Regeneration of virus-free grapevines using in vitro apical culture

Fragmented shoot apex culture of grapevine has been demonstrated to regenerate plants free from the graft-transmissible diseases leafroll, yellow speckle, fleck and summer mottle. This technique in combination with heat therapy of the cultures has also produced vines free from fanleaf virus. Regenerated plants free of yellow speckle were consistently obtained if cultures were incubated at 27/20 °C but not if incubated at 35 °C. The possible ways that the in vitro elimination of these diseases was achieved are considered.     

Evaluation of biological indexing and dsRNA analysis in grapevine virus elimination

Fragmented shoot apex culture (FSAC) was used to eliminate some diseases caused by viruses and virus-like agents from 24 varieties of imported grapevines. Each cultivar was examined by biological indexing before and after FSAC. Graft indexing revealed that leaf roll, stem pitting and yellow speckle diseases were common before FSAC. A correlation was observed between the incidence of leafroll disease and the presence of specific dsRNA species which were removed after FSAC. One of these species, an RNA of about one kilobase pair was associated with low yielding Sultana clones. Both biological indexing and dsRNA assay indicated no recurrence of the leafroll disease in material regenerated by FSAC even after 10 years in the field. It is concluded that dsRNA assay may be used as a faster and less expensive method than biological indexing in assessing the success of leafroll elimination by FSAC. The test also provides some information on the genome size of the viruses associated with leafroll disease. Graft indexing indicated that yellow speckle disease was resistant to elimination by FSAC, while stem pitting was removed from some of the vines and the grapevine fleck disease was eliminated from most sources.     

Incidence of Viruses and Nematode Vectors in Lebanese Vineyards

Surveys for virus diseases and nematode vectors were conducted in 95 commercial vineyards of four different Lebanese districts (Bekaa valley, Mount Lebanon, North and South Lebanon). Out of 915 randomly collected grapevine samples tested by ELISA, 511 (55.8%) were infected by one or more viruses. Grapevine virus A (30.9%) and Grapevine leafroll‐associated virus 3 (23.7%) were the prevailing viruses, followed by Grapevine fleck virus (15.1%), Grapevine leafroll‐associated virus 1 (10.6%) and Grapevine leafroll‐associated virus 2 (8.7%). Arabis mosaic virus was not found whereas Grapevine fanleaf virus (GFLV) and Grapevine virus B were little represented. The most important Lebanese grapevine varieties, i.e. Maghdouchi, Tfeifihi and Beitamouni, had average infection rates between 70% and 87%, whereas varieties of foreign origin had a better sanitary status with the exception of cvs Cinsaut and Thompson (c. 83% infection). Grapevine rupestris stem pitting‐associated virus was detected in 79 of 90 (87.8%) samples tested by RT‐PCR and closteroviruses were recorded in seven of 70 (10%) vines tested. One of these viruses was identified as Grapevine leafroll‐associated virus 5 by ELISA and partial genome sequencing. No nepoviruses other than GFLV were detected in any of 90 samples tested using three different sets of degenerate primers. Xiphinema index was found in 23 of 89 soil samples collected from vineyards, and in three of 15 samples collected primarily under fig trees in fields where no grapevines were grown.     

Transmission and differentiation of six viruses infecting hogweed (Heracleum sphondylium) in Scotland

Six viruses, code-named HV1-HV6, were transmitted manually and/or by aphids (Cavariella spp. from symptomless wild plants of hogweed (Heracleum sphondylium) in Scotland. HV1 was identified as parsnip yellow fleck virus (PYFV); anthriscus yellows virus, on which it depends for transmission by aphids, was presumably also present in the hogweed plants. HV2 was transmitted manually and by aphids and had very flexuous filamentous particles c. 700–750 nm long; it has affinities with the closteroviruses, and the name heracleum latent virus is proposed. HV3, HV4 and HV5 were transmitted manually, HV3 and HV5 also by aphids, but their particle morphology is unknown. HV6 was transmitted only by aphids and has very flexuous particles up to 1400 nm long; it is presumably a closterovirus distinct from HV2. All the viruses infected cultivated umbelliferous species experimentally but only PYFV is known to infect umbelliferous crops.     

