Search

Проаналізовано результати багаторічних досліджень вірусу некротичного пожовтіння жилок буряку, який спричиняє хворобу ризоманію, що є карантинним об’єктом. Описано молекулярно-біологічні властивості, особливості його геному та залежність патогенезу від генетичних варіацій вірусу. Висвітлено різноманіття патотипів вірусу і циркуляцію хвороби у світі та в Україні зокрема, а також способи її поширення. Проведено аналіз природних генів стійкості рослин цукрового буряку до ризоманії, The results of many years of research of beet necrotic yellow vein virus, which causes rhizomania, and is a quarantine object, are analyzed. The molecular-biological properties, features of its genome and dependence of pathogenesis on genetic variations of the virus are described. Diversity of virus pathotypes and disease circulation in the world and in Ukraine and ways of its spread are shown. Natural genes of sugar beet resistance to rhizomania were also analyzed., Проанализированы результаты многолетних исследований вируса некротического пожелтения жилок свеклы, вызывающего болезнь ризоманию, которая является карантинным объектом. Описаны молекулярно-биологические свойства, особенности его генома и зависимость патогенеза от генетических вариаций вируса. Показано разнообразие патотипив вируса и циркуляции болезни в мире и в Украине, способы ее распространения. Также проведен анализ природных генов устойчивости сахарной свеклы к ризомании.

Beet necrotic yellow vein virus (BNYVV) is causal agent of rhizomania disease, which is the most devastating viral disease in sugar beet production leading to a dramatic reduction in beet yield and sugar content. The virus is transmitted by the ubiquitous distributed soil-borne plasmodiophoromycete Polymyxa betae that infects the root tissue of young sugar beet plants. Rz1 is the major resistance gene widely used in most sugar beet varieties to control BNYVV. The strong selection pressure on the virus population promoted the development of strains that can overcome Rz1 resistance. Resistance-breaking has been associated with mutations in the RNA3-encoded pathogenicity factor P25 at amino acid positions 67–70 (tetrad) as well as with the presence of an additional RNA component (RNA5). However, respective studies investigating the resistance-breaking mechanism by a reverse genetic system are rather scarce. Therefore, we studied Rz1 resistance-breaking in sugar beet using a recently developed infectious clone of BNYVV A-type. A vector free infection system for the inoculation of young sugar beet seedlings was established. This assay allowed a clear separation between a susceptible and a Rz1 resistant genotype by measuring the virus content in lateral roots at 52 dpi. However, mechanical inoculation of sugar beet leaves led to the occurrence of genotype independent local lesions, suggesting that Rz1 mediates a root specific resistance toward BNYVV that is not active in leaves. Mutation analysis demonstrated that different motifs within the P25 tetrad enable increased virus replication in roots of the resistant genotype. The resistance-breaking ability was further confirmed by the visualization of BNYVV in lateral roots and leaves using a fluorescent-labeled complementary DNA clone of RNA2. Apart from that, reassortment experiments evidenced that RNA5 enables Rz1 resistance-breaking independent of the P25 tetrad motif. Finally, we could identify a new resistance-breaking mutation, which was selected by high-throughput sequencing of a clonal virus population after one host passage in a resistant genotype. Our results demonstrate the feasibility of the reverse genetic system for resistance-breaking analysis and illustrates the genome plasticity of BNYVV allowing the virus to adapt rapidly to sugar beet resistance traits.

Beet necrotic yellow vein virus (BNYVV) is the causal agent of rhizomania, a disease of global importance to the sugar beet industry. The most widely implemented resistance gene to rhizomania to date is Rz1, but resistance has been circumvented by resistance-breaking (RB) isolates worldwide. In an effort to gain greater understanding of the distribution of BNYVV and the nature of RB isolates in Minnesota and eastern North Dakota, sugar beet plants were grown in 594 soil samples obtained from production fields and subsequently were analyzed for the presence of BNYVV as well as coding variability in the viral P25 gene, the gene previously implicated in the RB pathotype. Baiting of virus from the soil with sugar beet varieties possessing no known resistance to rhizomania resulted in a disease incidence level of 10.6% in the region examined. Parallel baiting analysis of sugar beet genotypes possessing Rz1, the more recently introgressed Rz2, and with the combination of Rz1 + Rz2 resulted in a disease incidence level of 4.2, 1.0, and 0.8%, respectively. Virus sequences recovered from sugar beet bait plants possessing resistance genes Rz1 and/or Rz2 exhibited reduced genetic diversity in the P25 gene relative to those recovered from the susceptible genotype while confirming the hypervariable nature of the coding for amino acids (AAs) at position 67 and 68 in the P25 protein. In contrast to previous reports, we did not find an association between any one specific AA signature at these positions and the ability to circumvent Rz1-mediated resistance. The data document ongoing virulence development in BNYVV populations to previously resistant varieties and provide a baseline for the analysis of genetic change in the virus population that may accompany the implementation of new resistance genes to manage rhizomania.

