Investigations of carnation viruses

Carnation vein mottle virus (CarVMV) is rare in glasshouse carnations in Britain, although locally common in Dianthus barbatus in private gardens. In Sim carnations free from other viruses, CarVMV caused slight diffuse chlorotic mottling in the younger leaves, decreased flower yield by c. 22%, and caused flower breaking in cvs William Sim and Dusty. In non-Sim cultivars Pink Shibiuya, Orchid Beauty and Vesta, leaf symptoms and flower breaking were more pronounced. In mixed infections with carnation mottle virus, symptoms were much more severe. CarVMV was not eliminated from carnation or D. barbatus plants grown for 4 wk at 37^oC, and only rarely from cuttings then taken from them, but it was readily eliminated by meristem-tip culture. Myzus persicae adults or nymphs acquired and transmitted the virus within a total time of 4 min, and remained infective for 30–60 min if feeding, or for 75 min if starved. The carnation aphid, M. persicae f. dianthi, transmitted the virus much less efficiently. The virus was not transmitted by dodder (Cuscuta campestris), or through seed of D. barbatus or Chenopodium quinoa. The maximum infective dilution in sap of D. barbatus, carnation and C. quinoa ranged from 10^-2 to 10^-5. The virus withstood 10 min at 60 but not 65^oC, up to 9 days at c. 18^oC or 3–4 wk at c. 2^oC. CarVMV infected twenty-two of 107 plant species in six of thirty-seven families; suscepts were confined to the Chenopodiaceae, Caryophyllaceae and closely allied families. C. quinoa was the best local lesion assay host. Seedling clones of D. barbatus, selected as resistant to carnation mottle virus, proved the best indicator and propagation species. Up to 50 mg virus/kg tissue were obtained by butanol clarification followed by differential and density gradient centrifugation. The preparations contained a single sedimenting component, s_20w= 144S, and had flexuous filamentous particles, c. 790 times 12 run; the particles contained a single polypeptide, mol. wt 34800, and 5% of a single-stranded ribonucleic acid (RNA) with nucleotide base ratios of G21: A25: C25: U29. Serologically CarVMV was related distantly to turnip mosaic (cabbage black ring strain), pea mosaic, watermelon mosaic (Strain 2) and bean yellow mosaic viruses, more closely to pepper veinal mottle virus, but unrelated to twelve other potyviruses. CarVMV is not at present a danger to carnation crops in Britain, but the recent trend of sending carnation plants to overwinter outdoors in warmer countries involves potential risks of more rapid spread by effective vector races of M. persicae.     

The elimination of four viruses from carnation and sweet william by meristem-tip culture

Carnation mottle virus (CarMV), carnation ringspot virus (CRSV), carnation vein mottle virus (CarVMV) and carnation latent virus (CarLV) were all eliminated from both carnation (Dianthus caryophyllus) and sweet william (D. barbatus) plants by meristem-tip culture. Carnation mottle virus was more readily eliminated from D. barbatus than from carnation. Carnation vein mottle and carnation latent viruses were more readily eliminated from carnation than from sweet william: they are rarely found in carnation but CarVMV is found frequently in sweet william. Carnation ringspot was eliminated equally readily from both hosts.     

Isolation of Carnation Ringspot Virus from a Canal Near a Sewage Plant: cDNA Hybridization Analysis, Serology and Cytopathology

A small water sample of only 60 ml from the Oker Aue canal near Braunschweig was ultracentrifuged. The resuspended pellet was rubbed on Chenopodium quinoa leaves which responded with the formation of almost 200 local lesions. The virus causing these lesions was identified as carnation ringspot virus (CarRSV) by means of dot blot hybridization using random-primed cDNAs to the viral nucleic acids and by means of serology. Northern blot analysis revealed that the two RNA species of the virus which consisted of c. 3.7 and 1.5 Kb, respectively, have little or no base sequence homology. In immunoelectrophoresis at pH 7.0 the virus migrated towards the cathode. The isometric particles were 33 nm in diameter, round to slightly angular in outline, and showed a distinct granular or knobly surface structure. Virus particles occurred in the cytoplasm, nuclei, and vacuoles of infected cells which in addition contained amorphous granules in the cytoplasm and/or proliferated endoplasmic reticulum. Heavily, affected cells were necrotized and contained large virus particle aggregates which sometimes were crystallized. CarRSV is the third carnation virus, after carnation mottle and carnation Italian ringspot viruses, which was identified in a natural water.     

