Chromosome numbers of European blackberries (Rubus subg.Rubus,Rosaceae)

Chromosome numbers are presented for 32 collections of 29 European blackberry species (Rubus subg.Rubus) from Germany. One species is triploid (2n = 21), 27 species are tetraploid, (2n = 28), and one species is pentaploid (2n = 35). Chromosome numbers are reported for the first time ofR. adspersus, R. amisiensis, R. calvus, R. conothyrsoides, R. contractipes, R. demissus, R. elegantispinosus, R. ferocior, R. foliosus, R. hypomalacus, R. leucandrus, R. nemorosus, R. platyacanthus, R. praecox, R. rhombifolius, andR. rhytidophyllus. Chromosome numbers forR. dasyphyllus, R. gelertii, R. glandithyrsos, R. lamprocaulos, R. lindebergii, R. macrophyllus, R. montanus, R. muenteri, R. pedemontanus, R. polyanthemus, R. senticosus, R. silvaticus, andR. vigorosus are confirmed.     

Establishment and characterization of Rubus tissue culture systems for in vitro bioassays against phytotoxins from Rubus fungal pathogens

The Rubus species R. parviflorus, R. spectabilis and R. strigosus interfere with conifer seedling establishment on forest regeneration sites in Canada and the United States. As a first step towards microbial metabolite-based control, callus and cell suspension cultures of the Rubus species were developed as a bioassay system to detect phytotoxic compounds that may have relevance in a vegetation control context. Rapidly growing friable callus and suspension cultures were obtained from leaf disks of the three weedy Rubus species using similar culture media conditions (modified Murashige and Skoog) but required different plant growth regulators (R. parviflorus, 4.5 M 2,4-D; R. spectabilis, 26.9 M NAA/0.5 M zeatin; and R. strigosus, 12.4 M picloram). Cell growth and health attributes including callus circumference, degree of browning and suspension culture cell viability as measured by the TTC vital stain assay were developed and were rapid and convenient to use. We have established Rubus tissue culture systems that will make it possible for large scale screening of phytotoxic metabolites.     

Apomixis versus sexuality in blackberries (Rubus subgen.Rubus,Rosaceae)

InRubus L. a connection seems to exist between the degree of meiotic disturbances on the one hand, and the production of unreduced embryo sacs, pollen fertility and relative seed set on the other hand. Severe meiotic disturbances commonly encountered in apomictic taxa decrease pollen fertility and thereby seed set since pollen is necessary for endosperm development. By contrast interspecific hybrids between apomictic taxa appear to be sexual and exhibit high pollen fertilities, probably due to an improved meiosis. Thus, apomixis leads to a decreased fertility inRubus, not the opposite, as often discussed.     

Rubus laconensis and Rubus pignatti (Rosaceae), two new species from Sardinia (Italy)

Rubus is poorly represented in Sardinia, mainly by the widespread R. ulmifolius and further by two narrow endemic species. Two new species, R. laconensis and R. pignatti, here described and illustrated, grow in two neighbouring small areas of the Sarcidano country. Both are characterized, in particular, by fruits with a low number of large drupelets. R. laconensis is for the time being attributed to the R. caesius aggregate, and R. pignatti to R. subg. Rubus.     

Characterisation of Rubus niveus: a prerequisite to its biological control in oceanic islands

This study is intended to inform the search for a biocontrol solution to stem the spread of Rubus niveus, a morphologically-diverse and invasive Rubus species which has become an aggressive invader in areas of the world outside its native range. The fragile biodiversity of islands can be especially threatened by this plant – exemplars of this are the Galápagos archipelago and Hawaii. This report focuses on the morphological and molecular work carried out to establish the homogeneity or intra-specific variation of R. niveus populations within the Galápagos Islands, as well as their kinship with plants of the same name found in Asia. The premise of this study is that genetic closeness between these populations offers increased probability that pathogens of R. niveus in its native range will also provide effective control in the target environment. Increased knowledge of the genetic make-up of R. niveus in both its native and naturalised locations will also help to assess the feasibility of using organisms which have been found to be effective against other Rubus species, but not hitherto considered for R. niveus itself.     

