Details of DPV and References

DPV NO: 12 June 1970

Family: Virgaviridae

Genus: Tobravirus

Species: Tobacco rattle virus | Acronym: TRV

There is a more recent description of this virus: DPV 346

Tobacco rattle virus

B. D. Harrison Scottish Horticultural Research Institute, Invergowrie, Dundee, Scotland

Contents

Introduction

Described by Quanjer (1943).

Selected synonyms

(Most of the names listed below refer to isolates that differ to some extent from the type culture.)

Aster ringspot virus (Rev. appl. Mycol. 33: 675)

Belladonna mosaic virus (Rev. appl. Mycol. 22: 451)

Potato corky ringspot virus (Rev. appl. Mycol. 26: 76; 37: 501)

Potato stem mottle virus (stengelbont virus) (Rev. appl. Mycol. 27: 33)

Ratel virus (Rev. appl. Mycol. 25: 83)

Tabakmauche Virus (Behrens, 1899 and Rev. appl. Mycol. 37: 402)

Tabak Streifen und Kräuselkrankheit Virus (Rev. appl. Mycol. 7: 546)

(For the major variant, pepper ringspot virus, see under Strains)

An RNA-containing virus with straight tubular particles of two predominant lengths, the longer c. 190 nm and the shorter 45-115 nm, depending on the isolate. Some isolates are readily transmitted by inoculation of sap but others are not. The virus has a wide host range and is transmitted by soil-inhabiting nematodes (Trichodorus spp.).

Main Diseases

Causes one type of spraing (‘corky ringspot’, Pfropfenbildung) (Fig .4), and stem-mottle, in potato, notched leaf in gladiolus, ringspot in aster and pepper, yellow blotch in sugar beet, and unnamed diseases in lettuce, hyacinth, narcissus, tulip and several other ornamental species.

Geographical Distribution

Europe, USA, Brazil, Japan.

Host Range and Symptomatology

The host range is very wide. More than 400 species in more than 50 dicotyledonous and monocotyledonous families can be infected; in many the virus does not become systemic (Schmelzer, 1957; Uschdraweit & Valentin, 1956).

Diagnostic species

Chenopodium amaranticolor. Necrotic local lesions (Fig. 2), some tending to spread; not systemic.

Cucumis sativus (cucumber). Chlorotic or necrotic local lesions; not systemic.

Nicotiana clevelandii. Inoculated leaves remain symptomless, or develop chlorotic or necrotic lesions. Systemically infected leaves develop few symptoms or variable amounts of necrotic flecking and distortion.

Phaseolus vulgaris (French bean). Pin-point, necrotic local lesions develop in 1-3 days (Fig. 5); not systemic.

Propagation species

Nicotiana clevelandii is the best host for maintaining cultures and as a source of virus for purification.

Assay species

Chenopodium amaranticolor and Phaseolus vulgaris are the most useful plants for local lesion assay. Nicotiana tabacum (tobacco, Fig. 1) and N. glutinosa (Fig. 3) can be used with isolates that produce local lesions in these species. N. tabacum and Cucumis sativus are useful bait plants for testing transmission by vectors.

Strains

Many strains are described but few are reliably distinguished by symptoms in test plants. Some of the best characterized are:

PRN (Cadman & Harrison, 1959; Harrison & Nixon, 1959). Originally obtained from potato in Scotland. Now used as the type strain. Short particles c. 75 nm long.

CAM (Harrison & Woods, 1966) = pepper ringspot virus (Kitajima & Costa, 1969). Originally obtained from Bidens pilosa in Brazil. Infects N. tabacum without producing necrotic local lesions. Short particles 52 nm long.

California strains (Semancik, 1966). Originally obtained from Capsicum frutescens in California. Short particles 45 and 90 or 110 nm long.

Oregon strains. Originally obtained from potato in Oregon. Separated and characterized by Lister & Bracker (1969). One variant produces yellow ringspots in N. clevelandii, N. glutinosa, etc. Short particles 48 and 81, 90 or 100 nm long.

