Unusual filamentous virus-like particles in symptomless blackberry associated with severe leaf curling in the virus indicator Rubus macraei, and anomalous data using a degenerate badnavirus primer in PCR of Rubus species

Electron microscopy of ultrathin sections of leaves of symptomless Himalaya Giant blackberry and of the virus indicator species, Rubus macraei, showing severe leaf curl symptoms following graft inoculation with scions from this blackberry, detected highly flexuous virus‐like particles with an unusual ‘beaded’ structure. Such particles were restricted to a few vascular cells and were distinct from P‐protein common in some such cells. This virus, provisionally named Hawaiian rubus leaf curl virus (HRLCV), symptomlessly infected a wide range of Rubus species and cultivars. Badnavirus‐like bacilliform particles were observed in some cells of a single R. macraei plant showing leaf curl symptoms following graft inoculation with the causal agent of this disease symptom from Himalaya Giant blackberry after passage through red raspberry, but not in any other material. PCR with primer sets for the badnaviruses Rubus yellow net virus and Gooseberry veinbanding associated virus, showed that no Rubus sources studied contained these viruses. However, using a sequence‐specific primer set designed from the sequence of the product generated with a badnavirus degenerate primer set, a specific product was amplified from healthy plants of all of 16 raspberry cultivars and two Rubus species, but not from 16 blackberry cultivars (including cv. Himalaya Giant). All of these sources were free from viruses known to occur in Rubus. Sequence analysis of this product showed no homology with any known badnavirus, or with any other published sequences. It seems most likely therefore that a region of the raspberry genome has been amplified using the degenerate badnavirus primer set and that it is absent from the blackberry genome.     

The effect of fungicides, irrigation and plant density on the development of Peronospora sparsa, the cause of downy mildew in rose and blackberry

Downy mildew of rose (Rosa spp.) and blackberry (Rubus fructicosus), caused by Peronospora sparsa, can cause significant losses in production. In a series of experiments, fungicides with different modes of action to the commonly used phenylamide‐based products were examined for efficacy on both crops. Cymoxanil + mancozeb + oxadixyl and fluazinam gave good downy mildew control on both rose and blackberry. On outdoor, container‐grown rose, high volume sprays of fosetyl‐aluminium were also effective, but on young micropropagated blackberry plants, application as a drench treatment was better than as a spray. Good control was also achieved on blackberry with chlorothalonil and with metalaxyl in formulation with either thiram or mancozeb. There was no evidence of loss of control with phenylamide fungicides due to resistance. Irrigation regimes were also examined on blackberry and sub‐irrigation of plants in propagation on a sand bed led to significantly lower disease levels than those found where plants had been irrigated by overhead sprinklers. Reducing the density of container‐grown rose plants had a measurable effect on the progress of downy mildew, though this was small and temporary when compared with the effect of fungicide treatment. Fungicide programmes which gave season‐long control of rose downy mildew resulted in significantly improved extension growth.     

Purification and properties of blackberry chlorotic ringspot, a new virus species in subgroup 1 of the genus Ilarvirus found naturally infecting blackberry in the UK

A mechanically transmissible virus was isolated from Bedford Giant blackberry plants showing chlorotic mottling and ringspot symptoms growing in Scotland. It infected several herbaceous test plants, many of them symptomlessly. This virus was also transmitted to several Rubus species and cultivars by graft inoculation with scions from the field‐infected Bedford Giant plant. Most grafted plants were infected symptomlessly, but Himalaya Giant blackberry and the hybrid berry Tayberry developed symptoms similar to those in the infected Bedford Giant plant. In the sap of infected Chenopodium quinoa, the virus lost infectivity when diluted 10^?4 but not 10^?3, after 6 h and 48 h when kept at 20°C and 4°C, respectively, but was infective for more than 8 days when kept at ?15°C. Preparations of purified virus from infected C. quinoa or spinach sedimented as three major nucleoprotein components and consisted of quasi‐isometric particles that varied in size from 24 to 32 nm in diameter and that were not penetrated by negative stain. Such virus particle preparations contained a major polypeptide of ca 28 kDa and three single‐stranded RNA species of estimated size 3.2, 2.8 and 2.1 kb. The complete sequence of the largest RNA (RNA 1, 3478 nt) and the partial sequence of the other RNAs (1863 and 2102 nt long, respectively) were determined and compared with sequences in databases. These findings, together with the biological and biochemical properties of this virus, indicate that it should be regarded as a distinct species in subgroup 1 of the genus Ilarvirus even though it was serologically unrelated to existing members of this subgroup. The virus showed a very distant serological relationship with prune dwarf virus (PDV) but differed significantly from it in the amino acid sequence of its coat protein, experimental host range and symptomatology and was unrelated to PDV at the molecular level. The virus, tentatively named blackberry chlorotic ringspot virus, is therefore a newly described virus and the first ilarvirus found naturally infecting Rubus in the UK.     

