Carrot red leaf virus
P. M. Waterhouse
Scottish Crop Research Institute, Invergowrie, Dundee, Scotland
A. F. Murant
Scottish Crop Research Institute, Invergowrie, Dundee, Scotland
Described by Stubbs (1948
), Watson, Serjeant & Lennon
and Waterhouse & Murant (1981a
A virus with RNA-containing isometric particles c. 25 nm
in diameter, transmitted by the aphid Cavariella aegopodii in the persistent
(circulative) manner but not by inoculation with sap. It is apparently confined to
phloem tissue, and infects only species in the family Umbelliferae. Distributed
throughout the world, frequently in association with carrot mottle virus, which is
dependent on it for transmission by aphids.
In carrot (Daucus carota
), the virus causes reddening and
yellowing of leaves and some stunting of the plants. In nature, it
commonly occurs in association with carrot mottle virus
(Watson et al., 1964
; Murant, 1974
) to cause the disease
known as motley dwarf, in which the reddening, yellowing and
stunting is accentuated (see Fig.1 of Murant, 1974
) and there
is a variable amount of mottling. The motley dwarf disease was
first described and named by Stubbs (1948
), but the existence
of two virus components was not then recognized. The virus complex
caused serious losses in spring-sown carrots and in carrot seed crops
in Australia in the 1930s and 1940s (Stubbs, 1948
). It has also caused
serious yield losses in some years in Britain (Watson, 1960
; Watson &
) and in the USA (Krass & Schlegel, 1974
). A similar
disease complex exists in parsley (Petroselinum crispum
Britain (Frowd & Tomlinson, 1972
) but the relationship of the
causal viruses to carrot red leaf and carrot mottle
viruses remains to
be established (see below, under Strains).
Carrot motley dwarf disease is reported from Australia (Stubbs,
), USA (Stubbs, 1956
; Howell & Mink, 1974
; Krass &
), Japan (Komuro & Yamashita, 1956
), New Zealand
), Europe (Watson, 1960
; Heinze, 1968
) and Canada
). Both the carrot red leaf and carrot mottle virus
components are presumably present in all these countries.
Host Range and Symptomatology
Occurs naturally in cultivated and wild carrot and is also
reported from parsley (though see below under Strains), cow parsley
), and hogweed (Heracleum
). Experimentally, it is restricted to a few species
of Umbelliferae (Watson et al., 1964
; P. M. Waterhouse &
A. F. Murant, unpublished data), in which it typically induces stunting,
reddening and yellowing. Transmitted by aphids but not by inoculation
with sap. The following are useful test species.
- Diagnostic species
- Anthriscus cerefolium (chervil). Mild to severe stunting,
and curling of leaves, 10-16 days after inoculation (Fig.1). Similar
symptoms occur when carrot mottle virus is also present, but the
stunting may be more severe, the leaflets being slightly more reduced
in size than with carrot red leaf virus alone (Fig.1).
- Apium leptophyllum, syn. A. ammi (slender celery).
Severe stunting and reddening, visible 7-14 days after inoculation.
Similar symptoms occur when carrot mottle virus is also present.
- Coriandrum sativum (coriander). Severe stunting, bright
yellowing and reddening of the whole plant, visible 12-16 days after
inoculation (Fig.2). Similar symptoms occur when
carrot mottle virus
is also present.
- Apium graveolens (celery) and Petroselinum crispum
(parsley) are immune to most isolates (but see below under Strains);
they can, however, be infected (symptomlessly) by carrot mottle virus.
- Propagation species
- Daucus carota and Anthriscus cerefolium are
convenient for the maintenance of cultures; Anthriscus
cerefolium is used as a source of virus for purification.
- Assay species
- Anthriscus cerefolium, Apium leptophyllum and Coriandrum
sativum are used in tests for transmission by aphids.
Few isolates have been compared serologically and most isolates
described from carrot have similar host ranges. However, Frowd &
described an isolate from parsley, a species which
is immune to carrot isolates, and Ohki, Doi & Yora (1979
described an isolate from carrot in Japan which was unusual in
infecting celery (Apium graveolens
) and Cryptotaenia
. Although these isolates may well be host range variants
of carrot red leaf virus, there is as yet no serological evidence that
they are related to it.
Transmission by Vectors
Transmitted in the persistent (circulative) manner by the
carrot-willow aphid, Cavariella aegopodii
, but not by eight
other species tested: Anuraphis tulipae, Acyrthosiphon pisum,
Cavariella pastinacae, C. theobaldi, Hyadaphis foeniculi, Myzus
and two unidentified species of Macrosiphum
; Watson et al., 1964
; Murant, 1974
). The carrot virus from Japan (Ohki et al., 1979
transmitted both by Cavariella aegopodii
and by Semiaphis
In tests with C. aegopodii, the minimum acquisition access
period was about 30 min and the minimum inoculation access period
about 2 min (Elnagar & Murant, 1978a); the minimum latent
period was between 6 and 18 h (Watson et al., 1964; Elnagar
& Murant, 1978a). Aphids retained the ability to transmit
the virus after moulting (Stubbs, 1955; Watson et al., 1964;
Elnagar & Murant, 1978a) and continued to transmit it after
at least 12 serial daily transfers (Elnagar & Murant, 1978a).