Beet soilborne mosaic virus (BSBMV) is a rigid rod-shaped virus transmitted by Polymyxa betae. Particles were 19 nm wide and ranged from 50 to over 400 nm, but no consistent modal lengths could be determined. Nucleic acids extracted from virions were polyadenylated and typically separated into three or four discrete bands of variable size by agarose-formaldehyde gel electrophoresis. RNA 1 and 2, the largest of the RNAs, consistently averaged 6.7 and 4.6 kb, respectively. The sizes and number of smaller RNA species were variable. The molecular mass of the capsid protein of BSBMV was estimated to be 22.5 kDa. In Northern blots, probes specific to the 3′ end of individual beet necrotic yellow vein virus (BNYVV) RNAs 1–4 hybridized strongly with the corresponding BNYVV RNA species and weakly with BSBMV RNAs 1, 2, and 4. Probes specific to the 5′ end of BNYVV RNAs 1–4 hybridized with BNYVV but not with BSBMV. No cross-reaction between BNYVV and BSBMV was detected in Western blots. In greenhouse studies, root weights of BSBMV-infected plants were significantly lower than mock-inoculated controls but greater than root weights from plants infected with BNYVV. Results of serological, hybridization, and virulence experiments indicate that BSBMV is distinct from BNYVV. However, host range, capsid size, and the number, size, and polyadenylation of its RNAs indicate that BSBMV more closely resembles BNYVV than it does other members of the genus Furovirus.

A high degree of reciprocal cross-protection occurred in sugar beets between beet soilborne mosaic virus (BSBMV) and beet necrotic yellow vein virus (BNYVV) in greenhouse experiments. The incidence of cross-protection depended on the interval between inoculations with protecting and challenging virus; longer inoculation intervals enhanced the incidence of cross-protection. Cross-protection was most effective when the inoculation interval was between 5 and 10 days, a period during which virus accumulated to a maximum level in plants singly infected with BSBMV or BNYVV. Results obtained by enzyme-linked immunosorbent assay (ELISA) and Western blotting were consistent and indicated that cross-protection affected viral capsid protein accumulation. RNA of both protecting and challenging viruses was detected in doubly infected plants by using reverse transcription-polymerase chain reaction (RT-PCR), indicating that RNA of the challenge virus was present in protected plants, but capsid protein was undetected by serological tests. BSBMV is a closely related but serologically distinct virus that provides an excellent system for studies of cross-protection with BNYVV.

Beet necrotic yellow vein virus (BNYVV) causes rhizomania in beets. BNYVV is widespread in Europe and can be divided into an A and B group based on reverse transcription-polymerase chain reaction (RT-PCR) restriction fragment length polymorphism (RFLP) analysis (1). The vector of BNYVV, Polymyxa betae, is widespread in Denmark. However, surveys for BNYVV have regularly been carried out in Denmark since 1985, but it has not been detected until now. In the autumn of 2000, attention was drawn to a sugar beet field in the southeastern part of Denmark containing plants with unusually low sugar content. Typical BNYVV symptoms of bearded roots were observed on all plants sampled in the field. Root samples of 12 plants all tested positive for the presence of BNYVV by ELISA using antiserum from Adgen, Loewe, and Aschersleben, respectively. Immunosorbent electron microscopy with antiserum to BNYVV coat protein revealed decorated rod-shaped particles of lengths typical for BNYVV. Pulverized infected root tissue was used to mechanically inoculate Chenopodium quinoa and resulted in formation of few local chlorotic lesions. RNA was extracted from symptomatic C. quinoa and subjected to RT-PCR analysis using two primer sets specific for RNA 2 of BNYVV (1) and bands of the expected sizes were observed with both. The nucleotide sequence of a part of the PCR products corresponding to the N-terminus of the coat protein was determined and showed 99.5% identity to sequences derived from A group isolates (1). Finally, a bait-plant test was carried out, where sugar beet seeds were sown in 20 soil samples collected from different locations within the infested field. After 8 weeks of growth, all samples tested positive for the presence of BNYVV by ELISA, indicating that the entire field is infested and that viruliferous P. betae must have been present in several years. To our knowledge, this is the first record of BNYVV in Denmark. Reference: (1) M. Kruse et al. J. Gen. Virol. 75:1835, 1994.

The effects of cultivar mixtures and two irrigation frequency treatments were evaluated over two seasons for their impact on a complex of sugar beet root diseases in three fields infested with the fungal pathogens Aphanomyces cochliodes, Fusarium oxysporum f. sp. radicis-betae, Rhizoctonia solani, and the viral pathogen Beet necrotic yellow vein virus (BNYVV). Irrigations after emergence consisted of two or five (two 1994 studies) and three or six (1995 study) applications of water for dry and wet treatments, respectively. Cultivar treatments included MH9155, HH67, Ranger, Rhizosen, and four combinations of these same cultivars. Disease progress was monitored through destructive sampling of plants exhibiting foliar symptoms typical of root disease during the season. At harvest, data on root and sucrose yields, sucrose percentage, and a root disease index were collected. No significant irrigation × cultivar treatment interactions were observed. Few significant differences were observed between irrigation treatments involving measured yield components. Reduced irrigations however, resulted in significantly lower disease incidence in all three repeated experiments when cultivar treatments were combined. No added benefits were observed for increasing yield or decreasing root disease by planting mixed cultivars, compared to the same cultivars planted individually. Several regionally adapted cultivars performed as well or better than mixtures under the unusually high levels of disease pressure in test fields. When few alternative options are available, sugar beet growers may still benefit from reducing irrigations, and growing locally adapted cultivars in soils severely infested with root pathogens.

