Suppression of Resistance-Breaking Beet necrotic yellow vein virus Isolates by Beet oak-leaf virus in Sugar Beet.

Rhizomania, a serious disease of sugar beet (Beta vulgaris), is caused by Beet necrotic yellow vein virus (BNYVV). Resistance allele Rz1 has been widely incorporated into commercial cultivars. Recently, resistance-breaking isolates of BNYVV (RB-BNYVV) were identified and characterized. When the occurrence of RB-BNYVV was surveyed throughout the sugar-beet-growing areas in the United States, most soil samples contained Beet oak-leaf virus (BOLV) as well. BNYVV and BOLV often occurred in the same field and sometimes in the same sugar beet plant. The possibility of interactions between these two Polymyxa betae-transmitted sugar beet viruses was tested. Plants grown in soils infested with aviruliferous P. betae or carrying RB-BNYVV and BOLV, alone and in combination, were compared with plants grown in noninfested soil for differences in plant fresh weight and virus content as measured by enzyme-linked immunosorbent assay (ELISA). Rz1 and Rz2 resistance genes that condition resistance to BNYVV did not confer resistance to BOLV. BNYVV ELISA values were significantly higher in single infections than in mixed infections with BOLV in both the rhizomania-resistant and -susceptible cultivars. In contrast, ELISA values of BOLV were not significantly different between single and mixed infections in both the rhizomania-resistant and -susceptible cultivars. Results indicate that BOLV may suppress BNYVV in mixed infections. Soils infested with P. betae significantly reduced fresh weight of sugar beet seedlings regardless of whether they were with or without one or both viruses or resistance genes.

Distribution and Molecular Characterization of Resistance-Breaking Isolates of Beet necrotic yellow vein virus in the United States.

Beet necrotic yellow vein virus (BNYVV) is the causal agent of rhizomania in sugar beet (Beta vulgaris). The virus is transmitted by the plasmodiophorid Polymyxa betae. The disease is controlled primarily by the use of partially resistant cultivars. During 2003 and 2004 in the Imperial Valley of California, partially resistant sugar beet cultivars with Rz1 allele seemed to be compromised. Field trials at Salinas, CA have confirmed that Rz1 has been defeated by resistance-breaking isolates. Distinct BNYVV isolates have been identified from these plants. Rhizomania-infested sugar beet fields throughout the United States were surveyed in 2004-05. Soil surveys indicated that the resistance-breaking isolates not only existed in the Imperial Valley and San Joaquin Valley of California but also in Colorado, Idaho, Minnesota, Nebraska, and Oregon. Of the soil samples tested by baited plant technique, 92.5% produced infection with BNYVV in 'Beta 6600' (rz1rz1rz1), 77.5% in 'Beta 4430R' (Rz1rz1), 45.0% in 'Beta G017R' (Rz2rz2), and 15.0% in 'KWS Angelina' (Rz1rz1+Rz2rz2). Analyses of the deduced amino acid sequence of coat protein and P-25 protein of resistance-breaking BNYVV isolates revealed the high percentage of identity with non-resistance-breaking BNYVV isolates (99.9 and >98.0%, respectively). The variable amino acids in P-25 proteins were located at the residues of 67 and 68. In the United States, the two amino acids found in the non-resistance-breaking isolates were conserved (AC). The resistance-breaking isolates were variable including, AF, AL, SY, VC, VL, and AC. The change of these two amino acids cannot be depended upon to differentiate resistance-breaking and non-resistance-breaking isolates of BNYVV.

Stalk Rot of Sugar Beet Caused by Fusarium solani on the Pacific Coast.