Specificity of the effect of sap-transmissible viruses in increasing the accumulation of luteoviruses in co-infected plants

The concentration of potato leafroll luteovirus (PLRV) (c. 1300 ng/g leaf) in singly infected Nicotiana clevelandii plants was increased up to 10-fold in plants co-infected with each of several potyviruses, or with narcissus mosaic potexvirus, carrot mottle virus or each of three tobravirus isolates. With the tobraviruses, PLRV concentration was increased equally by co-infection with either NM-type isolates (coat protein-free cultures containing RNA-1) or M-type isolates (particle-producing cultures containing RNA-1 and RNA-2). In contrast, the accumulation of PLRV was not substantially affected by co-infection with either of two nepoviruses, cucumber mosaic cucumovirus, broad bean mottle bromovirus, alfalfa mosaic virus, pea enation mosaic virus or parsnip yellow fleck virus. The specificity of these interactions between PLRV and sap-transmissible viruses was retained in tests made in Nicotiana benthamiana and when beet western yellows luteovirus was used instead of PLRV.     

The influence of vines on an oligohaline marsh community: results of a removal and fertilization study

The effects of competitive suppression by vines on the non-vine plant community have received little attention in temperate habitats. This study investigated the impact vines have on their herbaceous hosts in a wetland community at two soil fertility levels. Plots in an oligohaline marsh were treated in a 2 × 2 factorial design with vine removal and fertilization over two growing seasons. There was no significant interaction between removal and fertilization treatments on any of the measured variables. Vine removal initially caused an increase in light penetration through the canopy, but by the end of the study, plots with vines removed had less light due to a 25% increase in biomass by the plants released from competition with vines. For plots with vines removed, species richness was higher during a brief period in the spring of the second year, but by the end of the study, richness in removal plots decreased relative to controls. Fertilization caused a 40% increase in biomass overall, although only two species, Sagittaria lancifolia L. and Polygonum punctatum Ell., showed dramatic increases. Despite fertilization causing a 40% decrease in light penetration to the ground, no change in species richness was observed. Overall, these results show that vine cover in this wetland suppresses non-vine species and reduces community biomass. Removal of vines increased biomass of non-vine dominants but resulted in only an ephemeral change in species richness. Fertilization did not increase the effects of vines on the non-vine community. Received: 14 November 1996 / Accepted: 10 June 1997     

Influences of herbaceous vines on community characteristics in pioneer succession stages

In the continental tropics, herbaceous vines and lianas are roughly equal in abundance, and the former are even more abundant in the temperate region. However, only little attention has been paid to the study of biological and ecological characteristics of herbaceous vines. In particular, research about effects of herbaceous vines acting as a biological control on plant communities has not been carried out in depth. Herbaceous vines are widely distributed and abundant also in humid subtropical areas, especially in the early stages of forest community succession, but their ecology is little known. The aim of our study is to understand the effect of local herbaceous vines on community characteristics in pioneer succession stages. The hypothesis was tested that herbaceous vines would have predominantly negative effects on co-occurring species, thereby reducing their diversity. Based on a quadrate method, a detailed survey of shrub and herb communities covered by herbaceous vines was conducted in the Jinyun Mountain Nature Reserve of Chongqing, SW China. The sample plots were selected based on the numbers and coverage of vines, distinguishing among high vine coverage plots, middle vine coverage plots and low vine coverage plots. All species in the plots with different herbaceous vine coverage were identified and measured. The measurements for each species included number, average height and coverage. Because of abundant tree seedlings in the habitat of forest edge plots, we only recorded the number of tree seedlings in those plots to evaluate the overall effects of vines on tree seedling regeneration. After the field investigation, herbaceous vines and other species in the plots were harvested respectively, then oven dried and weighed. The results showed that herbaceous vines had high productivity and produced a lot of branches, which caused above-ground competition and mechanical stress to other species. Herbaceous vines seriously affected species composition and species importance values of self-supporting species. In all three habitats, the number of species and families in low coverage samples was larger than that in high coverage samples, and furthermore the identities of species were different between them. Species richness significantly decreased with increasing herbaceous vine coverage, illustrating that some species disappeared. Herbaceous vines reduced species diversity of communities, and as a result, community complexity was decreased, which might also decrease community stability. Biomass of communities of self-supporting species significantly decreased with increasing herbaceous vine coverage, which suggested that herbaceous vines significantly decreased community productivity. The number of seedlings also significantly decreased with increasing herbaceous vine coverage, and seedlings were mainly distributed in lower coverage samples. Herbaceous vines reduced the light exposure in the understory, which may be the mechanistic explanation for the negative influence of vines on the performance of tree seedlings. It was concluded that herbaceous vines affected seedling quantitative dynamics and distribution, and inhibited the natural succession from shrub and herb communities to tree communities. Thus herbaceous vines not only had significant influences on community characteristics in pioneer succession stages, but also on subsequent succession stages.     

