82
June 1972
Family: Tombusviridae
Genus: Tombusvirus
Species: Cucumber necrosis virus
Acronym: CuNV


Cucumber necrosis virus

H. F. Dias
Research Station, Research Branch, Canada Department of Agriculture, Vineland Station, Ontario, Canada

C. D. McKeen
Research Station, Research Branch, Canada Department of Agriculture, Harrow, Ontario, Canada

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 McKeen (1959) and Dias & Doanne (1968).

An RNA-containing virus with isometric particles c. 31 nm in diameter. Soil-borne, transmitted by the chytrid fungus Olpidium cucurbitacearum and readily transmitted by mechanical inoculation to a wide range of plants. Found naturally only in greenhouse cucumbers.

Main Diseases

Causes necrotic spots, severe malformation of the leaves and stunting in greenhouse cucumbers.

Geographical Distribution

Canada.

Host Range and Symptomatology

Several species of Amaranthaceae, Chenopodiaceae, Compositae, Cucurbitaceae, Leguminosae and Solanaceae can be infected by mechanical inoculation of sap. Usually only inoculated leaves become infected and buff or necrotic local lesions appear 2-5 days after inoculation. The virus becomes systemic only in Cucumis sativus (cucumber) and, less frequently, in Vigna sinensis (cowpea) and Zinnia elegans; it does not become fully systemic in other Cucurbitaceae.

Diagnostic species

Cucumis sativus (cucumber). Necrotic lesions (Fig.3) develop in inoculated cotyledons in 3-4 days and enlarge to 3-5 mm; the cotyledons desiccate and die. Systemic symptoms may include: chlorotic or tan-coloured areas with pin-point necrotic centres which usually fall out leaving ‘shot-holes’ of various sizes (Fig.4); severely deformed leaves, sometimes with dark green enations; and small fruits, occasionally with a green mottle, developing on the stunted plants. Symptoms in summer are mild or indistinct.

Gomphrena globosa. Irregular greyish necrotic local lesions with reddish margins develop in 3-5 days (Fig.2).

Chenopodium amaranticolor. Local necrotic lesions (0.5-1 mm) develop in 2-5 days (Fig.1), the margins reddening as the leaves age. Heavily infected leaves desiccate and fall.

Nicotiana tabacum (tobacco). Greyish necrotic local spots up to 4 mm in diameter (Fig.5).

Propagation species

Cucumis sativus cv. Windermoor Wonder is a good host for maintaining cultures. Highly infectious inoculum is obtained from cotyledons 5-7 days after inoculation.

Assay species

Chenopodium amaranticolor and Gomphrena globosa are good local lesion hosts.

Strains

No strain differences reported.

Transmission by Vectors

Transmitted in soil by zoospores of the chytrid fungus Olpidium cucurbitacearum (Barr, 1968; Dias, 1970a). The virus is not carried internally by the resting spores of the fungus; virus-free zoospores are discharged both from resting spores and zoosporangia (Fig.6) formed in virus-infected roots. Virus released from roots adsorbs to zoospores after a 5-15 min contact period and enters roots at the same time as the zoospores (Dias, 1970b). The fungus appears to infect only species of the Cucurbitaceae. Olpidium brassicae, the vector of tobacco necrosis virus, does not transmit cucumber necrosis virus nor does O. cucurbitacearum transmit tobacco necrosis virus (Dias, 1970b). Attempts to transmit the virus by the melon aphid (Aphis gossypii) failed (McKeen, 1959).

Transmission through Seed

Not seed-transmitted in cucumber (McKeen, 1959).

Transmission by Dodder

Not reported.

Serology

Antisera with titres greater than 1/1000 are readily obtained by intramuscular injection. The virus forms a single precipitin band near the antigen well in gel-diffusion tests. This test reliably detects virus in crude sap.

Relationships

Cucumber necrosis virus is not serologically related to tobacco necrosis virus (Dias & Doanne, 1968) although both viruses have many similar properties. Cucumber necrosis virus should not be confused with viruses of the same name from the Netherlands (Van Koot & Van Dorst, 1955), Sweden (Rydén, 1966) and possibly USSR (Vovk & Nikiforova, 1961), which are strains of tobacco necrosis virus.