Epidemiology of Raspberry Bushy Dwarf Virus in the Czech Republic

Extensive monitoring of the raspberry bushy dwarf virus (RBDV) in cultivated raspberry, wild raspberry and blackberry was conducted in 1994‐99. RBDV was revealed by ELISA in 31.6% of field samples, 15.7% plants in germplasm collections and in 43.8% of propagated plants. Infected cultivars were Aborigen, Balzam, Brigantina, Bulharský Rubín, Canby, Comox, F‐103, Findus, Gatineau, Glen Moy, Granát, Heritage, Lloyd George, M‐101, Mája, Meeker, Norfolk Giant, Norna, NS?‐1D‐101, Skeena, Trent, Veten, ZamatoS? and Zeva. The virus was detected in 6.5 and 6.7% of wild raspberry and wild blackberry plants, respectively, at 22.8% and 11.4% of sampled locations. Vegetatively propagated plants seem to be the main source for virus spread in cultivated raspberry, rather than naturally infested wild Rubus populations.     

Genomic characterisation and the detection of raspberry chromatin in polyploid Rubus

 This paper reports genomic in situ hybridization (GISH) and fluorescent in situ hybridization (FISH) data for chromosomes of raspberry (Rubus idaeus 2n=2x=14), blackberry (Rubus aggregate, subgenus Eubatus. 2n=2–12x=14–84) and their allopolyploid derivatives used in fruit breeding programmes. GISH was used to discriminate labelled chromosomes of raspberry origin from those of blackberry origin in allopolyploid hybrid plants. The raspberry chromosomes were labelled by GISH at their centromeres, and 1 chromosome was also labelled over the short arm. In one allopentaploid plant a chromosome carried a terminal signal. Karyotype analysis indicated that this is a blackberry chromosome carrying a raspberry translocation. GISH analysis of an aneuoctaploid blackberry cv ‘Aurora’ (2n=8x=58) showed that both whole and translocated raspberry chromosomes were present. The basic Rubus genome has one ribosomal DNA (rDNA) locus, and in all but one case all levels of ploidy had the expected multiples of rDNA loci. Interestingly, in the blackberry cv ‘Aurora’, there were only six sites, two less than might be predicted from its aneuoctaploid chromosome number. Our results highlight the potential of GISH and FISH for genomic designation, physical mapping and introgression studies in Rosaceous fruit crops. Received: 20 February 1998 / Accepted: 12 May 1998     

Host specificity of the rust Phragmidium violaceurn, a potential biological control agent of European blackberry

The host specificity of the rust fungus Phragmidium violaceum, a potential biological control agent of European blackberry (Rubus fruticosus) was studied by inoculating a mixture of 15 isolates of the rust on 108 plants of importance to the Australasian region. A scale of infection types was developed based on the results of microscopic and macroscopic observations of the reaction of host and non-host plants to the rust. The results showed that P. violaceum has a limited host range in the genus Rubus. The rust was able to reproduce on 17 taxa of Rubus previously unrecorded as hosts, including Australasian species of Rubus subgenera Dalibarda and Lampobatus. All other taxa attacked were species of Rubus subgenus Eubatus and the majority were hybrid cultivars containing European blackberry species.     

Infectibility and sensitivity of UK raspberry, blackberry and hybrid berry cultivars to Rubus viruses

