Susceptibility of Onion Relatives (Allium spp.) to Iris yellow spot virus.

Iris yellow spot virus (IYSV; family Bunyaviridae, genus Tospovirus) is becoming an increasingly important constraint to the production of bulb and seed onions (Allium cepa L.) in many onion-growing regions of the continental United States and the world (4). During an evaluation of onion germplasm for susceptibility to IYSV, six other Allium species (A. altaicum, A. galanthum, A. roylei, A. schoenoprasum, A. tuberosum, and A. vavilovii) were also evaluated under natural field conditions. In July 2010, symptoms suggestive of IYSV infection (straw-colored necrotic lesions) were observed on leaves of these Allium spp. in experimental plots in Las Cruces, NM. IYSV was detected in symptomatic leaves of A. altaicum, A. vavilovii, A. tuberosum, A. schoenoprasum and A. roylei with a commercially available ELISA kit (Agdia Inc., Elkhart, IN). IYSV infection was confirmed by reverse transcription (RT)-PCR with forward and complementary primers 5'-CTCTTAAACACATTTAACAAGCAC-3' and 5'-TAAAACAAACATTCAAACAA-3' flanking the nucleocapsid (N) gene encoded by the small RNA of IYSV as previously described (1,3). Amplicons, approximately 1.1 kb long, were obtained from all symptomatic Allium spp. samples but not from healthy samples or water controls. Sequencing of selected amplicons confirmed IYSV infection. The highest nucleotide identity of 98% was shared with IYSV isolates from Japan (GenBank Accession No. AB180921). A. altaicum, A. vavilovii, and A. pskemense were previously reported from Washington to be susceptible to IYSV (2). Current findings expand the list of Allium spp. that are susceptible to IYSV and underscores the need for continued screening of other members of the genus to find sources of resistance to IYSV. References: (1) H. R. Pappu et al. Arch. Virol. 151:1015, 2006. (2) H. R. Pappu et al. Plant Dis. 90:378, 2006. (3) H. R. Pappu et al. Plant Dis. 92:588, 2008. (4) H. R. Pappu et al. Virus Res. 141:219, 2009.

First Report of Enterobacter cloacae Causing Onion Bulb Rot in the Columbia Basin of Washington State.

In August of 2006, onion plants of cv. Redwing exhibiting premature dieback and bulb rot were obtained from a commercial onion crop under center pivot irrigation in the Columbia Basin of Washington State. High temperatures during the summer were similar to those in 2004, which preceded significant outbreaks of Enterobacter rot of onion bulbs in storage. Fungal pathogens of onion were not observed. Bacteria from infected bulb tissue were isolated and purified on nutrient broth yeast extract (NBY) agar, and 537 isolates were evaluated for the ability to ferment glucose anaerobically. Of the facultative anaerobes (~50% of all isolates), 48 isolates were arginine dihydrolase positive, indole negative, and unable to degrade pectin, i.e., characteristics typical of the genus Enterobacter (2), which includes Enterobacter cloacae, a bacterial pathogen reported to cause onion bulb rot in California and Colorado (1,3). Sixteen of the putative Enterobacter isolates, along with four strains of E. cloacae known to be pathogenic on onion (1) (ATCC 23355 and ATCC 13047, 310 (H. F. Schwartz, Colorado State University), and E6 (J. Loper, USDA ARS), were tested for pathogenicity on onion bulbs (8 to 10 cm in diameter; cv. Tamara). The isolates were grown overnight in NBY broth at 28°C, harvested by centrifugation and resuspended to an OD600 = 0.3 (~108 CFU/ml) in sterile distilled water. After the outermost fleshy scale of each bulb was removed, each bulb was surface disinfected in 0.6% NaOCl for 2 min, dipped in sterile distilled water, and then dipped in 95% ethanol. Each bulb was air dried before a 0.5-ml aliquot of bacterial suspension was injected into the shoulder of the bulb with a 20-gauge needle. Three bulbs were inoculated for each isolate, placed in individual plastic bags, sealed, and incubated at 30°C in the dark. Three bulbs injected with water and three noninjected bulbs served as controls. After 14 days, each bulb was sliced through the center and rated for rot. Thirteen isolates induced rot symptoms on the inner fleshy scales of all inoculated bulbs. Of these, seven also caused tan-to-brown discoloration of the inner fleshy scales; similar symptoms were caused by the four pathogenic reference strains of E. cloacae (1). No symptoms were observed in any of the controls. Symptoms were not observed when the bacteria, prepared as described above, were infiltrated into onion leaves. Bacteria were reisolated from the symptomatic inoculated bulb tissue and confirmed to be Enterobacter spp. by the above physiological tests. In addition, an isolate designated ECWSU2 and the corresponding strain recovered from one of the inoculated symptomatic bulbs, along with the four reference strains, were evaluated for anaerobic growth on a variety of carbon sources by using API 50 CHE test strips (bio Mérieux Vitek, Inc., Hazlewood, MO). The physiological test data along with sequence analysis of a portion of the 16S rRNA gene of each isolate confirmed all of these isolates to be E. cloacae (4; Ribosomal Database Project [ http://rdp.cme.msu.edu/ ]). To our knowledge, this is the first report of E. cloacae causing a bulb rot of onion in Washington State. References: (1) A. L. Bishop and R. M. Davis. Plant Dis. 74:692, 1990. (2) J. G. Holt et al. Bergey's Manual of Determinative Bacteriology. Williams and Wilkins, Baltimore, MD, 1994. (3) H. F. Schwartz and K. Otto. Plant Dis. 84:808, 2000. (4) L. Verdonck et al. Int. J. Syst. Bacteriol. 37:4, 1987.