In 2006, symptoms of stalk blight (2) were observed on sugar beet (Beta vulgaris L.) plants from roots produced in Oregon that were being grown for seed production in a greenhouse in Salinas, CA using Salinas Valley soil. Symptoms included vascular and cortical browning, necrosis, and death of seed stalks. Isolations were made from the edge of stalk lesions and the crown. In addition to Fusarium oxysporum, the known cause of stalk blight (2), two isolates of F. solani were identified by morphology. For pathogenicity tests, sugar beet plants (FC606 [4]), grown in pasteurized potting mix and induced to flower by exposure at 4 to 7°C for 90 days (1) were used. Bolting plants were maintained in a greenhouse at 24 to 27°C. A 100-µl drop of a spore suspension (104 spores per ml) of each Fusarium isolate was placed on the surface of the seed stalk. The plant was stabbed through the drop with a sterile 18-gauge needle so that the drop was taken into the plant by hygroscopic pressure. Positive and negative control treatments were a stalk blight isolate of F. oxysporum from an Oregon seed production field and sterile water, respectively. Three plants were inoculated per isolate. Each inoculation site was wrapped loosely in Parafilm for 1 week to maintain a high humidity level around the site of inoculation, and seed stalks were covered in cloth bags (1). After 1 week, the Parafilm was removed and plants were examined weekly for symptoms. At 4 weeks, lesion size was measured. After 5 weeks, sections were taken from the seed stalk around the site of inoculation, surface disinfested with 0.5% NaOCl, and plated on potato dextrose agar to confirm the presence of the pathogen. The experiment was done twice. One of the two isolates of F. solani caused dark brown lesions on all inoculated seed stalks. On one plant, at 4 weeks after inoculation when the bag was being removed for observation, the seed stalk broke at the site of inoculation because of a spreading, brown lesion at the site. No lesions were observed on the water control plants. Brown lesions were observed on seed stalks inoculated with the known stalk blight isolate. Lesions were significantly (P = 0.001) larger with F. oxysporum than with F. solani when measured at 4 weeks (mean of 6.3 cm versus 2.2 cm, respectively). Lesions caused by F. solani showed a dark discoloration through the cortical tissue, as opposed to those caused by F. oxysporum, for which most of the initial discoloration was in the vascular bundles and epidermis. Fusarium isolates recovered from inoculated plants were morphologically similar to the isolates used for inoculation. Fusarium spp. were not isolated from the water control plants. While some F. solani isolates cause seedling or mature root disease in sugar beet (3), to our knowledge, this is the first report of a Fusarium species other than F. oxysporum causing a rot of sugar beet stalks. References: (1) E. Biancardi et al. Genetics and Breeding of Sugar Beet Science Publishers, Inc., Enfield, NH, 2005. (2) A. N. Mukhopadhay. Handbook of Diseases of Sugar Beet, Vol. 1. CRC Press, Boca Raton, FL 1987. (3) E. G. Ruppel. Plant Dis. 75:486, 1991. (4) G. A. Smith and E. G. Ruppel. Crop Sci. 19:300, 1980.

Occurrence of Resistance-Breaking Beet necrotic yellow vein virus of Sugar Beet.

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.

Inheritance of Powdery Mildew Resistance in Sugar Beet Derived from Beta vulgaris subsp. maritima.

Powdery mildew of sugar beet (Beta vulgaris), caused by Erysiphe polygoni, was introduced into North American in 1974. Since then, chemical control has been needed. Moderate resistance of a slow-mildewing type is known and has been used commercially. High resistance was identified recently in B. vulgaris subsp. maritima accessions WB97 and WB242 and has been backcrossed into sugar beet breeding lines. These enhanced lines were used as sources of powdery mildew resistance to determine the inheritance of resistance. Analyses of segregating testcross families showed that resistance from both sources is inherited as a single, dominant, major gene. The gene symbol Pm is proposed for the resistant allele. The allelism of the resistance from the two wild beet sources was not determined. Pm conditions a high level of resistance, but disease developed on matured leaves late in the season. This late development of mildew on lines and the slow-mildewing trait in susceptible, recurrent lines tended to obfuscate discrete disease ratings.

Specificity of TAS-ELISA for Beet Necrotic Yellow Vein Virus and Its Application for Determining Rhizomania Resistance in Field-Grown Sugar Beets.

Levels of beet necrotic yellow vein virus (BNYVV), as measured by triple-antibody sandwich-enzyme-linked immunosorbent assay (TAS-ELISA), were compared with biological evaluations in representative commercial and experimental sugar beet cultivars developed for production in the United States and ranging in their reactions to rhizomania from uniformly susceptible to highly resistant. TAS-ELISA was specific for BNYVV and did not react with related soilborne sugar beet viruses. Differences in absorbance (A405nm) values measured in eight cultivars closely correlated with the dosage and frequency of the Rz allele, which conditions resistance to BNYVV. A diploid (Rzrz) hybrid had a significantly lower absorbance value (less virus) than a similar triploid (Rzrzrz) hybrid. Cultivars that segregated (Rzrz:rzrz) had higher absorbance values than uniformly resistant (Rzrz) hybrids, as was expected. For all cultivars, absorbance values decreased as the season progressed. Absorbance value was significantly positively correlated with rhizomania disease index score and negatively correlated with individual root weight, plot root weight, and sugar yield. This information should be useful in resistance-breeding and -evaluation programs and in the sugar industry when considering cultivar choice, inoculum production, and future crop rotations.

First Report of Bacterial Leaf Spot of Swiss Chard Caused by Pseudomonas syringae pv. aptata in California.