Stability in Sap

In tobacco sap, the thermal inactivation point (10 min) is 75-80°C, the dilution end-point is 10-4-10-5 and the virus retains infectivity for about 30 days at room temperature. Very little infectivity is lost after 1 year in frozen sap or in desiccated leaves (McKeen, 1959). The virus is precipitated by 40% saturated ammonium sulphate and the infectivity of clarified extracts is unaffected between pH 4 and 8 (Dias & Doanne, 1968).

Purification

The following method gives highly infectious preparations: blend 100 g of fresh or frozen leaves with 200 ml 0.67 M sodium phosphate buffer pH 7.0 and 50 ml 0.1 M ascorbic acid, filter the sap through gauze and centrifuge at low speed (12,500 g). Clarify by adding 1/10th vol. chloroform followed by acidification (pH 5.3), and differential centrifugation (Dias & Doanne, 1968), or by adding ammonium sulphate to 40% saturation and subjecting the precipitate to differential centrifugation. Additional purification can be obtained by rate zonal density gradient centrifugation.

Properties of Particles

Dias & Doanne (1968) reported one component in purified preparations analysed by rate-zonal centrifugation in sucrose density gradients. Tremaine (1970) detected an accessory component in crude preparations which sedimented at one-third the rate of the main component.

Sedimentation coefficient (s20, w): 133 S (Dias & Doanne, 1968).

Molecular weight: 8.6 x 106 (J. H. Tremaine, unpublished).

Diffusion cofficient: 1.20 x 10-7 cm2/sec (J. H. Tremaine, unpublished).

Isoelectric point: pH 3.9 (J. H. Tremaine, unpublished).

Particle Structure

Particles are isometric, c. 31 nm in diameter (Fig.7), apparently angular in outline.

Particle Composition

16% RNA and 84% protein (J. H. Tremaine, unpublished). Approximately 391 amino acid residues per subunit. Amino acid composition: ala 41; arg 17; asx 46; cys 0; glx 24; gly 32; his 3; ile 20; leu 33; lys 16; met 1; phe 20; pro 23; ser 32: thr 31: trp 7; tyr 12; val 33 (Tremaine & Goldsack, 1968).

Relations with Cells and Tissues

No information.

Notes

The properties and host range of cucumber necrosis virus and tobacco necrosis virus, especially the cucumber strains, are similar; therefore, serological tests are the only reliable way of identifying the viruses.

References

  1. Barr, Can. J. Bot. 46: 1087, 1968.
  2. Dias, Virology 40: 828, 1970a.
  3. Dias, Virology 42: 204, 1970b.
  4. Dias & Doanne, Can. J. Bot. 46: 47, 1968.
  5. McKeen, Can. J. Bot. 37: 913, 1959.
  6. Rydén,Meddn St. VäxtskAnst. 13: 289, 1966.
  7. Tremaine, Virology 42: 611, 1970.
  8. Tremaine & Goldsack,Virology 35: 227, 1968.
  9. Van Koot & Van Dorst, Tijdschr. PlZiekt. 61: 163, 1955.
  10. Vovk & Nikiforova,Dokl. Akad. Nauk SSSR 137: 462, 1961.


Figure 1

Inoculated leaf of Chenopodium amaranticolor, showing small necrotic lesions.

Figure 2

Necrotic local lesions in Gomphrena globosa.

Figure 3

Necrotic local lesions in Cucumis sativus cv. Windermoor Wonder.

Figure 4

Systemically infected cucumber leaf showing necrotic areas and ‘shot holes’.

Figure 5

Necrotic lesions in inoculated leaf of Nicotiana tabacum cv. Harrow Velvet.

Figure 6

Zoosporangia of Olpidium cucurbitacearum within the root cells of cucumber. Note the variation in shape; one zoosporangium has a typical single exit tube.

Figure 7

Electron micrograph of virus particles from a purified preparation, mounted in 2% phosphotungstate. Bar represents 100 nm.