Rhizomania, caused by Beet necrotic yellow vein virus (BNYVV) and vectored by the soilborne fungus Polymyxa betae Keskin, is one of the most economically damaging diseases affecting sugar beet (Beta vulgaris L.). The virus likely originated in Europe and was first identified in California in 1983 (1). It has since spread among American sugar beet production regions in spite of vigorous sanitation efforts, quarantine, and disease monitoring (3). In the fall of 2002, mature sugar beet plants exhibiting typical rhizomania root symptoms, including proliferation of hairy roots, vascular discoloration, and some root constriction (2) were found in several fields scattered throughout central and eastern Michigan. Symptomatic beets were from numerous cultivars, all susceptible to rhizomania. Two to five sugar beet root samples were collected from each field and sent to the USDA-ARS in Salinas, CA for analysis. Hairy root tissue from symptomatic plants was used for mechanical inoculation of indicator plants. Mechanical inoculation produced necrotic lesions on Chenopodium quinoa and systemic infection of Beta vulgaris ssp. macrocarpa, both typical of BNYVV and identical to control inoculations with BNYVV. Symptomatic sugar beet roots were washed and tested using double antibody sandwich-enzyme linked immunosorbent assay (DAS-ELISA) for the presence of BNYVV using standard procedures and antiserum specific for BNYVV (3). Sugar beet roots were tested individually, and samples were considered positive when absorbance values were at least three times those of greenhouse-grown healthy sugar beet controls. Samples were tested from 16 fields, with 10 confirmed positive for BNYVV. Positive samples had mean absorbance values ranging from 0.341 to 1.631 (A405nm) after 30 min. The mean healthy control value was 0.097. Fields were considered positive if one beet tested positive for BNYVV, but in most cases, all beets tested from a field were uniformly positive or uniformly negative. In addition, soil-baiting experiments were conducted on seven of the fields. Sugar beet seedlings were grown in soil mixed with equal parts of sand for 6 weeks and were subsequently tested using DAS-ELISA for BNYVV. Results matched those of the root sampling. Fields testing positive for BNYVV were widely dispersed within a 100 square mile (160 km2) area including portions of Gratiot, Saginaw, Tuscola, and Sanilac counties in the central and eastern portions of the Lower Peninsula of Michigan. The confirmation of rhizomania in sugar beet from the Great Lakes Region marks the last major American sugar beet production region to be diagnosed with rhizomania disease, nearly 20 years after its discovery in California (1). In 2002, there were approximately 185,000 acres (approximately 75,00 ha) of sugar beet grown in the Great Lakes Region, (Michigan, Ohio, and southern Ontario, Canada). The wide geographic distribution of infested fields within the Michigan growing area suggests the entire region should monitor for symptoms, increase rotation to nonhost crops, and consider planting rhizomania resistant sugar beet cultivars to infested fields. References:(1) J. E. Duffus et al. Plant Dis. 68:251, 1984. (2) J. E. Duffus. Rhizomania. Pages 29–30 in: Compendium of Beet Diseases and Insects, E. D. Whitney and J. E. Duffus eds. The American Phytopathological Society, St. Paul, MN, 1986. (3) G. C. Wisler et al. Plant Dis. 83:864, 1999.

Rhizomania is an important virus disease of sugar beet and is caused by Beet necrotic yellow vein virus (BNYVV). During 2002-03, several sugar beet fields with cultivars partially resistant to BNYVV grown in the Imperial Valley of California were observed with severe rhizomania symptoms, suggesting that resistance conditioned by Rz1 had been compromised. Soil testing with sugar beet baiting plants followed by enzyme-linked immunosorbent assay (ELISA) was used to diagnose virus infection. Resistant varieties grown in BNYVV-infested soil from Salinas, CA, were ELISA-negative. In contrast, when grown in BNYVV-infested soil collected from the Imperial Valley, CA, all resistant varieties became infected and tested positive by ELISA. Based on host reaction, eight distinct BNYVV isolates have been identified from Imperial Valley soil (IV-BNYVV) by single local lesion isolation. Reverse transcription-polymerase chain reaction (RT-PCR) assays showed that the eight IV-BNYVV isolates did not contain RNA-5. Singlestrand conformation polymorphism banding patterns for the IV-BNYVV isolates were identical to A-type and different from P-type. Sequence alignments of PCR products from BNYVV RNA-1 near the 3′ end of IV-BNYVV isolates revealed that both IV-BNYVV and Salinas BNYVV isolates were similar to A-type and different from B-type. Our results suggest that the resistancebreaking BNYVV isolates from Imperial Valley likely evolved from existing A-type isolates.

