49
June 1971
Family: Bromoviridae
Genus: Bromovirus
Species: Cowpea chlorotic mottle virus
Acronym: CCMV


Cowpea chlorotic mottle virus

J. B. Bancroft
John Innes Institute, Norwich, Norfolk, England

Contents

Introduction
Main Diseases
Geographical Distribution
Host Range and Symptomatology
Strains
Transmission by Vectors
Transmission through Seed
Transmission by Grafting
Transmission by Dodder
Serology
Nucleic Acid Hybridization
Relationships
Stability in Sap
Purification
Properties of Particles
Particle Structure
Particle Composition
Properties of Infective Nucleic Acid
Molecular Structure
Genome Properties
Satellites
Relations with Cells and Tissues
Ecology and Control
Notes
References
Acknowledgements
Figures

Introduction

Described by Kuhn (1964b).

Selected synonyms

None

An RNA-containing virus with isometric particles about 25 nm in diameter. It is readily transmissible by inoculation with sap and infects mainly legumes. It is found in temperate regions and is transmitted by beetles.

Main Diseases

Causes a mosaic disease of cowpea (Kuhn, 1964b) and has also been obtained from mottled soybeans (Kuhn, 1968).

Geographical Distribution

Reported from the USA only.

Host Range and Symptomatology

Readily transmissible by inoculation of sap. Confined largely to the Leguminosae; limited numbers of species in the Cucurbitaceae, Solanaceae and Chenopodiaceae have also been infected (Kuhn, 1964b; Walters & Dodd, 1969). Symptoms range from a mosaic to veinal necrosis and local lesions depending on the species.

Diagnostic species

Causes systemic mottle in cowpea (Vigna unguiculata) (Fig.1) and local lesions in certain varieties of soybean (Glycine max) such as Lindarin (Fig.2) and in Chenopodium hybridum (Fig.3) and C. album.

Propagation species

Vigna unguiculata.

Assay species

Glycine max, Chenopodium album and Chenopodium hybridum.

Strains

In addition to the type strain (Kuhn, 1964b), a soybean strain (Kuhn, 1968) and a Desmodium strain (Walters & Dodd, 1969) have been described. The latter two strains produce a less intense mottle on cowpea than does the type strain.

Transmission by Vectors

The bean leaf beetle (Ceratoma trifurcata) and the spotted cucumber beetle (Diabotrica undecimpunctata) transmit the Desmodium strain (Walters & Dodd, 1969) after 24 hr acquisition and feeding times, and probably the other strains as well.

Transmission through Seed

None detected in cowpea (Kuhn, 1964b).

Transmission by Dodder

No information.

Serology

The virus is moderately immunogenic (Kuhn, 1964b; Bancroft et al., 1968). Gel-diffusion tests are best done using gels buffered to pH 5-6.

Relationships

The three reported strains appear to be very similar and perhaps identical serologically.

Stability in Sap

In cowpea sap, the thermal inactivation point (10 min) of the type strain is between 65 and 70°C, the dilution end-point is between 1 x 10-4 and 1 x 10-5 and the longevity in vitro is between 24 and 48 hr (Kuhn, 1964b). In sap of infected Pinto bean the Desmodium strain appears to have similar thermal inactivation and dilution end-points, but has a reported longevity in vitro of between 29 and 44 days (Walters & Dodd, 1969). Direct longevity comparisons between the two strains grown in the same host have not been made. Virus in shredded desiccated cowpea tissue loses infectivity in about 3 weeks (Kuhn, 1964b).

Purification

The virus occurs in concentrations of about 300 mg/kg of cowpea tissue. Tissue, collected about 2 weeks after inoculation, is blended in 0.2 M pH 4.5 acetate buffer, the sap is expressed through cheesecloth, allowed to stand for 3-16 hr at 4°C, and the virus is further purified and concentrated by 2 or 3 cycles of differential ultracentrifugation (Bancroft, Hills & Markham, 1967). Pellets are resuspended in 0.1 M pH 5.0 acetate buffer and virus is stored at 4°C.

Properties of Particles

Sedimentation coefficient (s20, w): (88.3-0.83c) S at pH 3-6 and (77.8-1.27c) S at pH 7 (Bancroft et al., 1968) where c = nucleoprotein concentration in mg/ml. The virus sediments as a single component at pH 3-6 but may degrade and sediment as two or more components at pH 7 or above depending on dialysis time and exact ionic conditions. The virus is inactivated at pH 7 in the absence of divalent cations.