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.     

Foliar indumentum in central‐European Rubus species (Rosaceae) and its contribution to the systematics of the group

The foliar indumentum of 94 taxa of Rubus has been surveyed by scanning electron microscopy. The analysed species belonged to five subgenera. Rubus subgen. Rubus with its three sections and 21 series was the most numerously represented. In all studied species, indumentum was present, but the species differed more or less from each other with respect to the density of the indumentum on the abaxial leaf surface. Three main hair types were present: simple eglandular (unbranched) trichomes, branched eglandular trichomes, and very short secretory uni‐seriate trichomes; subtypes were distinguished according to trichome size. Apart from investigating separate traits, we described character patterns for the best represented and homogeneous series: Discolores, Subthyrsoidei, Rhamnifolii and Rubus. Other large groups (Sylvatici, Glandulosi and Micantes) were more diverse.     

Incidence and genetic diversity of raspberry bushy dwarf virus (RBDV) in Rubus spp. in Turkey

Raspberry bushy dwarf virus (RBDV), recently renamed to Idaeovirus rubi, is one of the most common viruses infecting Rubus species worldwide but there is still a limited number of genome sequences available in the GenBank database and the majority of the sequences include partial sequences of RNA-1 and RNA-2. The distribution and incidence of RBDV in main raspberry and blackberry growing provinces in Turkey were monitored during 2015–2019 and 537 Rubus spp. samples were tested by both DAS-ELISA and RT-PCR. Among the tested samples, 36 samples tested positive for RBDV by DAS-ELISA and 67 samples by RT-PCR. There was relatively low nucleotide diversity among the Turkish isolates. Turkish isolates shared 93%–97.7%, 84.3%–98.9%, and 85%–99.2% nucleotide sequence identities with available sequences in the GenBank, in partial RNA-1, movement protein (MP) and coat protein (CP) genes, respectively. In the phylogenetic tree constructed for RNA-1, MP, and CP sequences, all Turkish raspberry isolates were clustered in a distinct clade. However, the blackberry isolates showed considerable variation in nucleotide sequences and were placed in three distinct groups. The divergent blackberry isolates showed high variability in MP (84.5%–89.3%) and CP (85.5%–89.7%) regions and were placed in a distinct group. The rest of blackberry isolates clustered together with sweet cherry RBDV isolates adjacent to the grapevine clade or together with raspberry isolates. The comparative analysis conducted on three RNA segments of RBDV highlighted the high sequence diversity of Turkish RBDV isolates. This study also emphasizes the importance of regular monitoring of RBDV infections in Turkey, with special regard to those Rubus spp. and grapevine accessions employed in conservation and selection programmes. In particular, the presence of new RBDV genetic variants and infection of Rubus species must be taken into account to choose a correct detection protocol and management strategy.     

Melting pot of biodiversity: first insights into the evolutionary patterns of the Colchic bramble flora (Rubus subgenus Rubus,Rosaceae)

The Caucasus is a biodiversity hotspot of global significance, containing a number of highly diverse and species‐rich plant taxa. The region is also thought to be an important evolutionary hotspot for Rubus subgenus Rubus (brambles). However, Caucasian brambles have only been poorly studied to date and our knowledge of their evolutionary mechanisms, systematics and taxonomic variability remains rudimentary. Therefore, the objectives of this study were to shed light on the evolution, diversity and reproduction modes of Rubus in one of the two Caucasian glacial refugia, Colchis. Flow cytometry measurements were used to estimate DNA ploidy, a flow cytometric seed screen was conducted to determine reproduction mode and Sanger sequencing of two non‐coding plastid regions was used to reveal phylogenetic patterns. The most common ploidy level was tetraploid, followed by diploid and (rarely) triploid. Intra‐individual variation in reproduction mode was low, as the morphoseries Glandulosi and Radula exhibited strict sexuality and other taxa were mostly apomictic. A few exceptions were observed that deserve further attention, e.g. sexuality induced hypothetically by haploid pollen or by environmental conditions, a high proportion of triploid embryos or polyspermy. Plastid haplotype variability revealed specific, ancient evolutionary patterns with limited involvement of extant diploid taxa and recent isolation from European brambles. We provide the first insight into the variability and evolution of Colchic brambles, which serves as a starting point for further systematic and evolutionary studies.     