Isolates unable to produce nucleoprotein particles (= NM = unstable isolates) can be obtained from any of the above strains by using inocula containing only 190 nm particles. They are poorly transmissible by mechanical inoculation unless inocula are made using phenol. They cause more necrosis in plants than do their parent cultures (Fig. 6) and are slow to become systemic (Cadman, 1962). Similar isolates are found in naturally infected plants.

Transmission by Vectors

At least 11 Trichodorus spp. (stubby root nematodes) are natural vectors in Europe or the USA but there is some evidence of specificity between virus strain and vector species (van Hoof, 1968; Taylor & Cadman, 1969). T. pachydermus and T. primitivus seem the most important vectors in Europe. Adults and larvae can transmit. The virus can be acquired by T. allius in 1 hr, inoculated in 1 hr and retained for 20 weeks by nematodes kept in soil without plants (Ayala & Allen, 1968).

Transmission through Seed

Occurs (1-6%) in some weeds, for example Capsella bursa-pastoris and Myosotis arvensis (Lister & Murant, 1967).

Transmission by Dodder

At least 6 Cuscuta spp. can transmit. The virus infects the dodder (Schmelzer, 1956).

Serology

Some strains are poorly immunogenic but antisera with titres of 1/1000 can be made against others. Precipitin tests in mixed liquids have been used most. One of two bands of precipitate develop in double diffusion tests in 1% agar gel.

Relationships

Antigenic differences are found between some isolates from W. Europe, USA and Japan, but isolates from Brazil differ more (Harrison & Woods, 1966). A distant serological relationship to pea early-browning virus was reported by Maat (1963).

Stability in Sap

In N. clevelandii sap, the thermal inactivation point (10 min) of several strains is 80-85°C, dilution end-point about 10^-6 and infectivity is retained at 20°C for more than 6 weeks. By contrast, RNA-producing isolates lose infectivity when sap is heated for 10 min at 60°C, diluted to 10^-2 or kept at 20°C for 3 h (Cadman, 1962).

Purification

Systemically infected N. clevelandii leaves yield 20-100 mg virus per kg leaf.

1. Store sap at -15°C. Thaw overnight at 20°C then purify by low- and high-speed centrifugation. Virus in pellets from the first high-speed centrifugation should be allowed to resuspend in 0.07 M phosphate buffer (pH 7.5) for 16 h at 5°C.

2. Grind cooled leaves in 0.01 M citric acid + phosphate buffer (pH 7.4, containing 0.1% sodium thioglycollate). Clarify extract by blending with 0.5 vol. of a 1:1 mixture of n-butanol + chloroform and freeze aqueous layer overnight. Thaw and purify virus by differential centrifugation, resuspending virus in 0.01 M phosphate buffer (pH 7.0) (Lister & Bracker, 1969).

Properties of Particles

All strains (except RNA-producing isolates) produce tubular particles of two or three predominant lengths, one c. 190 nm (L) and the other(s) 45-115 nm (S), depending on the isolate (Fig. 8). L particles are infective and induce synthesis of L-particle RNA. S particles are non-infective alone but carry the gene for the protein found in the virus particles. L and S are produced only when the inoculum contains both L and S or their RNAs (Lister, 1966; Frost, Harrison & Woods, 1967; Lister & Bracker, 1969; Sänger, 1969).

Sedimentation coefficient (s_{20, w}): c. 300 S (L), 155-243 S (S).

Molecular weight (daltons): c. 50 x 10⁶ (L), c. 12-29 x 10⁶ (S).

Electrophoretic mobility: -1.7 x 10^-5 cm² sec^-1 volt^-1 in 0.067 M phosphate buffer, pH 7.0 (L and S, PRN strain).

Absorbance at 260 nm (1 mg/ml, 1 cm light path): c. 3.0.

A₂₆₀/A₂₈₀: 1.15.

Ultraviolet radiation inactivates nucleoprotein particles irreversibly but some of the damage to extracted RNA is photoreactivable.