Six blackberry or hybrid berry cultivars and 19 raspberry cultivars were assessed for their infectibility with, and sensitivity to, graft inoculation with 10 distinct viruses found infecting Rubus in the UK. Cultivars were grafted with each of, two isolates of the pollen borne raspberry bushy dwarf virus (RBDV), five aphid borne viruses: black raspberry necrosis, raspberry leaf mottle (RLMV), raspberry leaf spot (RLSV), rubus yellow net and raspberry vein chlorosis (RVCV); and isolates of the nematode transmitted nepoviruses, arabis mosaic, raspberry ringspot, strawberry latent ringspot and tomato black ring. All tested cultivars were infectible with a resistance breaking isolate of RBDV but only about half of that number with the Scottish type isolate of the virus. The raspberry cvs Autumn Bliss, and occasionally Glen Garry and Glen Prosen, developed leaf yellowing symptoms following infection with RBDV, but none of the other infected cultivars showed obvious leaf symptoms when kept in a heated glasshouse during the growing season. All tested cultivars were infectible with each of the four viruses transmitted in nature by the aphid, Amphorophora idaei. Most were infected symptomlessly, but seven cultivars developed severe leaf spotting symptoms due to infection with RLMV or RLSV. All but one of the raspberry cultivars were infectible with RVCV, which is transmitted in nature by the aphid Aphis idaei, and almost all infected plants developed leaf symptoms; only one of the hybrid berry or blackberry cultivars tested was infected with RVCV. In tests with the four nepoviruses, all tested cultivars, except Tummelberry, were infectible with at least one or more of these viruses. However, cultivars responded differently to challenge inoculation with different isolates of individual nepoviruses. Several cultivars developed chlorotic leaf mottling following infection with some nepovirus isolates. The implications of these results for virus control are discussed in the light of the changing pattern of virus and virus vector incidence in the UK.     

Dispersal of Acalitus essigi to blackberry (Rubus fruticosus agg.) fruit

The eriophyoid mite, Acalitus essigi (Hassan), is hypothesised to be responsible for redberry disease of blackberry (Rubus fruticosus L. agg.) fruit. Infested fruit ripen unevenly with affected drupelets becoming hard, inedible and bright red, whereas unaffected drupelets ripen evenly. As a first step toward possible control of this disease, the method and timing of dispersal of A. essigi onto developing blackberry fruit was examined. No mites were found on unopened flower buds or open flowers. However, infestation of fruit was found to commence during the green stage of fruit development and significantly increase during the red fruit stage. Although redberry symptoms were not observed, experimental exclusion of A. essigi to prevent the mite moving up the pedicel of open flowers by a sticky barrier significantly reduced A. essigi populations within the resultant fruit by over five fold that of control fruit. Although very low levels of aerial dispersal onto fruit cannot be discounted it was concluded that non-aerial or crawling dispersal via the fruit pedicel was the dominant method of blackberry fruit infestation.     

Micropropagation of three berry fruit species using nodal segments from field-grown plants

Plant micropropagation of blueberry (Vaccinium corymbosum L. cv. Berkeley), blackberry (Rubus sp. cv. Smoothstem) and raspberry (Rubus idaeus L. cv. Gradina) was carried out from nodal segments of adult field‐grown plants. Hardwood and softwood cuttings were studied as explant sources. The cultures successfully established were softwood from all three species, and hardwood only from blueberry. Shoot‐bud establishment from blueberry was achieved by culturing explants in WPM salts with MS vitamins for 15 days, and then 30 days in the same medium with 18 mM Zeatin. The best results of multiplication were obtained in the same medium with 25 mM 2iP. For blackberry, shoot‐bud establishment was achieved by culturing explants in MS medium for 15 days, and then in the same medium with 4 mM BA and 0.25 mM IBA. This medium was also the best for blackberry multiplication. Raspberry explants (cvs Gradina and Willamette) were cultured in MS medium for 15 days and then transferred to MS medium supplemented with 4 μm BA and 0.25 mM IBA. After 30 days of culture, only ‘Gradina’ explants survived, from which shoot‐bud establishment was obtained in a modified MS medium (Anderson's macronutrients except calcium, with Sequestrene as the iron source) with the same growth regulators. Multiplication was achieved by subculturing explants in the same medium either with 4 mM BA plus 0.25 mM IBA or with 8 mM BA plus 0.25 mM IBA. Shoots of at least 1 cm in length from all species were rooted ex vitro in a mixture of peat and Perlite (1:1, v/v) in a mist chamber, and 100% of rooting plants were acclimated. Bacterial, fungal and viral diseases were detected in stock plants, while tests carried out in both shoots and regenerated plants revealed the absence of any kind of disease.     

Fruit quality attributes of blackberry (<Emphasis Type="Italic">Rubus sanctus</Emphasis>) mutants obtained by <Superscript>60</Superscript>Co gamma irradiation

Blackberry mutants were induced by the irradiation of blackberry shoots with 5 kilo radians of cobalt (⁶⁰Co) gamma irradiation. Seven mutant plants demonstrating improved plant growth properties were selected for evaluation of the fruit quality characteristics, such as size, shape, color, firmness, soluble solids content, juice acidity, nutritive value (e.g. ascorbic acid) and overall fruit flavor. The organoleptic properties, including texture, firmness, flavor, sweetness, acidity, color, and overall fruit flavor, were evaluated by a consumer panel. The results indicated that irradiation induced changes in some of the quality parameters of blackberry fruits, as well as increased the fruit shelf-life.     