During the 2001 growing season, a survey was conducted to determine the incidence of Beet necrotic yellow vein virus (BNYVV), Beet soilborne virus (BSBV), and Beet virus Q (BVQ) in Iran. A total of 2,816 random and 76 samples with rhizomania were collected from 131 fields in the main sugar beet cultivation areas of 13 provinces in Iran. All samples were tested using a tissue-blot immunoassay (TBIA) with commercial BNYVV (As-0799.1/CG6-F4), BSBV (As-0576.1), and BSBV/BVQ (As-0576.2) antisera provided by S. Winter (DSMZ, Braunschweig, Germany). For randomly collected samples, the highest incidence of virus infection was found for BNYVV (52.3%), followed by BSBV (9.5%) and BVQ (1.5%). Co-infection of BNYVV with BSBV or BVQ was 6.6% and 0.9%, respectively. Infection with both BSBV and BVQ was found in 16 (0.6%) samples. In addition, 0.4% (12) of the samples was infected with all three viruses. Our results indicated the presence of BVQ in samples from 10 fields located in Azarbayejan-e-gharbi, Esfahan, Fars, Kermanshah, Khorasan, Lorestan, and Semnan provinces of Iran, with or without rhizomania-like symptoms. The presence of viruses was confirmed using reverse transcription-polymerase chain reaction (RT-PCR) of RNA from 81, 19, and 14 root samples with positive reaction in TBIA to BNYVV, BSBV, and BVQ, respectively, with previously described primers (3,4). The primers specifically amplified fragments of 501 bp, 602 bp, 399 bp, and 291 bp of the BNYVV RNAs 1 and 4, BSBV RNA-2, and BVQ RNA-1, respectively. Our results indicated that the samples tested were also positive using RT-PCR. The putative vector for BNYVV, BSBV, and BVQ, Polymyxa betae, was also detected in 161 samples (from 127 fields) by amplification of a 170-bp fragment of the P. betae repetitive EcoRI-like fragments using previously described primers (4). RT-PCR products from 72 BNYVV-positive sugar beet root samples from 58 fields that also gave positive reactions in TBIA were analyzed using single-strand conformation polymorphism (SSCP) as previously described with extracts from root beards of the susceptible sugar beet cvs. OPUS and IC1 grown in the soils infested with BNYVV types A and B (provided by A. Meunier, Unite de Phytopathologie-UCL-AGRO-BAPA, Louvain-la-Neuve, Belgium) as positive controls (3). The patterns obtained with SSCP were uniform and showed widespread occurrence of BNYVV type A in almost all provinces surveyed. The fragments obtained for BNYVV RNAs 1 and 4 of an isolate from Qazvin (BNQ1) were sequenced (GenBank Accession Nos. AY703452 and AY703455) and compared with other sequences available in GenBank using Clustal W, which revealed 99.3 and 99.6% identity with the Japanese S (D84410) and Italian type A (AF197552) isolates, respectively. The economic importance of BVQ and its interactions with other sugar beet soilborne viruses remains a matter of debate. BNYVV and BSBV have been previously reported from Iran (1,2). To our knowledge, this is the first report of the natural occurrence of BVQ in sugar beets in Iran. References: (1) Sh. Farzadfar et al. Plant Dis. 86:187, 2002. (2) K. Izadpanah et al. Iran. J. Plant Pathol. 32:155, 1996. (3) R. Koenig et al. J. Gen. Virol. 76:2051, 1995. (4) A. Meunier et al. Appl. Environ. Microbiol. 69:2356, 2003.

Beet necrotic yellow vein virus (BNYVV), the casual agent of rhizomania disease, was identified in sugar beet plants from several fields in the Wielkopolska Region of Poland (1). In greenhouse studies, sugar beets were grown in the soil from one of these fields to bait soilborne viruses. Of 200 sugar beet plants, three developed symptoms of vein clearing, vein banding, and mosaic. Crude sap from symptomatic plants was used for mechanical inoculation of various plants species. In Chenopodium quinoa, C. amaranticolor, and Tetragonia expansa only local lesions were observed. Electron microscope examination of negatively stained leaf-dip preparations from symptomatic sugar beet plants showed a mixture of rod-shape particles from 70 to 400 nm long. Using double-antibody sandwich enzyme-linked immunosorbent assay tests, two symptomatic sugar beet plants gave positive reactions with antiserum against BNYVV (Bio-Rad, Hercules, CA) and a third plant gave a positive reaction with antisera against BNYVV and Beet soilborne virus (BSBV). Total RNA was extracted from roots and leaves of the symptomatic plants and used in a multiplex reverse transcription-polymerase chain reaction (mRT-PCR) assay. Specific primers were designed to amplify a fragment of the RNA1 for BSBV and RNA2 for BNYVV and Beet virus Q (BVQ) (2). Two mRT-PCR products amplified with the primers specific to BNYVV and BSBV were obtained and sequenced. A 274-nt amplicon sequence (GenBank Accession No. DQ012156) had 98% nucleotide sequence identity with the German BNYVV isolate F75 (GenBank Accession No. AF19754) and a 376-nt amplicon sequence (GenBank Accession No. AY999690) had 98% nucleotide and 98% amino acid sequence identity with the German BSBV isolate (GenBank Accession No. Z97873). The Polish BSBV isolate had 88% nucleotide and 62% amino acid sequence identity with BVQ, another pomovirus (GenBank Accession No. AJ 223596 formerly known as serotype Wierthe of BSBV (2). In 2005, mRT-PCR was used on samples collected from two fields of the Wielkopolska Region. Of 15 tested sugar beet plants, 12 gave positive reactions with primers specific for BSBV and nine with primers specific to BNYVV. To our knowledge, this is first report of BSBV in Poland. In Europe, BSBV was previously reported in England, the Netherlands, Belgium, Sweden, Germany, France, and Finland (2,3). References: (1) M. Jezewska and J. Piszczek. Phytopathol. Polonica, 21:165, 2001. (2) A. Maunier et al. Appl. Environ. Microbiol. 69:2356, 2003. (3) C. M. Rush and G. B. Heidel. Plant Dis. 79:868, 1995.