The Effect of the Sequence of Infection of the Causal Agents of Sweet Potato Virus Disease on Symptom Severity and Individual Virus Titres in Sweet Potato cv. Beauregard

Sweet potato virus disease (SPVD) is caused by dual infection of plants with Sweet Potato Feathery Mottle Virus (SPFMV) and Sweet Potato Chlorotic Stunt Virus (SPCSV). Because SPFMV and SPCSV are transmitted by aphids and whiteflies, respectively, infection in nature occurs independently rather than simultaneously. To investigate the effect of consecutive infection on symptom development and individual virus titres, plants infected with a single virus were later inoculated with the second virus. Symptoms were significantly more severe in plants infected with SPCSV followed by SPFMV compared to plants infected with SPFMV followed by SPCSV. Virus titres were not significantly different for SPCSV, but SPFMV titres, in plants infected with SPCSV followed by SPFMV, were significantly higher than all other treatments. The results indicate that the sequence of infection of sweetpotato plants with the causal agents of SPVD influence the severity of symptoms and SPFMV titres in SPVD affected plants.     

Sweepoviruses cause disease in sweet potato and related Ipomoea spp.: fulfilling Koch's postulates for a divergent group in the genus begomovirus

Sweet potato (Ipomoea batatas) and related Ipomoea species are frequently infected by monopartite begomoviruses (genus Begomovirus, family Geminiviridae), known as sweepoviruses. Unlike other geminiviruses, the genomes of sweepoviruses have been recalcitrant to rendering infectious clones to date. Thus, Koch's postulates have not been fullfilled for any of the viruses in this group. Three novel species of sweepoviruses have recently been described in Spain: Sweet potato leaf curl Lanzarote virus (SPLCLaV), Sweet potato leaf curl Spain virus (SPLCSV) and Sweet potato leaf curl Canary virus (SPLCCaV). Here we describe the generation of the first infectious clone of an isolate (ES:MAL:BG30:06) of SPLCLaV. The clone consisted of a complete tandem dimeric viral genome in a binary vector. Successful infection by agroinoculation of several species of Ipomoea (including sweet potato) and Nicotiana benthamiana was confirmed by PCR, dot blot and Southern blot hybridization. Symptoms observed in infected plants consisted of leaf curl, yellowing, growth reduction and vein yellowing. Two varieties of sweet potato, 'Beauregard' and 'Promesa', were infected by agroinoculation, and symptoms of leaf curl and interveinal loss of purple colouration were observed, respectively. The virus present in agroinfected plants was readily transmitted by the whitefly Bemisia tabaci to I. setosa plants. The progeny virus population present in agroinfected I. setosa and sweet potato plants was isolated and identity to the original isolate was confirmed by sequencing. Therefore, Koch's postulates were fulfilled for the first time for a sweepovirus.     

Partial Characterization of Artichoke Virus M

A virus with filamentous particles 697 nm in length was isolated from artichoke plants in Southern Italy and identified as a new possible member of Carlavirus group, for which the name artichoke virus M (AVM) is suggested. AVM could not be transmitted by sap inoculation to herbaceous hosts and was always present in artichoke in mixed infections with other viruses. Virus particles had a buoyant density in CsCl of 1.31 g × cm^?3 and contained a single species of nucleic acid with an apparent size of 7.5 Kb and a single coat protein species with a mol. wt of 31,000. The virus was distantly related serologically to carnation latent and poplar mosaic carlaviruses but not to other members of the group including the recently described artichoke latent S carlavirus. Cytological alterations consisted of complex cytoplasmic inclusions composed of deranged organelles, lipid droplets and accumulations of membranes.     