Influences of herbaceous vines on community characteristics in pioneer succession stages

In the continental tropics, herbaceous vines and lianas are roughly equal in abundance, and the former are even more abundant in the temperate region. However, only little attention has been paid to the study of biological and ecological characteristics of herbaceous vines. In particular, research about effects of herbaceous vines acting as a biological control on plant communities has not been carried out in depth. Herbaceous vines are widely distributed and abundant also in humid subtropical areas, especially in the early stages of forest community succession, but their ecology is little known. The aim of our study is to understand the effect of local herbaceous vines on community characteristics in pioneer succession stages. The hypothesis was tested that herbaceous vines would have predominantly negative effects on co-occurring species, thereby reducing their diversity. Based on a quadrate method, a detailed survey of shrub and herb communities covered by herbaceous vines was conducted in the Jinyun Mountain Nature Reserve of Chongqing, SW China. The sample plots were selected based on the numbers and coverage of vines, distinguishing among high vine coverage plots, middle vine coverage plots and low vine coverage plots. All species in the plots with different herbaceous vine coverage were identified and measured. The measurements for each species included number, average height and coverage. Because of abundant tree seedlings in the habitat of forest edge plots, we only recorded the number of tree seedlings in those plots to evaluate the overall effects of vines on tree seedling regeneration. After the field investigation, herbaceous vines and other species in the plots were harvested respectively, then oven dried and weighed. The results showed that herbaceous vines had high productivity and produced a lot of branches, which caused above-ground competition and mechanical stress to other species. Herbaceous vines seriously affected species composition and species importance values of self-supporting species. In all three habitats, the number of species and families in low coverage samples was larger than that in high coverage samples, and furthermore the identities of species were different between them. Species richness significantly decreased with increasing herbaceous vine coverage, illustrating that some species disappeared. Herbaceous vines reduced species diversity of communities, and as a result, community complexity was decreased, which might also decrease community stability. Biomass of communities of self-supporting species significantly decreased with increasing herbaceous vine coverage, which suggested that herbaceous vines significantly decreased community productivity. The number of seedlings also significantly decreased with increasing herbaceous vine coverage, and seedlings were mainly distributed in lower coverage samples. Herbaceous vines reduced the light exposure in the understory, which may be the mechanistic explanation for the negative influence of vines on the performance of tree seedlings. It was concluded that herbaceous vines affected seedling quantitative dynamics and distribution, and inhibited the natural succession from shrub and herb communities to tree communities. Thus herbaceous vines not only had significant influences on community characteristics in pioneer succession stages, but also on subsequent succession stages.     

