The occurrence and distribution of isolates of raspberry bushy dwarf virus in England

The distribution of distinct isolates of raspberry bushy dwarf virus (RBDV) in Rubus in England was studied. Isolates similar in Rubus host range to the Scottish type isolate (D200) were largely confined to the old red raspberry (Rubus idaeus) cv. Norfolk Giant, but were also encountered in a single plant of an unidentified raspberry cultivar and in a clump of wild R. idaeus. Outside East Mailing Research Station (EMRS) RBDV isolates with wider Rubus host ranges than that of the type isolate were found only and exclusively in hybrid berries (Loganberry, clones LY59 and L654, and Tayberry) in which infection ranged from < 1% to 100%. The significance of these findings is discussed.     

Association of raspberry bushy dwarf virus with raspberry yellows disease; reaction of Rubus species and cultivars, and the inheritance of resistance.

The agent of raspberry yellows disease is transmitted by grafting but not by aphids and is resistant to thermotherapy. Further studies showed that it is transmitted by inoculation of sap through seed; it is probably transmitted to plants by pollination. Raspberry bushy dwarf virus (RBDV) shares all these attributes and is known to infect all yellows-sensitive raspberry cultivars except Puyallup and Sumner; however, neither of these cultivars has been tested by graft inoculation with RBDV. RBDV commonly infects plants symptomiessly, even those of yellows-sensitive cultivars, but it induced yellows when inoculated either manually to Norfolk Giant raspberry or by grafting to a yellows-sensitive raspberry selection. The evidence suggests that RBDV is the causal agent of yellows disease but that symptom expression is greatiy dependent on genetic and environmental factors. Many red raspberry cultivars are resistant, probably immune, to the type culture of RBDV and this character was shown to be conferred by a single dominant gene designated Bu.     

Host range, properties and purification of raspberry bushy dwarf virus

Raspberry bushy dwarf virus (RBDV) was found in all plants of Lloyd George raspberry with bushy dwarf disease and occurred occasionally in plants of some other cultivars. It was transmitted by inoculation of sap to fifty-five other species in twelve families of flowering plants and infected most of them symptomlessly. It caused systemic symptoms in some species of Amaranthaceae, Chenopodiaceae and Cucurbitaceae, and necrotic local lesions in some Leguminosae. It did not induce bushy dwarf disease when returned to Lloyd George raspberry. Chenopodium quinoa was used for propagating the virus and Vigna cylindrica for local lesion assay. In C. quinoa sap, RBDV lost infectivity when diluted 10^-4, heated for 10 min at 65 °C or stored for 4 days at 22 °C. Preparations made by twice precipitating the virus at pH 4·8 and resuspending it at pH 7·0, followed by ultracentrifugation and exclusion chromatography in columns of 2 % agarose beads, contained isometric particles about 33 nm in diameter, which sedimented as two components, with sedimentation coefficients of 111 and 116S. Only a few particles, all of them disrupted, were seen in preparations mounted in phosphotungstate, but the particles were well preserved in uranyl formate provided that they were first dispersed in a saxlt such as MgCl₂ instead of distilled water. Many particles were oval in outline as though distorted during drying. No serological relationship was detected between RBDV and twenty-four other isometric viruses nor between RBDV and the filamentous virus apple chlorotic leafspot, to which it was previously thought to be related. An isolate of loganberry degeneration virus was serologically indistinguishable from RBDV.     