The virus was not transmitted to the progeny of viruliferous aphids
and there is no evidence for multiplication of the virus in the vector
(Elnagar & Murant, 1978a, 1978b). Aphids injected with extracts
of viruliferous donor aphids transmitted carrot red leaf virus readily
to test plants (Elnagar & Murant, 1978b).
Carrot red leaf virus acts as a helper for transmission of carrot
mottle virus by aphids (Watson et al., 1964). In mixedly
infected plants, some carrot mottle virus nucleic acid becomes
packaged in coat protein of carrot red leaf virus (Waterhouse, 1981;
Waterhouse & Murant, 1981b); these particles are transmitted
by C. aegopodii in the same way as carrot red leaf virus
particles and have the same vector relations (Elnagar & Murant,
Transmission through Seed
Not seed-transmitted in carrot (Stubbs, 1948
; Krass &
; A. F. Murant, unpublished data) or in Anthriscus
A rabbit injected intradermally with 100 µg purified virus
yielded antiserum with a titre of 1/512 in double diffusion tests in
agarose gel (Waterhouse & Murant, 1981a
). A single line of
precipitate is produced in such tests but they must be done with
partially purified virus because the concentration of virus in plant
extracts is too low. However, the virus can be detected in
unconcentrated extracts from plants or groups of aphids by using
enzyme-linked immunosorbent assay (ELISA) or immunosorbent electron
microscopy (ISEM) (Waterhouse & Murant, 1981a
of antigen-antibody reactions by sucrose density gradient
centrifugation is a useful technique for determining serological
relationships (Waterhouse & Murant, 1981a
The properties of carrot red leaf virus (induction of reddening
and yellowing symptoms, restriction to phloem tissue,
non-transmissibility by mechanical inoculation, high vector
specificity, persistent (circulative) relation with the aphid
vector and possession of 25 nm isometric particles) place it in
the luteovirus group
. It is distantly serologically related to
each of seven luteoviruses tested (Waterhouse & Murant,
), most closely to barley yellow dwarf virus
and perhaps beet western yellows virus
, more distantly to tobacco
, potato leafroll
and bean leafroll
viruses, and very
distantly to barley yellow dwarf virus
(MAV strain) and soybean
Stability in Sap
Infectivity of purified preparations survived after 1 month
at -15°C (Waterhouse, 1981
The following method is useful (Waterhouse & Murant,
). Homogenise infected chervil plants in 0.1 M sodium
citrate buffer, pH 6.0, containing 1% (w/v) thioglycerol
(3 ml/g tissue). Centrifuge the resultant slurry at 3000
for 15 min and discard the supernatant fluid.
Resuspend the pellets, mainly fibrous material, in extraction
buffer containing 0.02% (w/v) sodium azide and 1.5% (w/v)
Driselase (a product that contains pectinase and cellulase enzymes);
then shake the suspension at 28°C for 16-20 h. After incubation
add Triton X-100 to 1% (v/v) and stir at room temperature for 30 min.
Subject the suspension to two cycles of differential centrifugation:
in each cycle, first centrifuge the preparation at 8000
for 15 min and then float the supernatant fluid
on a layer of 20% (w/v) sucrose occupying about one quarter of the
tube, and centrifuge for 3.5 h at 140,000 g
Resuspend the pellets from high speed centrifugation in 0.006 M
phosphate buffer, pH 7.0. The virus may be further purified by
sucrose density gradient centrifugation. About 1 mg virus may be
obtained from 1 kg leaf material.
Properties of Particles
The particles form a single sedimenting and buoyant density
Sedimentation coefficient (s20,w): 104 S.
Buoyant density in CsCl: 1.403 g/cm3.
(values not corrected for light-scattering).
Absorption coefficient (A260,0.1%,1 cm):
c. 7 .0, estimated from the A260/A280
ratio and buoyant density.
The particles are isometric, c.
25 nm in diameter,
many appearing hexagonal in outline (Fig.3
Particle CompositionNucleic acid:
RNA, single-stranded, about 28% of particle
weight, estimated from the buoyant density of the particles in CsCl.
Electrophoresis in polyacrylamide gels in non-denaturing conditions
revealed only one species, of M. Wt c.