Beet necrotic yellow vein virus (BNYVV) is the type member of the genus Benyvirus and the causal agent of rhizomania disease of sugar beet. Since 1999, BNYVV is becoming one of the most important viruses of sugar beet in Poland. BNYVV is represented by three types, frequently A and B and rarely P, however, in Poland, type A only was recorded (2). In 2005, a survey was conducted to determine the incidence of types A and B of BNYVV in Poland. Thirty samples of sugar beet plants with rhizomania-like symptoms were collected from six fields in the western, southern, and northern areas of Poland. All samples were analyzed by double antibody sandwich-enzyme linked immunosorbent assay (DAS-ELISA) with commercial antiserum (Bio-Rad, Hercules, CA). Infection of BNYVV was found in 26 samples. The presence of the virus in these samples was confirmed by reverse transcription-polymerase chain reaction (RT-PCR). The total RNA extracted from sugar beet was tested with specific primers designed to amplify a fragment of the RNA2 for BNYVV (1). Multiplex (m) RT-PCR with two sets of primers, rhizo AF/rhizo AR and rhizo BF/rhizo BR (2), was used to distinguish A and B types of BNYVV. Two obtained mRT-PCR products were sequenced and compared with other sequences available in GenBank. A 121-nt amplicon sequence (GenBank Accession No. DQ 228872) had 100% nucleotide and amino acid sequence identity with all BNYVV type B sequences (e.g., Stourton-GenBank Accession No. AY682695) (2). A 171-nt amplicon sequence (GenBank Accession No. DQ228871) had 100% nucleotide and amino acid sequence identity with all BNYVV type A sequences (e.g., Ravenna-GenBank Accession No. AY 654282) (2). Type A was detected in 23 BNYVV-infected sugar beet plants from five fields located in western and southern Poland while type B was found in three samples from one field in northern Poland. To my knowledge, this is the first report of the natural occurrence of BNYVV type B in Poland. References: (1) A. Maunier et al. Appl. Environ. Microbiol. 69:2356, 2003. (2) C. Ratti et al. J. Virol. Methods 124:41, 2005.

In October of 2005, sugar beet (Beta vulgaris L.) plants exhibiting symptoms of rhizomania caused by Beet necrotic yellow vein virus (BNYVV) (3) were observed in a production field near Greeley, CO. The roots of seven plants exhibiting moderate to severe symptoms characteristic of this disease were tested using double-antibody sandwich enzyme-linked immunosorbent assay with anti-BNYVV antiserum from rabbits. Of these, only two roots exhibiting the mildest symptoms tested positive for BNYVV (all roots tested negative for the presence of the related Beet soilborne mosaic virus (BSBMV). ‘Hairy’ lateral roots characteristic of the disease were combined from the remaining five roots, ground in phosphate buffer, and the supernatant from the suspension was mechanically applied to leaves of Chenopodium quinoa in an effort to isolate an infectious agent. Five days postinoculation (dpi), yellow lesions with necrotic centers were visible on inoculated leaves, well in advance of those typically observed for BNYVV or BSBMV. Lesions exhibiting a similar rate of development on C. quinoa subsequently were induced from extracts of root vascular tissue prepared from four of seven additional beet roots tested from this location. Transfer of the infection from the C. quinoa lesions to 32 healthy C. quinoa and 10 sugar beet plants (hybrid ACH9369; American Crystal Sugar Co., Moorhead MN) resulted in 100% infection. Inoculated leaves of C. quinoa exhibited a high density of necrotic local lesions within 3 dpi, whereas inoculated leaves of sugar beet exhibited pinpoint, necrotic to diffuse, chlorotic local lesions evident by 5 dpi. Electron microscopic examination of fixed, ultra-thin sections of symptomatic C. quinoa leaf tissue revealed aggregates of virus-like particles of icosahedral symmetry within the cell cytoplasm. Following a virus minipreparation procedure, nucleic acid extracted from the partially purified virus was found to be single-stranded RNA by ribonuclease digestion and alone was infectious when inoculated to C. quinoa leaves. The apparently monopartite RNA genome was 3.5 kb long and a candidate for the single coat protein (CP) had a mass of ˜25 kDa. The sole reference set found in the literature for a virus naturally occurring on sugar beet with similar characteristics was that for Beet black scorch virus (BBSV), a virus recently accepted by the ICTV into the genus Necrovirus within the family Tombusviridae (2). Prior to this communication, BBSV has only been reported in China where it was first documented affecting sugar beet in the late 1980s (1). Using the published sequence of BBSV (Genbank Accession No. AY626780), DNA primers directed to the 3′ half of the BBSV genome were used in reverse transcription-polymerase chain reaction to produce an amplicon from the unknown virus. Sequencing the amplicon revealed 88.8% nucleotide sequence identity to the BBSV CP gene and 97% amino acid sequence identity to the predicted CP gene product. Combined, the nucleotide sequence and physical characteristics confirm the presence of BBSV in a U.S. sugarbeet field for the first time. To our knowledge, this is the first report of the occurrence of BBSV outside of China. References: (1) Y. Cao et al. Arch. Virol. 147:2431, 2002. (2) C. M. Fauquet et al. Eighth Report of the International Committee on the Taxonomy of Viruses. Academic Press, New York, 2005. (3) C. M. Rush. Ann. Rev. Phytopathol. 41:567, 2003.

During storage of sugar beet, respiration and rots consume sucrose and produce invert sugar. Diseases that occur in the field can affect the magnitude of these losses. This research examines the storage of roots with rhizomania (caused by Beet necrotic yellow vein virus) and the effectiveness of rhizomania-resistant hybrids in reducing postharvest losses. Roots of susceptible hybrids from sites with rhizomania had respiration rates 30 days after harvest (DAH) that ranged from 0.68 to 2.79 mg of CO2 kg–1 h–1 higher than roots of the resistant hybrids. This difference ranged from 2.60 to 13.88 mg of CO2 kg–1 h–1 120 DAH. Roots of resistant hybrids from sites with rhizomania had 18 kg more sucrose per ton than roots from susceptible hybrids 30 DAH, with this difference increasing to 55 kg Mg–1 120 DAH. The invert sugar concentration of susceptible hybrids from sites with rhizomania ranged from 8.38 to 287 g per 100 g of sucrose higher than that for resistant hybrids 120 DAH. In contrast, differences between susceptible and resistant hybrids in respiration rate, sucrose loss, and invert sugar concentration in the absence of rhizomania were relatively small. Storage losses due to rhizomania can be minimized by planting resistant hybrids and processing roots from fields with rhizomania soon after harvest.

