|Family: Unallocated ssRNA+ viruses|
Species: Southern bean mosaic virus
|This is a revised version of DPV 274|
Department of Disease and Stress Biology, John Innes Centre, Norwich Research Park, Colney, Norwich NR4 7UH, UK
Host Range and Symptomatology
Transmission by Vectors
Transmission through Seed
Transmission by Grafting
Transmission by Dodder
Nucleic Acid Hybridization
Stability in Sap
Properties of Particles
Properties of Infective Nucleic Acid
Relations with Cells and Tissues
Ecology and Control
Described by Zaumeyer & Harter, 1942; 1943.
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.
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.
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).
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.
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).
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).
Diffusion coefficient (D20,w × 10-7cm2/sec): 1.34 (Miller & Price, 1946a).
Absorption coefficient A260 (0.1%, 1 cm): 5.85.
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).
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).
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).
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.
Local lesions on the primary leaf of Phaseolus vulgaris (cv. Pinto) inoculated with the type strain.
Systemic mosaic induced in leaf of P. vulgaris (cv. Black Turtle Soup) infected with strain M.
Virus particles from a purified preparation negatively stained in 2% uranyl acetate. Bar represents 100 nm.
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.
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).
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).