Relative Prevalence of Grapevine Leafroll-Associated Virus Species in Wine Grape-Growing Regions of California

Some diseases manifest as one characteristic set of symptoms to the host, but can be caused by multiple pathogens. Control treatments based on plant symptoms can make it difficult to effectively manage such diseases, as the biology of the underlying pathogens can vary. Grapevine leafroll disease affects grapes worldwide, and is associated with several viral species in the family Closteroviridae. Whereas some of the viruses associated with this disease are transmitted by insect vectors, others are only graft-transmissible. In three regions of California, we surveyed vineyards containing diseased vines and screened symptomatic plants for all known viral species associated with grapevine leafroll disease. Relative incidence of each virus species differed among the three regions regions, particularly in relation to species with known vectors compared with those only known to be graft-transmitted. In one region, the pathogen population was dominated by species not known to have an insect vector. In contrast, populations in the other surveyed regions were dominated by virus species that are vector-transmissible. Our survey did not detect viruses associated with grapevine leafroll disease at some sites with characteristic disease symptoms. This could be explained either by undescribed genetic diversity among these viruses that prevented detection with available molecular tools at the time the survey was performed, or a misidentification of visual symptoms that may have had other underlying causes. Based on the differences in relative prevalence of each virus species among regions and among vineyards within regions, we expect that region and site-specific management strategies are needed for effective disease control.     

Influence of Virus on the Chlorophyll, Carotenoid and Polyamine Contents in Grapevine Microcuttings

We have studied the effect of different types of virus infections on the content in chlorophylls, carotenoids and free polyamines in shoots of vine cv. Albariño cultured in vitro. The ratio of chlorophyll a/c hlorophyll b was maintained practically constant in the different cases studied. A significant increase was observed in the polyamines, especially putrescine, in shoots infected with the different types of virus, mainly in the cases of grapevine fleck disease+grapevine leafroll disease type I and grapevine stem pitting.     

Grapevine Viruses' Detection and Sanitary Selection in Grapevine Germplasm of Calabria (Southern Italy)

A survey of grapevine viruses present in the region of Calabria (southern Italy) was carried out, and the sanitary selection was conducted on various indigenous varieties. Serological (ELISA) and molecular (multiplex RT‐PCR) tests were used to detect the viruses included in the Italian certification programme: Arabis mosaic virus (ArMV), Grapevine fanleaf virus (GFLV), Grapevine leafroll associated virus 1 (GLRaV‐1), Grapevine leafroll associated virus 2 (GLRaV‐2), Grapevine leafroll associated virus 3 (GLRaV‐3), Grapevine virus A (GVA), Grapevine virus B (GVB) and Grapevine fleck virus (GFkV). The frequency with which the above viruses have been detected was 37.4, 32.6, 12.8, 7.7, 7.3, 1.9 and 0.3%, respectively, for GVA, GLRaV‐3, GFLV, GFKV, GLRaV‐1, GLRaV‐2 and GVB. ArMV was never found. The sanitary selection allowed for the detection of 6 putative clones of ‘Arvino’, 2 of ‘Magliocco dolce’ and 2 of the rootstock ‘17–37’ free of the above‐mentioned viruses. The necessary process for the commercialization of these clones as ‘certified’ propagation material was accomplished, and their official approval by the Italian Ministry of Agriculture is currently in progress.     

Detection of Grapevine Viruses in Poland

During a 3‐year study, grapevines from 23 vineyards in Poland were surveyed for virus diseases and tested to determine the prevalence of the most economically important viruses by RT‐PCR. The rate of positive samples was 2.2% for grapevine leafroll‐associated virus 1 (GLRaV‐1), 1.9% for grapevine leafroll‐associated virus 2 (GLRaV‐2), 1.5% grapevine leafroll‐associated virus 3 (GLRaV‐3), 1.9% for grapevine virus A (GVA), 0.2% for grapevine virus B (GVB), 0.2% for grapevine virus E (GVE), 0.65% for grapevine fanleaf virus (GFLV), 20.4% for grapevine fleck virus (GFkV) and 71.9% for grapevine rupestris stem pitting‐associated virus (GRSPaV). These viruses were found to occur as single or mixed infections of different combinations in individual grapevines. The overall viral infection rate in the surveyed grapevines was 82.6%. GRSPaV is the most widely distributed virus of all the viruses currently detected in the region. DNA sequencing confirmed the identification of the viruses in selected samples, and analysis indicated that the Polish isolates shared a close molecular identity with the corresponding isolates in GenBank. To our knowledge, this is the first detection of GLRaV‐1, ‐2, ‐3, GVA, GVB, GVE, GFLV, GFkV and GRSPaV in Poland.     