Particle Structure

Particles are helically constructed. The pitch of the helix is 2.5 nm and it has a central canal of diameter c. 5 nm. Number of protein subunits per turn = n + 1/3 (n =c. 25) (Nixon & Harrison, 1959; Offord, 1966).

Particle Composition

RNA: Single-stranded; molecular weight 2.3 x 10⁶ (L particles) and 0.6-1.3 x 10⁶ (S particles); about 5% of particle weight. Sedimentation coefficients are 26 S (L) and c. 12-20 S (S) in pH 7.4 buffer (0.01 M Tris + 0.01 M KCl + 10^-4 M MgCl₂). Molar percentages of nucleotides:

G 25; A 29; C 17; U 29 (California isolate) (Semancik & Kajiyama, 1967).

Protein: Subunits have a molecular weight of 2.4 x 10⁴ and contain about 218 amino acid residues (Offord & Harris, 1965). Amino acid composition of two California isolates is given by Semancik (1966).

Relations with Cells and Tissues

Most tissues of some species become infected. L particles of strain CAM become radially arranged around mitochondria (Fig. 7), whereas S particles are dispersed in the cytoplasm (Harrison & Roberts, 1968; Kitajima & Costa, 1969). Other strains behave differently (de Zoeten, 1966). In N. clevelandii, X-bodies largely composed of abnormal mitochondria can be induced by a RNA-producing isolate and by strain PRN (Harrison, Stefanac & Roberts, 1970).

Notes

Tobacco rattle virus mainly occurs on light sandy or peaty soils, which are the preferred habitat of its nematode vectors. It may be patchily distributed within a field. It can be distinguished from pea early-browning virus by failure to infect pea systemically, by having slightly shorter L particles and by serological tests. Also, most isolates of pea early-browning virus produce large lesions in Phaseolus vulgaris.

Acknowledgements

Photographs: courtesy of the Scottish Horticulltural Research Institute.

Figures

References list for DPV: Tobacco rattle virus (12)

1. Ayala & Allen, J. Agric. Univ. P. Rico 52: 101, 1968.
2. Behrens, Landwn Vers Stnen 52: 442, 1899.
3. Cadman, Nature, Lond. 193: 49, 1962.
4. Cadman & Harrison, Ann. appl. Biol. 47: 542, 1959.
5. de Zoeten, Phytopathology 56: 744, 1966.
6. Frost, Harrison & Woods, J. gen. Virol. 1: 57, 1967.
7. Harrison & Nixon, J. gen. Microbiol. 21: 569, 1959.
8. Harrison & Roberts, J. gen. Virol. 3: 121, 1968.
9. Harrison & Woods, Virology 28: 610, 1966.
10. Harrison, Stefanac & Roberts, J. gen. Virol. 6: 127, 1970.
11. Kitajima & Costa, J. gen. Virol. 4: 177, 1969.
12. Lister, Virology 28: 350, 1966.
13. Lister & Bracker, Virology 37: 262, 1969.
14. Lister & Murant, Ann. appl. Biol. 59: 49, 1967.
15. Maat, Neth. J. Pl. Path. 69: 287, 1963.
16. Nixon & Harrison, J. gen. Microbiol. 21: 582, 1959.
17. Offord, J. molec. Biol. 17: 370, 1966.
18. Offord & Harris, Fed. European Biochem. Soc. 2nd meeting, abstracts: 216, 1965.
19. Quanjer, Tijdschr. PlZiekt. 49: 37, 1943.
20. Sänger, J. Virol. 3: 304, 1969.
21. Schmelzer, Phytopath. Z. 28: 1, 1956.
22. Schmelzer, Phytopath. Z. 30: 281, 1957.
23. Semancik, Phytopathology 56: 1190, 1966.
24. Semancik & Kajiyama, Virology 33: 523, 1967.
25. Taylor & Cadman, in Viruses, Vectors and Vegetation, New York, Interscience Publishers, p. 55, 1969.
26. Uschdraweit & Valentin, NachrBl. dt. PflSchutzdienst, Braunschweig 8: 132, 1956.
27. van Hoof, Nematologica 14: 20, 1968.