In a survey of soilborne viruses infecting sugar beet in central Spain, Beet black scorch virus (BBSV) was detected in field grown sugar beets with symptoms of rhizomania disease. BBSV was found in all analyzed sugar beet producing regions from central Spain, as well as in bait plants grown in soils with a history of rhizomania from several Western European countries, thereby constituting the first report of BBSV in Europe. BBSV was transferred to Chenopodium quinoa, where it caused chlorotic local lesions from which virus particles were purified. The nucleotide sequence of the 3′-untranslated region of the genomic RNA was determined for 13 European isolates, and sequences were highly similar to those reported for Chinese and U.S. isolates. Sequence comparisons revealed three clusters of sequences, one including most European isolates, one including one European and two Chinese isolates, and the third including the U.S. isolate. BBSV was detected in a number of samples with rhizomania symptoms in which Beet necrotic yellow vein virus went undetected. However, its role in rhizomania disease in Europe, if any, remains to be established.

To reduce storage losses and improve resistance to rhizomania caused by Beet necrotic yellow vein virus (BNYVV), studies were initiated to establish a storage cultivar selection program. In 2006 and 2007, 30 or more commercial sugar beet (Beta vulgaris) cultivars were grown in soil naturally infested with BNYVV. At harvest, two root samples from each plot were collected and used to establish percent sugar. Additional samples were placed on top of an indoor pile (set point 1.7°C) and inside an outdoor pile in a randomized complete block design with four replications. After 142 and 159 days in indoor storage, sucrose reduction ranged from 13 to 90% in 2007 and 57 to 100% in 2008. Outdoor storage sucrose reduction ranged from 13 to 32% in 2007 and 28 to 60% in 2008. An average of 31 and 45% of the root surface was covered with fungal growth in 2007 and 2008, respectively. Cultivars that retained the most sucrose had resistance to BNYVV and the least fungal growth and weight loss. Indoor storage with BNYVV-infested roots allowed for the most consistent cultivar separation and will potentially lead to selection of cultivars for improved storability and rhizomania resistance.

Beet necrotic yellow vein virus (BNYVV) is an economically important pathogen of sugar beet (Beta vulgaris var. saccharifera) in several European, and Asian countries and in the United States (3). The virus is transmitted by the soil-inhabiting plasmodiophorid Polymyxa betae and causes the rhizomania disease of sugar beet. In November 2012, plants of B. vulgaris subsp. vulgaris cv. Boro (red table beet) exhibiting mainly severe characteristic root symptom of rhizomania were found in a commercial field located in the municipality of São José do Rio Pardo, State of São Paulo, Brazil. No characteristic virus-inducing foliar symptom was observed on diseased plants. The incidence of diseased plants was around 70% in the two visited crops. As the hairy root symptom is indicative of infection by BNYVV, the present study aimed to detect and identify this virus associated with the diseased plants. Preliminary leaf dip analysis by transmission electron microscopy revealed the presence of very few benyvirus-like particles. Total RNA was extracted from roots of three symptomatic plants and one asymptomatic plant according to Toth et al. (3). One-step reverse-transcription–polymerase chain reaction (RT-PCR) was performed as described by Morris et al. (2) with primers that amplify part of the coat protein gene at RNA2. The initial assumption that the hairy root symptom was associated with BNYVV infection was confirmed by the amplification of a fragment of ~500 bp from all three symptomatic samples. No amplicon was obtained from the asymptomatic control plant. Amplicons were directly sequenced, and the consensus nucleotide and deduced amino acid sequences showed 100% identity. The nucleotide sequence for one amplicon (Accession No. KM433683) was compared with other sequences deposited in GenBank. The nucleotide (468 nt) and deduced amino acid (156 aa) sequences shared 93 to 100 and 97 to 99% identity, respectively with the corresponding nucleotide and amino acid sequences for other isolates of type A of BNYVV. The virus was transmitted to three of 10 red table beet plants inoculated with contaminated soil, and infection was confirmed by nested RT-PCR, as described by Morris et al. (1), and nucleotide sequencing. This is the first report on the occurrence of BNYVV in Brazil, which certainly will affect the yield of red table beet in the producing region. Therefore, mapping of the occurrence of BNYVV in red table beet-producing areas in Brazil for containment of the spread of the virus is urgent. In the meantime, precautions should be taken to control the movement of contaminated soil and beet roots, carrots, or any vegetable grown on infested land that might introduce the virus to still virus-free regions. References: (1) J. Morris et al. J. Virol. Methods 95:163, 2001. (2) D. D. Sutic et al. Handbook of Plant Virus Diseases. CRC Press, Boca Raton, Florida, 1999. (3) I. K. Toth et al. Methods for the Detection and Quantification of Erwinia carotovora subsp. atroseptica (Pectobacterium carotovorum subsb. atrosepticum) on Potatoes: A Laboratory Manual. Scottish Crop Research Institute, Dundee, Scotland, 2002.

