 1995) and comprises 4109 nucleotides. A value of 4136 nucleotides was determined for the Arkansas strain (accession number AF055887) and a resistance-breaking strain (accession number AF055888) (Lee & Anderson, 1998). The molar percentage of nucleotides is G 25.7; A 23.8; C 24.5; U 26.0. RNA preparations also contain one subgenomic RNA component as well as heterogeneous RNA (Rutgers et al., 1980; Ghosh et al., 1981).

Protein:
Coat protein can be isolated by first swelling the virus (Hsu et al., 1976) at 10-20 mg/ml in 0.1 M sodium phosphate buffer, 0.01 M EDTA (pH. 8.0) for at least 1 h at 0 °C and then adding an equal volume of 4 M LiCl and freezing overnight at -20 °C. After thawing, the RNA precipitate is removed by low speed centrifugation and the protein-containing supernatant fluid dialysed against 0.01 M phosphate buffer (pH 7.5) containing 0.01 M EDTA to remove LiCl.

The particles contain a single protein species of Mr 28,000 and comprising 79% of the particle weight; the amino acid composition differs for the B and M strains (Tremaine, 1966; Ghabrial et al., 1967).

Genome Properties

The single RNA species has a covalently linked 5'-terminal protein, which has Mr of 12,300 and is required for infectivity (Veerisetty & Sehgal, 1980; Mang et al., 1982). No polyadenylated terminal region or tRNA like structure has been detected (Ghosh et al., 1979).

The nucleotide sequence obtained by Lee & Anderson (1998) for two Arkansas isolates of SBMV contains four possible open reading frames (ORFs) (Fig. 4). ORF1 has a coding capacity for a protein of Mr 17,200, which, by analogy with other sobemoviruses, is the cell-to-cell movement protein (Sivakumaran et al., 1998) and is also the suppressor of RNA silencing (Voinnet et al., 1999). ORF2 encodes a polyprotein of Mr 106,500, which contains sequence motifs for a serine proteinase, the genome-linked protein and for the RNA-dependent RNA polymerase and is processed to give the genome-linked protein (Van der Wilk et al., 1998). ORF3 is located within ORF2, and encodes a protein of Mr about 14,900, which is of unknown function. The ORF4 product, Mr 28,800, is the coat protein. The nucleotide sequence obtained for the type strain of SBMV by Othman & Hull (1995) is 91.3% identical to that of Lee & Anderson (1998), but single nucleotide deletions at several positions in the genome resulted in considerable losses of amino acid sequence identity. As a result, this sequence does not have an equivalent to ORF3, and the products of the equivalents of ORFs 1, 2 and 4 have predicted Mr of 11,684, 96,481 and 28,107 respectively.

The nucleotide sequence of ORF4 of SBMV has considerable homology with that of SCpMV which is reflected in the similarities in particle structure. The sequence of the rest of the SBMV genome differs significantly from that of SCpMV which justifies them being separate species (Othman & Hull, 1995).

Translation of unfractionated RNA in the wheat embryo or rabbit reticulocyte systems results in two related products, of Mr 105,000 and 75,000, and two distinctive products, the coat protein and a protein of Mr 14,000 not related to the genome-linked protein (Salerno-Rife et al., 1980). Fractionation of the RNA by density gradient centrifugation yields the full length RNA, a 0.7 to 0.9 × 106 component and a 0.3 to 0.4 × 106 component, as well as heterogeneous populations of intermediate and smaller size (Rutgers et al., 1980; Mang et al., 1982). By analogy with SCpMV (Hacker & Sivakumaran, 1997), there is probably one subgenomic RNA from which the coat protein is expressed, whereas ORFs 1 and 2 are expressed from the genomic RNA; expression of ORF 3 has not been demonstrated.

Relations with Cells and Tissues

Virus particles are found in cytoplasm and nuclei of infected mesophyll cells (De Zoeten & Gaard, 1969); crystallization of virus particles in mesophyll cells is rare in bean (Weintraub & Ragetli, 1970) (Fig. 5). Mitochondria may contain dense bodies, and extrusions of chloroplasts may also occur (Weintraub & Ragetli, 1970). Removal of ribosomes by in situ RNAse treatment (Hatta & Francki, 1981) clearly revealed randomly distributed virus particles in the cytoplasm. Particles may also be found in the phloem (Fig. 6).

Ecology and Control

The virus is usually introduced into the field through infected seed and then transmitted to other plants by beetles; some soil transmission may occur to healthy plants grown in soil from which infected plants had been recently removed. Control is by insecticidal treatment against the beetle vector and by the use of resistant varieties; however, there are some resistance-breaking strains of the virus.

Notes

The virus can be distinguished from other legume-infecting viruses by its morphology, sedimentation rate, serology and narrow host range. It is distinguished from SCpMV by its inability to cause systemic infection of cowpea.

Figures

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Local lesions on the primary leaf of Phaseolus vulgaris (cv. Pinto) inoculated with the type strain.

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Systemic mosaic induced in leaf of P. vulgaris (cv. Black Turtle Soup) infected with strain M.

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Virus particles from a purified preparation negatively stained in 2% uranyl acetate. Bar represents 100 nm.

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Genome organization of SBMV. The complete line represents the RNA genome with the 5' and 3' ends indicated; the scale at the bottom is of nucleotides. The open reading frames (ORFs) are shown by boxes with the positions of three functional domains, serine protease, genome-linked protein (VPg) and RNA-dependent RNA polymerase (RdRp) in ORF2 being indicated; ORF4 encodes the coat protein.

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Section of P. vulgaris mesophyll leaf cell infected with SBMV showing individual and crystalline arrays of particles in the cytoplasm and nucleus (arrows). Bar represents 1 µm. (From Weintraub & Ragetli, 1970).

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Section of P. vulgaris phloem cell 16 days after infection with SBMV showing crystalline arrays of particles. Bar represents 1 µm. (From Weintraub & Ragetli, 1970).

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