Further studies on resistance to Leptosphaeria coniothyrium in the red raspberry and related species

Rubus pileatus and its F₁ hybrids with raspberry (R. idaeus) were resistant to cane blight (Leptosphaeriu conioth-yriurn), but little resistance was obtained in subsequent backcross generations apart from a hybrid identified in the second backcross. Two hybrids from backcrossing R. coreanus to raspberry also showed resistance. R. pileatus and its F, hybrids produced hard growth, unlike that of raspberries, which may have been associated with a form of resistance that could not easily be transferred to commercial raspberry cultivars. Some of the genotypes used as parents showed intermediate levels of resistance and it is possible that the highly resistant genotype identified in the second backcross arose from a recombination of genes for resistance. Plants with pubescent canes (gene H) were up to 20% more resistant to mycelial inoculation than those with non-pubescent canes (gene h), and the percentage of machine-harvester inflicted wounds with disease symptoms that resulted from natural infection was also less in genotypes with pubescent canes. Many genotypes with intermediate levels of resistance suffered only limited damage from mycelial inoculation and so there are good prospects for breeding cultivars with an effective resistance, despite the limited value of R. pileatus as a donor species.     

Isolates of raspberry bushy dwarf virus differing in Rubus host range

Isolates of raspberry bushy dwarf virus (RBDV) occurring in the field at East Mailing Research Station (EMRS), and an isolate from raspberry seed imported from the USSR, were found to differ from the Scottish type isolate (D200) of RBDV in that they infected red raspberry cultivars that are resistant, possibly immune, to isolate D200. Of several red raspberry, blackberry and hybrid berry cultivars and EMRS raspberry selections graft-inoculated with these recently discovered RBDV isolates only two raspberry cvs (Haida and Rannaya Sladkaya) and one EMRS selection did not become infected. Differences in the conclusions reached in two previous studies on the inheritance of resistance to RBDV in raspberry can be explained by the use of virus isolates that differed in Rubus host range.     

Molecular detection and characterisation of black raspberry necrosis virus and raspberry bushy dwarf virus isolates in wild raspberries

Virus‐derived small interfering RNAs (siRNAs) were extracted from leaves of wild raspberries (Rubus idaeus) sampled from three different regions in Finland and subjected to deep sequencing. Assembly of the siRNA reads to contigs and their comparison to sequences in databases revealed the presence of the bipartite positive‐sense single‐stranded RNA viruses, raspberry bushy dwarf virus (RBDV, genus Idaeovirus), and black raspberry necrosis virus (BRNV, family Secoviridae) in 19 and 26 samples, respectively, including 15 plants coinfected with both viruses. Coverage with siRNA reads [21 and 22 nucleotides (nt)] was higher in BRNV‐FI (Finland) RNA1 (79%) than RNA2 (45%). In RBDV, the coverage of siRNA reads was 89% and 90% for RNA1 and RNA2, respectively. Average depth of coverage was 1.6–4.9 for BRNV and 16.5–36.5 for RBDV. PCR primers designed for RBDV and BRNV based on the contigs were used for screening wild raspberry and a few cultivated raspberry samples from different regions. Furthermore, the sequences of BRNV RNA1 and RNA2 were determined by amplification and sequencing of overlapping contigs (length 1000–1200 nt) except for the 3′ and 5′ ends of RNA1 and RNA2 covered by primers. RNA1 of the Finnish BRNV isolate (BRNV‐FI) was 80% and 86% identical to BRNV‐NA (USA) and BRNV‐Alyth (UK), respectively, whereas the identity of NA and Alyth was 79%. RNA2 of BRNV‐FI was 84% and 80% identical to BRNV‐NA and BRNV‐Alyth, respectively, whereas NA and Alyth were 82% identical. Hence, the strains detected in Finland differ from those reported in the UK and USA. Our results reveal the presence of BRNV in Finland for the first time. The virus is common in wild raspberries and nearly identical isolates are found in cultivated raspberries as well. The results show that wild raspberries in Finland are commonly infected with RBDV or BRNV or both viruses and thus are likely to serve as reservoirs of RBDV and BRNV for cultivated Rubus spp.     

Further studies on breeding for resistance to Botrytis cinerea in red raspberry canes

A high level of resistance to cane botrytis caused by Botrytis cinerea was transferred from Rubus pileatus to the red raspberry through three generations of backcrossing. The strength of the resistance showed little diminution through these generations and it was therefore thought that a major gene for resistance had been transferred. But discontinuity in the resistance levels observed was detected in plants of only one group of the progenies and so evidence for a major gene is lacking. The results showed the considerable influence of gene H, which confers pubescence, and emphasised that resistance must be separately assessed for pubescent and non-pubescent segregates.     