Development and use of simple sequence repeat SSR markers in Rubus species

The isolation of polymorphic codominant microsatellite markers in Rubus and in particular red raspberry will provide a tool to investigate gene flow between cultivated and wild raspberries. Microsatellite loci were isolated by screening a PstI size selected genomic library with AC₍₁₃₎ and AG₍₁₃₎. Positive clones were sequenced and primer pairs designed to the sequences flanking identified SSRs. One primer of each pair was fluorescently labelled to facilitate polymerase chain reaction (PCR) product identification on an automated DNA sequencer. We describe 10 polymorphic microsatellite loci developed and demonstrate their usefulness in different Rubus species.     

Identification of Rubus yellow net virus as a distinct badnavirus and its detection by PCR in Rubus species and in aphids

Rubus yellow net virus (RYNV) infects Rubus species and cultivars worldwide and is an essential component of raspberry veinbanding mosaic (RVBMD), a virus disease complex that causes serious decline in plant vigour and productivity. The virus is transmitted, probably in a semi‐persistent manner, by the large raspberry aphid, Amphorophora idaei in Europe, and A. agathonica in North America. The particles of RYNV are bacilliform in shape and measure 80–150 × 25–30 nm, similar to those of badnaviruses. A1.7 kb fragment of the viral DNA was amplified by PCR and then directly sequenced. Analysis of this sequence suggests that RYNV is possibly a distinct species in the genus Badnavirus and is most closely related to Gooseberry vein banding associated virus (GVBAV) and Spiraea yellow leaf spot virus, two other badnaviruses described recently. Using the sequence derived from the PCR‐amplified viral DNA fragment, RYNV‐specific primers were designed and used in PCR to assay for RYNV in a range of Rubus germplasm infected with RYNV, with other unrelated viruses and virus‐like diseases found in Rubus, and in healthy plants. RYNV was detected in all glasshouse cultures of RYNV‐infected plants, whether alone or in complex infections with other viruses, but not from healthy Rubus plants, nor from plants infected with other viruses. It was also detected in field‐grown raspberry plants with and without symptoms of RVBMD and in raspberry plants infected with RYNV by viruliferous A. idaei. RYNV was also detected by PCR in A. idaei following access feeds on RYNV‐infected plants of 1 h or more. PCR failed to amplify DNA from gooseberry infected with GVBAV confirming the specificity of the RYNV analysis. PCR detection of RYNV in dormant raspberry buds allows assays to be made outside the natural growing season, providing a useful application for plant introduction and quarantine programmes.     

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 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.     

Accessory Buds in the Genus Rubus with Particular Reference to R. idaeus L.

Accessory (secondary) axillary buds occurred in 21 out of 32Rubus species, including wild raspberry, R. idaeus L. The adaptivevalue of the secondary-bud character is discussed. Disbuddingexperiments on three raspberry cultivars and their inbred derivativesrevealed considerable variability in proportion of secondarylaterals developing and in numbers of flowers per secondarylateral, suggesting the possibility of selection for high secondarycrop potential.     

Mycoplasma-like bodies associated with Rubus stunt disease

Mycoplasma-like bodies were found by electron microscopy in the sieve tubes of blackberry, loganberry and raspberry plants showing symptoms of Rubus stunt disease, but not in those of normal plants. Watering diseased blackberry plants with aureomycin caused remission of the symptoms and greatly decreased the number of mycoplasma-like bodies.     