Properties of a carlavirus causing a latent infection of pepino (Solanum muricatum)

Pepino (Solanum muricatum) cuttings imported from Chile contained a latent virus which was transmitted by inoculation of sap to Chenopodium quinoa but not to 21 other species. The virus was transmitted by the aphid, Myzus persicae. In C. quinoa sap, the virus lost infectivity when diluted between 10^-3 and 10^-4, heated for 10 min between 65 and 70 °C, or stored at room temperature for 4 to 6 days. The virus particles were straight or slightly flexuous filaments 660 to 680 nm long. Up to 15 mg virus per 100 g C. quinoa leaves was obtained by clarification with a mixture of chloroform and carbon tetrachloride. Purified preparations had A_max/A_min= 1.11, A₂₆₀/A₂₈₀= 1–30, A^0.₂₆₀^1%= 2.8, and contained a single sedimenting component with a sedimentation coeficient of 149s and a buoyant density in CsCl of 1–318. The virus particles contained 5.5% of single-stranded RNA of mol. wt 2.4×10⁶ (estimated by gel electrophoresis of undenatured RNA) and sedimentation coefficient 38.5S, and a single polypeptide of mol. wt 33 000. The virus is distantly serologically related to potato S and carnation latent viruses and is considered a new member of the carlavirus group. The name pepino latent virus is proposed. The cryptogram for this virus is R/1: 2.4/5–5: E/E: S/Ve/Ap.     

The occurrence, hosts and properties of lilac chlorotic leafspot virus, a newly recognised virus from Syringa vulgaris

Lilac chlorotic leafspot virus (LCLV), a hitherto undescribed virus, was isolated from three of 65 lilacs (Syringa vulgaris) with chlorotic leafspotting symptoms growing in S.E. England. The virus was transmitted readily by sap-inoculation to 21 of 52 species from eight of 20 families, but it was not seed-borne in four hosts or transmitted in the semi-persistent manner by any of four aphid species. The virus was moderately stable in vitro; sap from Chenopodium quinoa was infective after 10 min at 60 but not 65 ^oC, after 8–16 days at 20 ^oC or 25–30 wk at 2 ^oC, and after dilution to 10^-3 but not 10^-4. Up to 180 mg of purified virus per kg leaf tissue were obtained from C. quinoa by clarification of buffered leaf extracts with 8% (v/v) n-butanol, followed by one cycle of differential centrifugation and molecular permeation chromatography on controlled pore glass beads (700 Å, 120–200 mesh). LCLV has fragile flexuous filamentous particles which, when intact, mostly measured c. 12-5 times 1500–1600 nm; the helical substructure (pitch c. 3–7 nm) was clearly visible on some particles mounted in uranyl acetate. The particles sedimented as a single component (sedimentation coefficient 96 S; buoyant density 1–302 g cm^-3) and contained c. 5% nucleic acid and a single polypeptide of mol. wt 27 times 10³. Although these properties place LCLV in the closterovirus group, the virus showed no serological relationship to any of six closteroviruses (beet yellows, beet yellow stunt, carnation necrotic fleck, apple chlorotic leafspot, apple stem grooving and potato virus T) and differed from other recognised or possible members of this group in host range and/or symptoms induced in indicator species. The infrequent occurrence of LCLV in lilac in S.E. England indicates that the virus could probably be eradicated by selecting only virus-free plant material for propagation.     

THE TRANSMISSION BY MITES, HOST-RANGE AND PROPERTIES OF RYEGRASS MOSAIC VIRUS

A virus that causes chlorotic streaks on ryegrass leaves was transmitted by the eriophyid mite Abacarus hystrix (Nalepa). Virus-free mites acquired the virus in 2 hr. feeding on infected ryegrass and the proportion that became infective increased with increased feeding time up to 12 hr.; vectors lost infectivity within 24 hr. of leaving the infected leaves. All instars of A. hystrix transmitted the virus.
The virus was transmitted by manual inoculation of sap to other species of Gramineae, including oats, rice, cocksfoot and meadow fescue, but none of these hosts seemed to contain as much virus as ryegrass; their saps did not precipitate specifically with antiserum prepared against the virus in ryegrass, whereas sap from infected ryegrass precipitated up to a dilution of 1/32. Infective sap of S22 Italian ryegrass contained flexuous rod-shaped particles; the dilution end-point of the virus was about 1 in 1000; the virus was inactivated when held for 10 min. at 60°C. and most of its infectivity was lost after 24 hr. at room temperature.     

Immunodiagnosis of plant viruses by a virobacterial agglutination test

A new virobacterial agglutination (VBA) test for the immunodiagnosis of plant viruses is described. The test is based on the agglutination of Staphylococcus aureus cells by virus particles after treatment of the cells with homologous antiserum. The agglutination occurs within 1–5 min. The sensitivity of the test is 0·1-0·4 μg virus/ml and is not affected by the shape of the virus particle. The use of affinity purified antibodies for sensitisation of S. aureus cells increases the sensitivity of the reaction 50-fold and enables the detection of tobacco mosaic and cucumber green mottle mosaic viruses at a concentration of 2 ng/ml. The VBA test allows the estimation of potato viruses X, S, M and Y in the eyes and sprouts of infected tubers and in the leaves of infected plants. The diagnosis of carnation mottle virus in carnation plants and of mushroom viruses in mushroom (Agaricus bisporus) fruit-bodies and mycelium are also described.     