Natural infection with raspberry bushy dwarf virus (RBDV) of the putatively RBDV-resistant red raspberry cultivar Glen Moy, and the demonstration that it does not contain the RBDV resistance gene, Bu

When released to commerce in 1981, the red raspberry cv. Glen Moy was reported to be immune to the Scottish type isolate of raspberry bushy dwarf virus (RBDV-D200). Field observations of this cultivar in localities where RBDV was prevalent tended to support this claim of its resistance, but in the past 6–10 yr, RBDV infection has been reported in this cultivar in Australasia, USA and in several commercial crops in England. Therefore, experiments were made to investigate the reason(s) for this apparent anomaly using RBDV-infected material, putatively of cv. Glen Moy, from two locations in southern England and one each from Australia, New Zealand (NZ) and the USA. Genetic fingerprinting of genomic DNA from samples of these five RBDV-infected raspberry sources confirmed their identity as cv. Glen Moy. Comparisons of some serological and genomic properties of the five Glen Moy RBDV isolates indicated that, whilst they shared many properties with previously well characterised isolates of this virus, they were distinguishable from them. Characterisation of the isolate from NZ maintained in raspberry showed that it did not have a Rubus host range characteristic of resistance-breaking (RB) isolates, indicating that for this location, and probably also for those of Australia and the USA, RB isolates were not the cause of infection in cv. Glen Moy. When virus-tested plants of cv. Glen Moy and 45 progeny seedlings from the cross between cv. Glen Moy and the RBDV-susceptible cv. Autumn Bliss were graft inoculated with RBDV-D200, all grafted plants became infected indicating that cv. Glen Moy does not contain the RBDV resistance gene, Bu. Possible reasons for the previously reported resistance of cv. Glen Moy to RBDV are discussed.     

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.     

Spread of raspberry bushy dwarf virus by pollination, its association with crumbly fruit, and problems of control

Raspberry bushy dwarf virus (RBDV) was transmitted to raspberry seed both through the pollen and through the ovule and it infected plants pollinated with infected pollen. It did not infect plants prevented from flowering, and transmission through pollen seems to be the only method of spread in the field; in the proximity of infectors, most plants became infected during the first two or three flowering seasons. Plants containing RBDV showed no obvious symptoms, but healthy or infected flowers pollinated with infected pollen produced ‘crumbly’ fruit, containing a high proportion of aborted drupelets. RBDV was difficult to eliminate from infected raspberry by heat therapy. Raspberry cultivars that fail to become infected naturally were also immune to infection by grafting. Use of immune cultivars offers the only method of control and, because infected plants may produce crumbly fruit, future cultivars should if possible possess immunity to RBDV.     