The collection and selection in Europe of isolates of Phragmidium violaceum (Uredinales) pathogenic to species of European blackberry naturalised in Australia

Phragmidium violaceum is a rust fungus with potential for the biological control of European blackberry (Rubus fruticosus) in Australia. The collection, selection, purification and multiplication in Europe of isolates of the rust is described. Species of European blackberry naturalised in Australia showed different levels of susceptibility when inoculated with a pool of 15 isolates highly pathogenic to Rubus procerus. Selection of individual isolates on the four most widespread blackberry species showed that only two of the isolates would be required to obtain the best attack on these four species.     

Diagnosis and management of Sclerotinia stem rot (white mould) of lentils in Greece

The paper describes the field-level symptoms, the identification and management of Sclerotinia stem rot of lentil caused by the soilborne plant pathogen Sclerotinia sclerotiorum in Greece. Regarding symptoms at a field level, initially plants before flowering turn yellow with roots and the base of the plants become brown; then rotten plants exhibit a dry stem and die. On the diseased tissue, at the base of the stem, the typical white mycelium and the resting bodies (sclerotia) were observed. According to our pathogenicity studies in vitro, on the infected plant tissues the fungus first develop a characteristic fluffy white mycelium which will give rise to large black sclerotia, the most obvious evidence of plants infected with S. sclerotiorum. Finally, concerning evaluation of fungicides, isolates of S. sclerotiorum were sensitive to thiophanate-methyl and to triazole fungicides. Thiophanate-methyl and triazole fungicides proved to be most effective in controlling the disease emerged from mycelium or sclerotia.     

Genetic relationships among wild and cultivated blackberries (Rubus caucasicus L.) based on amplified fragment length polymorphism markers

Abstract

Rubus is a large genus of flowering plants in the rose family, Rosaceae, subfamily Rosoideae. The blackberries, as well as various other Rubus species with mounding or rambling growth habits, are often called brambles. Little information is available on the genetic diversity of wild-grown blackberries. The objective of this study was to determine the genetic relationships among nine promising (high-yield capacity, free of pest and diseases, better fruit traits) wild blackberry (Rubus caucasicus L.) selections and the well-known cultivar, “Chester” by using amplified fragment length polymorphism (AFLP) markers. Genotypes were evaluated with three selective primer-enzyme combinations, producing a total of 223 AFLP fragments with 53% polymorphism ratio. Clustering of genotypes using unweighted pair-group method of arithmetic average cluster analysis clearly separated groups of wild blackberry genotypes while the variety “Chester” was clustered independently. Wild selections represented a distinct germplasm source on the basis of the estimated genetic distance among them. Genetic diversity data from this study will be helpful in using and exploiting the wild genetic material for breeding purposes as well as for further research.     

DNA fingerprinting reveals significant amounts of genetic variation in a wild raspberry Rubus idaeus population

Establishing the genotypic distribution in natural plant populations is an important part of ecological studies concerning plant growth, reproduction and turn-over. Restriction enzyme-digested DNA samples, isolated from 24 plants of a natural Rubus idaeus population, were analysed with DNA fingerprinting using the M13 repeat sequence as well as a synthetic (AC)/(TG) polydinucleotide as hybridization probes. All the examined samples exhibit unique DNA fingerprint patterns, suggesting that vegetative reproduction may be considerably more restricted in wild R. idaeus populations than previously assumed. By comparison, all samples of the apomictic blackberry species Rubus nessensis, collected on the same location, were completely identical.     

Differential Organ Distribution,Pathogenicity and Benomyl Sensitivity of Colletotrichum spp. from Blackberry Plants in Northern Colombia

Blackberry anthracnose, caused by Colletotrichum spp., is an important disease of cultivated blackberry in the world. In Colombia, it is the number one limiting factor for commercial production. This study was conducted to determine the species of Colletotrichum infecting blackberry plants as well as the organ distribution, pathogenicity and response to benomyl of the isolated strains. Sixty isolates from stems (n = 20), thorns (n = 20) and inflorescences (n = 20) were identified as Colletotrichum acutatum and Colletotrichum gloeosporioides by a species‐specific polymerase chain reaction (PCR). Both Colletotrichum species were found in the same plant but on different organs. Colletotrichum gloeosporioides species predominated in thorn lesions (n = 16) and C. acutatum in stems (n = 15) and inflorescence (n = 15). Pathogenicity assays on detached blackberry organs demonstrated differences between the two species with an average period of lesion development of 8.7 days for C. gloeosporioides and 10.3 days for C. acutatum. Wound inoculated organs had 90% disease development compared to 17.5% in non‐wounded. All C. acutatum isolates (n = 34) were benomyl tolerant, whereas C. gloeosporioides isolates (n = 26) were 30.7% sensitive and 69.2% moderately tolerant. Phylogenetic analysis with ITS sequences of a subset of 18 strains showed that strains classified as C. gloeosporioides had 100% identity to Colletotrichum kahawae, which belongs to the C. gloeosporioides species complex, whereas C. acutatum strains clustered into two different groups, with high similarity to the A2 and the A4 molecular groups. These data demonstrate for the first time the differential distribution of both species complexes in blackberry plant organs and further clarifies the taxonomy of the strains.     