Molecular weight (daltons): 4.6 x 106 (Bancroft et al., 1968).

Isoelectric point: pH 3.6 at an ionic strength of 0.1.

Electrophoretic mobility: -6 x 10-5 cm2 sec-1 volt-1 at pH 5 and ionic strength of 0.1.

Absorbance at 260 nm (1 mg/ml, 1 cm light path): 5.87 (uncorrected for light scattering).

A260/A280: 1.7.

Buoyant density: 1.382 g/ml in CsCl, in which some of the virus particles seem to aggregate in groups of four (Bancroft et al., 1968).

Particle Structure

Particles are isometric (Fig.4) about 25 nm in diameter in 1% pH 4.7 uranyl acetate, and are built of 180 structure units in hexamer-pentamer clusters (Bancroft et al., 1967). The stained particles have a small, sometimes noticeable, electron-dense central area about 5 to 7.5 nm in diameter.

Particle Composition

RNA: M. Wt is 1.1 x 106. Smaller RNA molecules, perhaps derived from the larger, and with M. Wt c. 0.3 x 106 and 0.7 x 106, occur together in some particles. Single-stranded. Molar percentages of nucleotides: G26; A25; C20; U28. RNA is 24% of particle weight. Sedimentation coefficients (s20, w) in 0.01 M KCl, 0.01 M tris pH 7.4, 5 x 10-4 M MgCl2 are 23.1, 18.2 and 12.6 S for the 3 major classes of RNA (Bancroft et al., 1968).

Protein: Subunits have M. Wt c. 19,400 and contain 181 amino acid residues (J. B. Bancroft, unpublished) having one less tryptophan residue than originally reported (Bancroft et al., 1968).

Relations with Cells and Tissues

The virus is found in the nucleus, cytoplasm and vacuoles of infected cells (Hills & Plaskitt, 1968). It can be obtained from the roots, leaves, sepals, pistils, stamens and petals, but not from the pollen, of infected plants (Gay, 1969). It can also be detected in the seed coats, but not in the cotyledons or embryos, of immature undried seeds (Gay, 1969).

Notes

The virus, which is apparently common in cowpea, at least in Georgia, USA, is physically similar, but serologically unrelated, to brome mosaic and broad bean mottle viruses, neither of which produces symptoms in cowpea. Among other serologically unrelated viruses occurring in cowpea, cowpea mosaic virus (Shepherd, 1964) has isometric nucleoprotein particles sedimenting at 91 and 112 S and does not produce local lesions in soybean or Chenopodium album; the cowpea strain of southern bean mosaic virus (Shepherd & Fulton, 1962) has isometric particles sedimenting at 115 S and retains infectivity in sap after heating for 10 min at 85°C; and cowpea aphid-borne mosaic virus (Anderson, 1959; Lovisolo & Conti, 1966) has filamentous particles about 750 nm long and resembles bean yellow mosaic virus. Kuhn (1964a) has presented a scheme for differentiating some of the viruses which produce mosaic in cowpea.

References

  1. Anderson, Phytopathology 49: 117, 1959.
  2. Bancroft, Hills & Markham, Virology 31: 354, 1967.
  3. Bancroft, Hiebert, Rees & Markham, Virology 34: 224, 1968.
  4. Gay, Phytopathology 59: 802, 1969.
  5. Hills & Plaskitt, J. Ultrastruct. Res. 25: 323, 1968.
  6. Kuhn, Phytopathology 54: 739, 1964a.
  7. Kuhn, Phytopathology 54: 853, 1964b.
  8. Kuhn, Phytopathology 58: 1441, 1968.
  9. Lovisolo & Conti, Neth. J. Pl. Path. 72: 265, 1966.
  10. Shepherd, Phytopathology 54: 466, 1964.
  11. Shepherd & Fulton, Phytopathology 52: 789, 1962.
  12. Walters & Dodd, Phytopathology 59: 1055, 1969.


Figure 1

Systemically infected leaf of cowpea (Vigna unguiculata).

Figure 2

Local lesions in soybean (Glycine max) cv. Lindarin.

Figure 3

Local lesions in Chenopodium hybridum.

Figure 4

Virus particles from a purified preparation in 1% pH 4.7 uranyl acetate. Bar represents 100 nm. Enlarged particle (inset) shows detailed structure.