Properties of a resistance-breaking strain of potato virus X

During indexing of a potato germplasm collection from Bolivia, a strain of potato virus X (PVX), X_HB, which failed to cause local lesions in inoculated leaves of Gomphrena globosa was found in 7% of the clones. X_HB was transmitted by inoculation of sap to 56 species from 11 families out of 64 species from 12 families tested. It was best propagated in Nicotiana glutinosa or N. debneyi; Montia perfolia and Petunia hybrida were useful as local lesion hosts. Inoculated leaves of G. globosa plants kept at 10°, 14°, 18°, 22°, or 26 °C after inoculation were always infected symptomlessly. X_HB caused a mild mosaic, systemic chlorotic blotching or symptomless infection in 16 wild potato species and eight Andean potato cultivars, systemic necrotic symptoms in clone A6 and cultivar Mi Peru, and bright yellow leaf markings in cultivar Renacimiento. It caused necrotic local lesions in inoculated leaves of British potato cultivars with the PVX hypersensitivity gene Nb but then invaded the plants systemically without causing further necrosis; with gene Nx systemic invasion occurred but no necrotic symptoms developed. These reactions resemble those of PVX strain group four. X_HB differed from other known strains of PVX in readily infecting PVX-immune clones 44/1016/10, G. 4298.69 and USDA 41956, cultivars Saphir and Saco, and Solanum acaule PI 230554. X_HB had slightly flexuous filamentous particles with a normal length of 516 nm. It was transmitted readily by plant contact and it partially protected G. globosa leaves from infection with X_CP, a group two strain of PVX. Sap from infected N. glutinosa was infective after dilution to 10^--⁶ but not 10^--⁷ after 10 min at 75° but not 80 °C and after 1 yr at 20 °C. X_HB was readily purified from infected N. debneyi leaves by precipitation with polyethylene glycol followed by differential centrifugation. Microprecipitin tests showed that X_HB and X_CP are closely related serologically.     

Properties, relationships and transmission of a strain of raspberry ringspot virus infecting raspberry cultivars immune to the common Scottish strain*

Plants of Lloyd George and Seedling M raspberry (Rubus idaeus L.) were found in eastern Scotland infected with raspberry ringspot (RRV), a virus to which these varieties were previously considered immune. Most RRV isolates from affected plants caused milder symptoms in herbaceous test plants than did the type isolates of the common Scottish and English strains. In graft-transmission tests the Lloyd George strain of RRV infected all the raspberry cultivars tested, including those immune to the common Scottish strain. No consistent differences were found between isolates of the two strains in in vitro properties or serological behaviour. Both strains were transmitted in seed of Stellaria media and in soil containing Longidorus elongatus. Possible reasons why the new strain is uncommon in Scotland are discussed.     

Studies on the occurrence of tomato bushy stunt virus in English rivers

Tomato bushy stunt virus (TBSV) of unknown source was isolated from water of the River Thames, near Oxford. The isolate designated TBSV-T was mechanically transmissible to several tomato (Lycopersicon esculentum) cvs and to other species including Petunia hybrida, pepper (Capsicum annuum). eggplant (Solanum melongena), Nicotiana clevelandii, Chenopodium amaranticolor and C. quinoa in which it caused systemic symptoms. It caused no infection of globe artichoke (Cynara scolymus) or Pelargonium domesticum. The virus was not adsorbed to soil and could be isolated from leachate of soil in which systemically-infected tomato or C. quinoa plants were grown. Tomato plants became infected when grown in soil watered with virus suspensions. TBSV-T was infective after 10 min at 80°C but not at 90°C and when diluted to 10^-5 but not to 10^-6. Purified virus preparations contained C. 30 nm isometric particles. In gel-diffusion serological tests, TBSV-T reacted with homologous anti-serum and with antiserum to petunia asteroid mosaic virus but not to pelargonium leaf curl virus. Seed-borne infection (50–65%) of TBSV was demonstrated in plants grown from seed of symptomlessly-infected tomato fruit. TBSV was isolated from symptomlessly-infected tomato fruit imported from Morocco during October-April 1981. One of the isolates (TBSV-M) was indistinguishable from TBSV-T in host range, symptomatology and serological reactions. TBSV was also found in tomato plants growing extraneously in primary settlement beds at sewage works; such plants having been derived from undigested seeds in sewage. Because of its ‘alimentary-resistance’ in man, it is possible that one ecological route whereby TBSV enters rivers is by man's consumption of TBSV-infected tomatoes and eventual sewage dispersal into rivers.     