From 1999 through 2003, a previously unreported disease was found on commercial Swiss chard (Beta vulgaris subsp. cicla) in the Salinas Valley, (Monterey County) California. Each year the disease occurred sporadically throughout the long growing season from April through September. Initial symptoms were water-soaked leaf spots that measured 2 to 3 mm in diameter. As disease developed, spots became circular to ellipsoid, 3 to 8 mm in diameter, and tan with distinct brown-to-black borders. Spots were visible from the adaxial and abaxial sides. Cream-colored bacterial colonies were consistently isolated from spots. Strains were fluorescent on King's medium B, levan positive, oxidase negative, and arginine dihydrolase negative. Strains did not rot potato slices but induced a hypersensitive reaction on tobacco (Nicotiana tabacum cv. Turk). The isolates, therefore, belong in LOPAT group 1 (1). Fatty acid methyl esters (FAME) analysis (MIS-TSBA version 4.10, MIDI Inc., Newark, DE) gave variable results that included Pseudomonas syringae, P. cichorii, and P. viridiflava with similarity indices ranging from 0.91 to 0.95. BOX-polymerase chain reaction (PCR) analysis gave identical banding patterns for the chard isolates and for known P. syringae pv. aptata strains, including the pathotype strain CFBP1617 (2). The bacteria were identified as P. syringae. Pathogenicity of 11 strains was tested by growing inoculum in nutrient broth shake cultures for 48 h, diluting to 10 × 6 CFU/ml, and spraying onto 5-week-old plants of Swiss chard cvs. Red, White, Silverado, and CXS2547. Untreated control plants were sprayed with sterile nutrient broth. After 7 to 10 days in a greenhouse (24 to 26°C), leaf spots similar to those observed in the field developed on all inoculated plants. Strains were reisolated from the spots and identified as P. syringae. Control plants remained symptomless. To investigate the host range of this pathogen, the same procedures were used to inoculate three strains onto other Chenopodiaceae plants: five cultivars of sugar beet (B. vulgaris), and one cultivar each of spinach (Spinacia oleracea) and Swiss chard. In addition, five chard strains and strain CFBP1617 were inoculated onto two cultivars of sunflower (Helianthus annuus), and one cultivar each of cantaloupe (Cucumis melo), sugar beet, spinach, and Swiss chard. All Swiss chard, cantaloupe, sunflower, and sugar beet plants developed leaf spots after 7 days. The pathogen was reisolated from spots and confirmed to be the same bacterium using BOX-PCR analysis. Spinach and untreated controls failed to show symptoms. All inoculation experiments were done at least twice and the results were the same. The phenotypic data, fatty acid and genetic analyses, and pathogenicity tests indicated that these strains are P. syringae pv. aptata. To our knowledge this is the first report of bacterial leaf spot of commercially grown Swiss chard in California caused by P. syringae pv. aptata. The disease was particularly damaging when it developed in Swiss chard fields planted for "baby leaf" fresh market products. Such crops are placed on 2-m wide beds, planted with high seed densities, and are sprinkler irrigated. This disease has been reported from Asia, Australia, Europe, and other U.S. states. References: (1) R. A. Lelliott et al. J. Appl. Bacteriol. 29:470, 1966. (2) J. L. W. Rademaker et al. Mol. Microbiol. Ecol. Man. 3.4.3:1-27, 1998.

Interactions Between Beet necrotic yellow vein virus and Beet soilborne mosaic virus in Sugar Beet.

Soils naturally infested with cultures of aviruliferous Polymyxa betae and viruliferous P. betae carrying two sugar beet benyviruses, Beet necrotic yellow vein virus (BNYVV) and Beet soilborne mosaic virus (BSBMV), alone and in combination, were compared with noninfested soil for their effects on seedling emergence, plant fresh weight, and virus content as measured by enzyme-linked immunosorbent assay (ELISA). Studies examined sugar beet with and without resistance to the disease rhizomania, caused by BNYVV. The Rz gene, conferring resistance to BNYVV, did not confer resistance to BSBMV. BSBMV ELISA values were significantly higher in single infections than in mixed infections with BNYVV, in both the rhizomania-resistant and -susceptible cultivars. In contrast, ELISA values of BNYVV were high (8 to 14 times the healthy mean) in single and mixed infections in the rhizomania-susceptible cultivar, but were low (approximately three times the healthy mean) in the rhizomania-resistant cultivar. Results indicate BNYVV may suppress BSBMV in mixed infections, even in rhizomania-resistant cultivars in which ELISA values for BNYVV are extremely low. Soils infested with P. betae, and with one or both viruses, showed significantly reduced fresh weight of seedlings, and aviruliferous P. betae significantly decreased sugar beet growth in assays.