1.8 x 106
Protein: Protein of M. Wt c. 25,000 was observed
by electrophoresis in polyacrylamide/SDS gels. Three minor
slower-migrating components were found but their origin is unknown
Relations with Cells and Tissues
The virus particles are confined to the phloem tissue; they
occur especially in companion cells (Fig.5
) and sieve tubes, but
also in phloem parenchyma cells (Murant & Roberts, 1979
were found in the cytoplasm and vacuoles but not in the nuclei of
infected cells. A similar tissue distribution was reported for the
Japanese carrot virus studied by Ohki et al. (1979)
also observed cytoplasmic vesicular structures containing nucleic
Carrot red leaf virus is the only definitive luteovirus known to
infect species of Umbelliferae. The symptoms it causes in carrot,
whether alone or in association with carrot mottle virus
, are not
easily confused with those caused by any other virus but may be
mistaken for those of a deficiency disorder (Stubbs, 1948
with those caused by carrot fly (Psila rosae
treatments applied against carrot fly are also effective against
. Carrot red leaf and carrot mottle
viruses are therefore now uncommon in areas where routine treatments
against carrot fly are employed. The viruses do not infect willow,
the winter host of C. aegopodii
, but survive between crops
in weeds such as cow parsley or wild carrot, in volunteer carrots
or in overwintering carrot seed crops (Watson & Serjeant, 1964
Howell & Mink, 1977a
). The virus may also be carried
in roots transplanted into new areas for seed production (Stubbs,
; Howell & Mink, 1976
). The epidemiology of motley
dwarf disease was investigated by Watson & Serjeant (1964)
and Howell & Mink (1977a
- Anon., Rep. Dep. scient. ind. Res. N.Z. 1959, 112 pp., 1959.
- Elnagar & Murant, Ann. appl. Biol. 89: 237, 1978a.
- Elnagar & Murant, Ann. appl. Biol. 89: 245, 1978b.
- Frowd & Tomlinson, Ann. appl. Biol. 72: 177, 1972.
- Heinze, Z. PflKrankh. Pflpath. PflSchutz 75: 513, 1968.
- Horne & Ronchetti, J. Ultrastruct. Res. 47: 361, 1974.
- Howell & Mink, Pl. Dis. Reptr. 58: 766, 1974.
- Howell & Mink, Pl. Dis. Reptr 60: 1047, 1976.
- Howell & Mink, Pl. Dis. Reptr 61: 217, 1977a.
- Howell & Mink, Pl. Dis. Reptr 61: 841, 1977b.
- Komuro & Yamashita, Ann. phytopath. Soc. Japan 20: 155, 1956.
- Krass & Schlegel, Phytopathology 64: 151, 1974.
- Murant, CMI/AAB Descr. Pl. Viruses 137, 4 pp., 1974.
- Murant, Can. Pl. Dis. Surv. 55: 103, 1975.
- Murant & Roberts, Ann. appl. Biol. 92: 343, 1979.
- Ohki, Doi & Yora, Ann. phytopath. Soc. Japan 45: 74, 1979.
- Stubbs, Aust. J. sci. Res. B 1: 303, 1948.
- Stubbs, Aust. J. sci. Res. B 5: 399, 1952.
- Stubbs, J. Aust. Inst. agric. Sci 21: 267, 1955.
- Stubbs, Pl. Dis. Reptr 40: 763, 1956.
- Waterhouse, Ph.D. Thesis, University of Dundee, 244 pp., 1981.
- Waterhouse & Murant, Ann. appl. Biol. 97: 191, 1981a.
- Waterhouse & Murant, Abstr. 5th int. Congr. Virol., Strasbourg, France, 1981: 212, 1981b.
- Watson, Pl. Path. 9: 133, 1960.
- Watson & Serjeant, Ann. appl. Biol. 53: 77, 1964.
- Watson, Serjeant & Lennon, Ann. appl. Biol. 54: 153, 1964.
Chervil, Anthriscus cerefolium: (left) healthy,
(upper right) infected with carrot red leaf virus,
(lower right) infected with both carrot red leaf and
carrot mottle viruses.
Coriander, Coriandrum sativum: (left) healthy,
(right) infected with carrot red leaf virus.
Particles of carrot red leaf virus. Prepared by the
method of Horne & Ronchetti (1974) with 2% ammonium molybdate,
pH 6.5, and 2% uranyl acetate. Bar represents 100 nm.
(From Waterhouse & Murant, 1981a).
Schlieren patterns: (above) moving boundary
sedimentation in 0.006 M phosphate buffer, pH 7.0, at 20°C.
Photograph taken 8 min after a speed of 30,000 rev./min was reached;
(below) equilibrium banding after centrifugation for
16 h at 20°C at 44,000 rev./min in a CsCl solution of
initial density 1.44 g/cm3. (From Waterhouse
& Murant, 1981a).
Particles of carrot red leaf virus in a companion
cell from a thin section of an infected chervil leaf. The
virus particles are readily distinguished from the ribosomes
in the adjacent parenchyma cell which contains no virus-like
particles. (From Murant & Roberts, 1979). Bar
represents 500 nm.