Chloroplast DNA diversity in the genusRubus (Rosaceae) revealed by Southern hybridization

The variability in chloroplast DNA type of 20Rubus genotypes was examined by Southern hybridization. DNA extracted from theRubus accessions was digested with two restriction enzymes (EcoRI and EcoRV) and heterologous chloroplast DNA sequences from barley and pea were used as probes to detectRubus chloroplast DNA sequences on Southern blots ofRubus total DNA. Restriction fragment length polymorphism was detected and a total of 92 restriction fragments were generated by the probe/enzyme combinations examined. Cladistic principles based on the parsimony assumption were used to assemble a phylogenetic tree based on chloroplast restriction fragment length data. The phylogenetic tree grouped the taxonomically defined species and is in general agreement with information based on morphological criteria. However, the Japanese red raspberryR. illecebrosus was shown to have diverged considerably in terms of evolutionary time from other species in subg.Idaeobatus. Furthermore, the molecular approach provides a quantitative estimate of the relationship between species that is difficult to obtain from morphological data. In order to complement the chloroplast DNA information a ribosomal DNA probe was also included in the analysis and provided further information on the phylogenetic relationships withinRubus.     

Isolation and characterization of simple sequence repeat loci in Rubus hochstetterorum and their use in other species from the Rosaceae family

The identification of microsatellite loci in Rubus hochstetterorum provides an important tool for the characterization and conservation of wild populations of this species. Cross‐species amplification of markers may be of particular interest for the study of other Rubus species. In this study, 41 simple sequence repeat markers were identified in a genomic library of R. hochstetterorum. Fifteen of the identified microsatellite loci were characterized in a set of 30 samples and revealed to be polymorphic with three to 19 alleles per locus. All the identified markers allowed cross‐species amplification in at least one of the other three tested species from the Rosaceae family.     

Small-scale spatial pattern and size structure of Rubus hirtus in a canopy gap

Abstract. The response of a Rubus hirtus population to the creation of a new forest gap was analysed. All individuals in two 30-m² plots – one in a gap and one with full canopy cover – were mapped and the length of Rubus shoots measured. This was done 1, 2, 3, 4, and 7 yr after creation of the gap. Spatial patterns were analysed using Ripley's K-method. Although the density of Rubus in the gap increased from 3 to 24 rooting points/m², the change in spatial pattern of individuals over 7 yr was very limited. During the first four years a clumped pattern appeared at distances of > 0.5 m. After seven years the pattern was random at all distances. Young shoots spread in all directions with equal probability. The expansion was achieved by establishment of new ramets, increase in the number of shoots per rooting point playing a minor role. Individual plants contributed to gap filling very unequally; few of them produced many rooting points. The average size of canes and the proportion of tip-rooting canes were highest in the second year after the gap formation.     

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     

Die Embryologie von Rubus idaeus und von Bastarden zwischen Rubus caesius und Rubus idaeus

Zusammenfassung 1. Die embryologische Untersuchung verschiedener Pflanzen von Rubus idaeus zeigte vollkommen normale Archespor- und Embryosackentwicklung.2. Die drei untersuchten triploiden Bastarde R. caesius × R. idaeus bilden in geringem Maß auch Embryosäcke durch Diplosporie, während die nach meiotischen Teilungen entstandenen Embryosäcke oft degenerieren und wohl immer funktionsuntüchtig sind.3. Da von den beiden Elternpflanzen nur R. caesius Diplosporie zeigt, muß diese Erscheinung durch den R. caesius-Gameten auf den Bastard übertragen worden sein. Dadurch ist erwiesen, daß Diplosporie bei Rubus genetisch bedingt ist.4. Schließlich wird ein hypothetisches Vererbungsschema der Diplosporie diskutiert unter der Annahme, daß sich Diplosporie und Sexualität mehr oder weniger intermediär vererben.Mit 5 Textabbildungen.Herr Prof. Dr. A. Frey-Wyssling, Vorstand des Pflanzenphysiologischen Instituts der E.T.H., hat diese Arbeit durch manche Anregung gefördert und durch die Überlassung der Hilfsmittel des Instituts ermöglicht. Ich bin Herrn Prof. Frey-Wyssling dafür sehr zu Dank verpflichtet.