Occurrence and Distribution of Grapevine Leafroll-associated Viruses 1, 2, 3 and 7 in Turkey

Grapevines in central Anatolia region of Turkey were surveyed for the prevalence of grapevine leafroll viruses. The field study and collection of samples were conducted in nine major grapevine‐growing areas. Samples collected from 622 vines were tested for Grapevine leafroll‐associated virus 1, 2, 3 and 7 (GLRaV‐1, ‐2, ‐3 and ‐7). According to diagnostic tests and surveys, 27 of 41 cultivars and 95 of 622 samples (15.27%) were found to be infected at least one virus. GLRaV‐1 (8.36%) was found to be the most frequently encountered virus associated with leafroll disease of grapes, followed by GLRaV‐3 (5.78%), GLRAV‐7 (3.86%) and GLRAV‐2 (2.41%).     

Characterisation of an Australian isolate of tomato yellow top virus

An Australian isolate of tomato yellow top virus (TYTV-A) was transmitted in the persistent manner by the aphid Myzus persicae. Its host range was mainly restricted to the Solanaceae, though Capsella bursa-pastoris and Gomphrena globosa were symptomlessly infected. TYTV-A was purified from Physalis Joridana, using an enzyme-assisted method in which the initial tissue homogenate was incubated with cellulase. Yields of purified virus were 100–900 μg/kg tissue and depended on the age of the infected plants. Maximum yields were obtained 4–5 wk after inoculation. The particles of TYTV-A were c. 24 nm in diameter, had a buoyant density of 1.34 in caesium sulphate and a coat protein mol. wt of c. 25.7 × 10³. TYTV-A was shown to be closely related serologically to potato leafroll virus (PLRV), TYTV from New Zealand and more distantly related to several other luteoviruses. An antiserum to TYTV-A was used in enzyme-linked immunosorbent assay tests to detect TYTV in field-infected tomato plants and also luteoviruses from potato plants with leafroll symptoms. It is clear that TYTV-A is a luteovirus closely related to PLRV.     

A Carlavirus and a Rhabdovirus Infecting Lonicera X brownii cv. Dropmore Scarlet in Western Canada

Chlorotic and necrotic spots, yellow mottle, and vein banding leaf symptoms were seen on two honeysuckle vines in the Botanical Garden, University of British Columbia. Electron microscopic examination showed the presence of rhabdovirus and carlavirus particles. This is the first report of a rhabdovirus in honeysuckle. The carlavirus may be honeysuckle latent virus, although it was transmitted to only three of its nine reported herbaceous hosts, and it was not transmitted by the reported aphid vector, Hydaphis foeniculi . Both types of virus particles were seen in mesophyll, phloem parenchyma and companion cells of honeysuckle. The rhabdovirus was not transmitted to any other host and, in honeysuckle, was found only in the Spring. As its concentration declined, the number of carlavirus particles increased.     

Purification, properties and transmission of parsnip yellow fleck, a semi-persistent, aphid-borne virus

Parsnip yellow fleck virus (PYFV) is the commonest cause of virus-like symptoms in parsnip plants in Britain: it is sap-transmissible but systemically infects few species outside the Umbelliferae. It has isometric particles 29–31 mμ in diameter, a sedimentation coefficient of 167s, and loses infectivity in sap after dilution to 10^-3-10^-4, heating for 10 min at 57·5–65°C, or storage at room temperature for 4–7 days. Two isolates, from parsnip and Anthriscus sylvestris respectively, are only distantly serologically related. The aphid Cavariella aegopodii transmits PYFV in a semi-persistent manner from A. sylvestris but not from parsnip. Transmission by aphids apparently depends on the presence in A. sylvestris or other source plants of a second virus, anthriscus yellows (AYV), which is persistent in the vector and not manually transmissible. PYFV was therefore not transmitted by aphids from manually inoculated plants or from parsnip or other plants immune to AYV. In controlled experiments, C. aegopodii transmitted PYFV (both A. sylvestris and parsnip isolates) from chervil plants inoculated separately with PYFV and AYV, but not from plants inoculated only with PYFV.