Root rots in sugar beet storage can lead to multimillion dollar losses because of reduced sucrose recovery. Thus, studies were conducted to establish additional fungicide treatments for sugar beet storage and a greater understanding of the fungi involved in the sugar beet storage rot complex in Idaho. A water control treatment and three fungicides (Mertect [product at 0.065 ml/kg of roots; 42.3% thiabendazole {vol/vol}], Propulse [product at 0.049 ml/kg of roots; 17.4% fluopyram and 17.4% prothioconazole {vol/vol}], and Stadium [product at 0.13 ml/kg of roots; 12.51% azoxystrobin, 12.51% fludioxonil, and 9.76% difenoconozole {vol/vol}]) were investigated for the ability to control fungal rots of sugar beet roots held up to 148 days in storage during the 2012 and 2013 storage seasons. At the end of September into October, roots were harvested weekly for 5 weeks from each of two sugar beet fields in Idaho, treated with the appropriate fungicide, and placed on top of a commercial indoor sugar beet storage pile until early February. Differences (P < 0.0001 to 0.0150) among fungicide treatments were evident. Propulse- and Stadium-treated roots had 84 to 100% less fungal growth versus the control roots, whereas fungal growth on Mertect-treated roots was not different from the control roots in 7 of 12 comparisons for roots harvested each of the first 3 weeks in both years of this study. The Propulse- and Stadium-treated roots also reduced (P < 0.0001 to 0.0146; based on weeks 1, 3, and 4 in 2012 and weeks 1, 3, 4, and 5 in 2013) sucrose loss by 14 to 46% versus the control roots, whereas roots treated with Mertect did not change sucrose loss compared with the control roots in 7 of 10 evaluations. The predominant fungi isolated from symptomatic roots were an Athelia-like sp., Botrytis cinerea, Penicillium spp., and Phoma betae. If Propulse and Stadium are labeled for use on sugar beet in storage, these fungicides should be considered for root rot control in commercial sugar beet storage and on roots held for vernalization for seed production of this biennial plant species.

Rhizomania, caused by Beet necrotic yellow vein virus (BNYVV) is an economically important disease of sugar beet. Management includes the use of single resistance (Rz) genes. However, strains with the ability to overcome the widely used Rz1 resistance gene have already been reported. The mechanisms associated with these genes are unknown, therefore increased understanding of the host defense response may lead to more durable sources of rhizomania resistance. Non-targeted metabolome profiling, coupled with targeted phytohormone quantification, was applied to characterize the metabolic and signaling pathways associated with BNYVV infections in sugar beet. Clear differences in metabolites were identified following pathogen infection independent of plant genotype particularly those associated with host lipid composition or other membrane associated functions.

Sugar beet can be infected by many different viruses that can reduce yield; beet necrotic yellow vein virus (BNYVV) is one of the most economically important viruses of this crop plant. This report describes a new reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay for identification of BNYVV. In addition, a novel immunocapture (IC) RT-LAMP assay for rapid and easy detection (without RNA extraction) of BNYVV was developed here and compared with DAS-ELISA and RT-LAMP assays. Our results show that the IC-RT-LAMP assay is a highly reliable alternative assay for identification of BNYVV.

Beet necrotic yellow vein virus (BNYVV) is the most devastating pathogen of sugar beet worldwide. This virus has been reported in the majority of sugar beet growing regions of Iran as well. For the present study, we collected samples from different sugar beet varieties with suspected symptoms of BNYVV from the main important sugar beet growing regions in eight provinces of Iran. Infection of collected samples to BNYVV was tested by ELISA and RT-PCR. Upon testing of 167 collected samples of BNYVV suspected through ELISA and RT-PCR, 115 (68.9%) were infected. Different incidences of BNYVV through surveyed provinces may represent the presence of diverse infective viral sources or resistance genes in tested sugar beet varieties which need further attempts to develop control strategies. Results also showed that BNYVV has been recently distributed throughout some surveyed regions. Otherwise, trace infection or resistance to BNYVV infection in some varieties of distinct regions may represent proper sources of re...

&ldquo, Rhizomania&rdquo, of sugar beet is a soilborne disease complex comprised of beet necrotic yellow vein virus (BNYVV) and its plasmodiophorid vector, Polymyxa betae. Although BNYVV is considered the causal agent of rhizomania, additional viruses frequently accompany BNYVV in diseased roots. In an effort to better understand the virus cohort present in sugar beet roots exhibiting rhizomania disease symptoms, five independent RNA samples prepared from diseased beet seedlings reared in a greenhouse or from field-grown adult sugar beet plants and enriched for virus particles were subjected to RNAseq. In all but a healthy control sample, the technique was successful at identifying BNYVV and provided sequence reads of sufficient quantity and overlap to assemble &gt, 98% of the published genome of the virus. Utilizing the derived consensus sequence of BNYVV, infectious RNA was produced from cDNA clones of RNAs 1 and 2. The approach also enabled the detection of beet soilborne mosaic virus (BSBMV), beet soilborne virus (BSBV), beet black scorch virus (BBSV), and beet virus Q (BVQ), with near-complete genome assembly afforded to BSBMV and BBSV. In one field sample, a novel virus sequence of 3682 nt was assembled with significant sequence similarity and open reading frame (ORF) organization to members within the subgenus Alphanecrovirus (genus Necrovirus, family Tombusviridae). Construction of a DNA clone based on this sequence led to the production of the novel RNA genome in vitro that was capable of inducing local lesion formation on leaves of Chenopodium quinoa. Additionally, two previously unreported satellite viruses were revealed in the study, one possessing weak similarity to satellite maize white line mosaic virus and a second possessing moderate similarity to satellite tobacco necrosis virus C. Taken together, the approach provides an efficient pipeline to characterize variation in the BNYVV genome and to document the presence of other viruses potentially associated with disease severity or the ability to overcome resistance genes used for sugar beet rhizomania disease management.