The genome of black raspberry (Rubus occidentalis)

Black raspberry (Rubus occidentalis) is an important specialty fruit crop in the US Pacific Northwest that can hybridize with the globally commercialized red raspberry (R. idaeus). Here we report a 243 Mb draft genome of black raspberry that will serve as a useful reference for the Rosaceae and Rubus fruit crops (raspberry, blackberry, and their hybrids). The black raspberry genome is largely collinear to the diploid woodland strawberry (Fragaria vesca) with a conserved karyotype and few notable structural rearrangements. Centromeric satellite repeats are widely dispersed across the black raspberry genome, in contrast to the tight association with the centromere observed in most plants. Among the 28 005 predicted protein‐coding genes, we identified 290 very recent small‐scale gene duplicates enriched for sugar metabolism, fruit development, and anthocyanin related genes which may be related to key agronomic traits during black raspberry domestication. This contrasts patterns of recent duplications in the wild woodland strawberry F. vesca, which show no patterns of enrichment, suggesting gene duplications contributed to domestication traits. Expression profiles from a fruit ripening series and roots exposed to Verticillium dahliae shed insight into fruit development and disease response, respectively. The resources presented here will expedite the development of improved black and red raspberry, blackberry and other Rubus cultivars.     

A strain of radish mosaic virus occurring in turnip in Yugoslavia

Fields of turnip (Brassica rapa L. var. rapa) in Yugoslavia often contained plants infected with radish mosaic virus (RaMV). Yugoslav isolates resemble the type strain in host range, symptoms, physical properties, particle morphology and flea-beetle transmission and are serologically closely related to the type strain, but did not infect radish and are designated as the European strain. Light microscopy of infected turnip and some other cruciferous species revealed the characteristic inclusion bodies in the cytoplasm.     

Priming fingerprint induced by Bacillus amyloliquefaciens QV15, a common pattern in Arabidopsis thaliana and in field-grown blackberry

The aim of this study is focused on determining the Bacillus amyloliquefaciens QV15 priming fingerprint in two different plant species, Arabidopsis and blackberry as a crop of agronomic interest, associated with protection upon pathogen challenge. To achieve this goal, Arabidopsis thaliana plants were challenged with Pseudomonas syringae DC3000 under controlled conditions, and field-grown blackberries were challenged by a powdery Mildew outbreak, finding plant protection in plants treated with QV15, in both conditions. Changes in ROS scavenging enzymes’ activity, defense-related enzymes’ activity and gene expression were evaluated in both plant species, before and after pathogen challenge, revealing the ability of this strain to prime both. As a result of this analysis, the priming fingerprint induced by QV15 was defined by a decrease in ROS scavenging enzymes’ activity in pre- and post-challenged plants, an increase in glucanase and chitinase activity after pathogen challenge, significantly increasing the expression of PR1, indicating a salicylic acid (SA)-mediated pathway activation. These results suggest an excellent potential of B. amyloliquefaciens QV15 to protect different plant species against different pathogens in field conditions.     

Trade-off among antiherbivore defences in a south american blackberry (Rubus bogotensis)

Summary The idea of trade-offs among antiherbivore defences in plants is examined using data from a South American blackberry (Rubus bogotensis). Two distinct morphs of R. bogotensis, one with glandular trichomes and one without, were compared with respect to leaf toughness, number of prickles and prickle length. The two morphs were sympatric and grew under similar environmental conditions. The morph lacking trichomes had significantly tougher leaves and also tended to have more and longer prickles. Bioassay showed that Ithomiid larvae fed to a lesser extent on tough leaves than on more tender ones. Correlations between antiherbivore defences within each phenotype revealed three significant or almost significant negative relationships. The comparisons support the hypothesis that trade-offs exist among antiherbivore defences.