Studies on the inheritance in the red raspberry of immunities from three nematode-borne viruses

Lethal or chronic diseases of the raspberry caused by the nematodeborne viruses raspberry ringspot, arabis mosaic and tomato black ring can cause serious reductions in the productivity of raspberry plantations, but the existence of clear-cut immunities from these diseases provides a basis for control through plant breeding. The inheritance of these immunities was studied by means of graft tests on families of raspberry seedlings. Immunity from each virus was found to be dominant to susceptibility, but there was evidence that more than one gene was concerned in each case: while it was not possible to decide whether the second gene was a dominant complementary or a linked recessive affecting the viability of the immune segregates, the frequent occurrence in the raspberry of aberrant segregation ratios due to such lethal genes makes the latter explanation the more probable. There was also evidence of linkage between the genes for the three immunities. The experiment confirmed the practicability of breeding to incorporate genes for immunities from these three viruses into new raspberry varieties.     

Further studies on the effect of resistance to Amphorophora idaei in raspberry (Rubus idaeus) on the spread of aphid-borne viruses

The rate of spread of viruses transmitted by the aphid Amphorophora idaei into genotypes of raspberry differing in resistance to infestation by A. idaei was studied in a field experiment which exposed plants to large numbers of infective aphids. Under these conditions, genotypes that are readily colonised by A. idaei were totally infected with virus after two to three growing seasons, whereas genotypes with a high degree of resistance were substantially free of virus after four growing seasons but 56% of plants were infected after seven seasons. Genotypes with intermediate resistance were also substantially free of virus after three seasons but 76% of plants were virus infected after seven seasons. The effectiveness of resistance to A. idaei in raspberry in restricting spread of viruses transmitted by this aphid is discussed.     

Biology of the European large raspberry aphid (Amphorophora idaei): its role in virus transmission and resistance breakdown in red raspberry

1 The European large raspberry aphid Amphorophora idaei Börner is the most important vector of viral diseases afflicting commercially grown red raspberry ( Rubus idaeus L.) in Northern Europe, with European raspberry production amounting to 416 000 tonnes per annum. This review synthesizes existing knowledge on its biology and interactions with other organisms, including its host plant and the viral pathogens it vectors.
2 Information about trophic interactions with other insect herbivores and natural enemies is reviewed. Vine weevils Otiorhynchus sulcatus compromise aphid resistance in some raspberry cultivars, increasing A.   idaei abundance by 80%. Parasitoids show mixed success in parasitizing A.   idaei , although Aphidius ervi attack rates more than doubled when A.   idaei fed on a partially susceptible raspberry cultivar, compared with a resistant variety. These findings are discussed in the context of potential biological control as part of an integrated pest and disease management framework.
3  Amphorophora idaei transmits four known viruses: Black raspberry necrosis virus, Raspberry leaf mottle virus, Raspberry leaf spot virus and Rubus yellow net virus , with A.   idaei taking as little as 2 min to transmit some viruses.
4 Existing control strategies, including resistant cultivars, insecticides and eradication of disease from parent plants, are described. In particular, strong selection pressures have resulted in A .  idaei overcoming genetic resistance in many raspberry cultivars and most insecticides are now ineffective.
5 Future directions for the sustained control of A.   idaei are suggested, taking into consideration the possible effects of climate change and also changes in agronomic practices in U.K. agriculture.     

Variation in reaction to barley yellow dwarf virus in ryegrass and its inheritance

Genotypes of Italian and perennial ryegrass differed greatly in their reaction to infection with barley yellow dwarf virus (BYDV). One genotype of perennial ryegrass appeared unaffected, whilst the yield and height of other genotypes were reduced. Progeny of the Italian ryegrass genotypes showed a similar degree of variation. Tolerance, assessed by the effect of the virus on yield, was inherited in a wholly additive manner. Symptoms were rare and their severity was under both additive and non-additive genetic control and strongly correlated with increasing severity of reaction. The utilization of the variation in BYDV reaction in breeding programmes may prove difficult because plants with the highest performance tended to suffer most from BYDV infection, and half-sib progeny ranked in a different order of tolerance in the glasshouse from that of their respective parent genotypes in the field. It is suggested that the observed variation in BYDV response results from a readjustment in the balance of genotypically controlled and environmentally conditioned variation in plant growth.