Rhizomania disease, caused by the Beet necrotic yellow vein virus (BNYVV), is considered as one of the major constraints for sugar beet production, worldwide. As a result of the introgression of major resistance genes (Holly, Rz2) in commercially available sugar beet varieties, the virus has endured strong selection pressure since the 90s'. Understanding the virus response and diversity to sugar beet resistance is a key factor for a sustainable management of only few resistance genes. Here we report rhizomania surveys conducted in a rhizomania hot spot, the Pithiviers area (France) during a 4-year period and complementary to the study of Schirmer et al. (2005). The study aimed at evaluating the intra- and inter-field BNYVV diversity in response to different sources of resistance and over the growing season. To follow rhizomania development over the sugar beet growing season, extensive field samplings combined with field assays were performed in this study. The evolution of the BNYVV diversity was assessed at intra- and inter-field levels, with sugar beet cultivars containing different resistance genes (Rz1, Rz1 + Heterodera schachtii resistance and Rz1Rz2). Intra-field diversity was analyzed at the beginning and the end of the growing season of each field. From more than one thousand field samples, the simultaneous presence of the different A, B and P types of BNYVV was confirmed, with 21 variants identified at positions 67–70 of the p25 tetrad. The first variant, AYHR, was found most commonly followed by SYHG. Numerous mixed infections (9.93% of the samples), mostly of B-type with P-type, have also been evidenced. Different tetrads associated with the A- or B-type were also found with a fifth RNA-genome component known to allow more aggressiveness to BNYVV on sugar beet roots. Cultivars with Rz1+Rz2 resistant genes showed few root symptoms even if the BNYVV titre was quite high according to the BNYVV type present. The virus infectious potential in the soil at the end of the growing season with such cultivars was also lower despite a wider diversity at the BNYVV RNA3 sequence level. Rz1+Rz2 cultivars also exhibited a lower presence of Beet soil-borne virus (BSBV), a P. betae-transmitted Pomovirus. Cultivars with Rz1 and nematode (N) resistance genes cultivated in field infected with nematodes showed lower BNYVV titre than those with Rz1 or Rz1+Rz2 cultivars. Overall, the population structure of BNYVV in France is shown to be different from that previously evidenced in different world areas. Implications for long-term management of the resistance to rhizomania is discussed.

Rhizomania is one of serious threat to sugar beet production in Morocco and in several parts of the world. This disease led to a statistically significant decrease in the quality and yield of sugar beet plantations. Therefore, this study aimed at comparing the efficacy of six commonly used RNA extraction methods for the detection, recovery of RNA of beet necrotic yellow vein virus (BNYVV) and removal of amplification inhibitors by reverse transcription-polymerase chain reaction (RT-PCR). The efficiency of these extraction methods was then compared to that of a commercial isolation kit with high content of phenolic compounds. The results showed that the extraction with the lithium chloride technique, the commercial kit, and direct and membrane spotting crude extract methods were found effective in yielding a higher purity and a higher concentration of RNA when compared to the other tested methods. Extraction with the lithium chloride technique and the Qiagen kit (RNeasy Plant Mini Kit) allowed the most intense band, whereas the CTAB method has generated the least intense band. Furthermore, the silica capture extraction method did not yield any RNA after extraction and electrophoresis. Consequently, it was concluded that, of these six methods, the lithium chloride technique and the Qiagen kit are the most appropriate for the extraction of viral RNA from sugar beet samples prior to RT-PCR for detecting BNYVV.

The reform of the European Union's sugar regime caused potential decreasing beet prices. Therefore, the Speeding Up Sugar Yield (SUSY) project was initiated. At the start, a 3 × 15 target was formulated: in 2015 the national average sugar yield in the Netherlands equals 15 t/ha (60% of the sugar beet potential) and the total variable costs 15 euro/t sugar beet, aspiring a saving on total variable costs and a strong increase in sugar yield. Based on their average sugar yield in 2000-2004, 26 pairs of "type top" (high yielding) and "type average" (average yielding) growers were selected from all sugar beet growing regions in the Netherlands. On the fields of those farmers, all measures of sugar beet cultivation were investigated, including cost calculation and recording phytopathological, agronomical and soil characteristics in 2006 and 2007. Although there was no significant difference in total variable costs, the "type top" growers yielded significantly 20% more sugar in each year compared to the "type average" growers. Therefore, the most profitable strategy for the growers is maximizing sugar yield and optimizing costs. The difference in sugar yield between growers could be explained by pests and diseases (50%), weed control (30%), soil structure (25%) and sowing date (14%), all interacting with each other. The SUSY-project revealed the effect of the grower's management on sugar yield. As a follow up for the SUSY-project, a growers' guide "Suikerbietsignalen" was published, Best Practice study groups of growers were formed and trainings and workshops were given and field days organized. Further, the benchmarking and feedback on the crop management recordings and the extension on variety choice, sowing performance, foliar fungi control and harvest losses were intensified. On the research part, a resistance breaking strain of the Beet Necrotic Yellow Vein Virus (BNYVV) and a new foliar fungus, Stemphylium beticola, were identified and options for control were tested, and implemented in growers practices. The joint efforts of sugar industry, sugar beet research and growers resulted in a raise in sugar yield from 10.6 t/ha in 2002-2006 to 13.8 t/ha in 2012-2016.