PROPERTIES AND HOST RANGE OF TURNIP CRINKLE, ROSETTE AND YELLOW MOSAIC VIRUSES

Three isolates of turnip yellow mosaic virus and two other flea-beetle transmitted viruses, turnip crinkle and turnip rosette, have many similar properties: thermal inactivation end-point between 80 and 90° C.; dilution end-point greater than 10^-4; longevity in vitro at about 20° C. at least 30 days. All were transmitted by mechanical inoculation to a wide range of cruciferous host plants, including many weeds. Turnip yellow mosaic virus infected only Reseda odorata outside the Cruciferae , whereas rosette virus infected a few and crinkle virus many non-cruciferous hosts.     

THE EFFECT OF DIFFERENT HOST PLANTS OF POTATO VIRUS C IN DETERMINING ITS TRANSMISSION BY APHIDS

A stock of potato virus C derived from Edgecote Purple potatoes in 1945 was not then transmitted by aphids, although more than 2000 aphids were used in conditions optimal for transmitting the serologically related potato virus Y. This stock of virus C has been propagated continuously since, by manual inoculation in a series of Nicotiana glutinosa and N. tabacum , and in 1955 it was transmitted by the aphid Myzus persicae (Sulz.): about one in twenty of the aphids transmitted it compared with one in two for potato virus Y.
Virus C derived from the Edgecote Purple potatoes in 1955 was not transmitted by aphids; both stocks of virus C produced only local lesions in Majestic potato leaves, and gave similar symptoms in tobacco.
When inoculated to Majestic potatoes and then returned to tobacco plants, potato virus C usually ceased to be aphid transmitted and did not recover this property in any of the subsequent subcultures.
Transmission from stock by aphids did not isolate a strain of virus C which was any more readily transmitted by aphids, indeed, for the first two or three subcultures, aphids usually transmitted more readily from plants inoculated manually. But the few isolates which remained aphid transmissible, after a second passage through potato, were rather readily transmitted.
These results suggest that the ability of a virus to be aphid transmitted is, at least in part, determined by the host plant in which it is multiplying, but the nature of the changes which determine this ability are unknown.     

Use of meristem tip culture to eliminate carnation latent virus from carnation plants

A successful protocol for meristem tip culture to eliminate carnation latent virus from carnation cv. scania has been described . The virus was found to be mechanically transmissible to Chenopodium quinoa, C. amaranticolor, Dianthus barbatus and Saponaria vaccaria. Murashige and Skoog'smedium (MS) supplemented with NAA (1.0 microM) and Kn (20.0 microM) proved best for meristem establishment and microshoots were rooted in MS medium supplemented with IBA (5.0 microM). Meristems measuring 0.1 and-0.2 mm yielded virus free plants and larger meristems were not effective.     

昆明地区香石竹病毒病流行状况调查及脱病毒苗的制备

对昆明地区3种不同生产模式下的香石竹（Dianthus caryophyllus L.)进行了调查,采集样本146号,利用酶联免疫法和电镜检测法对样本感染香石竹病毒的情况进行检测,结果表明昆明地区主要流行的香石竹为香石竹斑驳病毒和香石竹坏死斑点病毒,以带香石竹斑驳病毒的香石竹品种“俏新朗”为实验材料,研究了直接剥茎尖法,高温处理结合剥茎尖法和病毒痤处理结合剥茎尖法3种方法在脱病毒效率和茎尖成苗率的差异,实验结果表明以加热处理结合剥茎尖法脱病毒效果最好,0.2mm茎尖脱病毒率可达77．78％,加5％病毒座处理对脱病毒有一定的影响,直接剥茎尖法脱病毒效果最差。     

Identification and distribution of viruses infecting sweet potato in Kenya

Four hundred and forty-eight symptomatic and 638 asymptomatic samples were collected from sweet potato fields throughout Kenya and analysed serologically using antibodies to Sweet potato feathery mottle virus (SPFMV), Sweet potato chlorotic stunt virus (SPCSV), Sweet potato mild mottle virus (SPMMV), Cucumber mosaic virus (CMV), Sweet potato chlorotic fleck virus (SPCFV), Sweet potato latent virus (SwPLV), Sweet potato caulimo-like virus (SPCaLV), Sweet potato mild speckling virus (SPMSV) and C-6 virus in enzyme-linked immunosorbent assays (ELISA). Only SPFMV, SPMMV, SPCSV, and SPCFV were detected. Ninety-two percent and 25% of the symptomatic and asymptomatic plants respectively tested positive for at least one of these viruses. Virus-infected plants were collected from 89% of the fields. SPFMV was the most common and the most widespread, detected in 74% of the symptomatic plants and 86% of fields surveyed. SPCSV was also very common, being detected in 38% of the symptomatic plants and in 50% of the fields surveyed. SPMMV and SPCFV were detected in only 11% and 3% of the symptomatic plant samples respectively. Eight different combinations of these four viruses were found in individual plants. The combination SPFMV and SPCSV was the most common, observed in 22% of symptomatic plants. Virus combinations were rare in the asymptomatic plants tested. Incidence of virus infection was highest (18%) in Kisii district of Nyanza province and lowest (1%) in Kilifi and Malindi districts of Coast province.     

Switching on RNA silencing suppressor activity by restoring argonaute binding to a viral protein

We found that Sweet potato feathery mottle virus (SPFMV) P1, a close homologue of Sweet potato mild mottle virus P1, did not have any silencing suppressor activity. Remodeling the Argonaute (AGO) binding domain of SPFMV P1 by the introduction of two additional WG/GW motifs converted it to a silencing suppressor with AGO binding capacity. To our knowledge, this is the first instance of the transformation of a viral protein of unknown function to a functional silencing suppressor.     

Isolation of Carnation Italian Ringspot Virus from a Creek in a Forested Area South West of Bonn

During a survey of plant viruses in small ponds and creeks in a forested area near Bad Münstereifel about 30 km south-west of Bonn several viruses were isolated. One of these was identified as carnation Italian ringspot virus which previously has been isolated only twice from carnations originating from Italy and USA and which infected carnations only with difficulty. The role of this virus in the forest ecosystem is so far unknown.     

Viruses infecting sweet potato in Rwanda: occurrence and distribution

A survey of sweet potato virus diseases was conducted in the major sweet potato production areas in low, medium and high altitude zones of Rwanda. A total of 205 symptomatic and 103 asymptomatic samples were collected from 51 sweet potato fields and assayed for Sweet potato feathery mottle virus (SPFMV), Sweet potato chlorotic stunt virus (SPCSV), Sweet potato mild mottle virus (SPMMV), Sweet potato chlorotic fleck virus (SPCFV), Sweet potato latent virus (SwPLV), Sweet potato caulimo‐like virus (SPCaLV) and Cucumber mosaic virus (CMV) using nitrocellulose membrane enzyme‐linked immunosorbent assay. The viruses detected in the samples were SPFMV, SPMMV, SPCSV, SPCFV and SwPLV. Viruses were detected in 83% and 31% of the symptomatic and asymptomatic samples, respectively. SPFMV was detected in 49% of the samples. SPCSV, the second most common virus, was detected in 28% of samples collected from 73% of the fields. About 19% of the samples were tested positive for SPMMV. Thirteen combinations of multiple virus infections were detected in the samples. Viruses were detected in samples from all the fields surveyed, and the frequency of detection was greatest in samples from low altitude zones.     

RED CLOVER MOTTLE VIRUS

A virus, provisionally named red clover mottle virus (RCMV), isolated from red clover plants in England, seems distinct from any previously described. It was transmitted by mechanical inoculation of sap to many legumes and to Gomphrena globosa L., but it was not transmitted by six aphid species, or through soil or through seeds.
RCMV is inactivated in 10 min. between 60 and 63°C., and in 8 days at 18°C., but survives for long periods at -20; sap was not infective when diluted more than 1/1000. The virus is soluble in the pH range (4–7) in which it is stable. It was precipitated without inactivation by 50% saturated ammonium sulphate solution, but it was inactivated by ethanol or acetone. Partially purified preparations contained polygonal particles about 28 mμ in diameter. Serological tests showed no antigens in common with broad bean mottle, true broad bean mosaic or lucerne mosaic viruses.