First Report of Brown Root Rot of Alfalfa Caused by Phoma sclerotioides in Colorado and New Mexico.

Brown root rot (BRR), caused by the fungal pathogen Phoma sclerotioides G. Preuss ex Sacc. (synonym Plenodomus meliloti Dearn. & G.B. Sanford), is associated with winterkill, slow emergence from winter dormancy, and yield loss of alfalfa (Medicago sativa L.) (1,2). BRR is a problem in regions with severe winters and is common in Alaska and Alberta, Saskatchewan and Manitoba, Canada. It was first observed in the continental United States in Wyoming during 1996 (2) and has subsequently been found in Idaho, Minnesota, Montana, New Hampshire, New York, Vermont, and Wisconsin. In the intermountain valleys of northern New Mexico and western Colorado, winters can be severe; alfalfa winterkill events occur periodically, but it is unknown if BRR is present. In May 2006, alfalfa plants were collected from production fields in Huerfano, Otero, and Rio Grande counties in Colorado and Rio Arriba and Taos counties in New Mexico and assessed for BRR. Two to three fields were sampled per county and 20 or 40 plants were collected per field. All fields existed for at least two winters. Fields sampled in Rio Grande County exhibited severe winterkill, with most plants completely girdled by crown lesions. Plants from other fields exhibited a range of root and crown rots. Isolation of P. sclerotioides was attempted from all plants with a previously described protocol (4). The pathogen was isolated from crown lesions of one alfalfa plant each from Rio Grande and Taos counties. Both lesions extended into the cortex. On potato dextrose agar and water agar with barley (4), single-conidium cultures of each isolate produced large pycnidia (0.35 to 0.80 mm in diameter) with multiple beaks, white cirri darkening to yellow with age, and unicellular, hyaline, ovoid conidia 5 to 7 µm long by 2 µm wide. Diagnostic PCR of the cultures using P. sclerotioides-specific primers (3) resulted in a single amplicon of expected size (500 bp). The internal transcribed spacer (ITS) 1, 5.8S, and ITS2 of the rDNA were amplified and sequenced using primers ITS1 and ITS4. The ITS sequences (GenBank Accession Nos. EU265669 and EU265670) were >98% identical to P. sclerotioides ATCC isolate 56515 over 503 bp of aligned sequence. Potted 'Vernal' alfalfa was inoculated 4 months after seeding, kept at 4°C for 5.5 weeks, 0 to -2°C for 12 weeks, and 4°C for 3 weeks. Of the 14 plants inoculated with the Colorado isolate, 11 developed cortical lesions and 8 winterkilled. Of the 23 plants inoculated with the New Mexico isolate, 22 developed cortical lesions and 16 winterkilled. Lesions were light to very dark brown, sometimes with a darker border and often containing abundant pycnidia. Winterkill was associated with lesions girdling the crown. P. sclerotioides was isolated from the lesions. To our knowledge, this is the southernmost report of BRR in North America and the first report of BRR in New Mexico and Colorado. The incidence and severity of BRR in the region surveyed appear to be considerably lower than in the more northern regions. References: (1) B. Berkenkamp et al. Can. J. Plant Sci. 71:211, 1991. (2) C. R. Hollingsworth et al. Can. J. Plant Pathol. 25:215, 2003. (3) R. C. Larsen et al. Plant Dis. 86:928, 2002. (4) M. J. Wunsch et al. Plant Dis. 91:1293, 2007.

First Report of Iris yellow spot virus on Onion in New York.

Iris yellow spot virus (IYSV [family Bunyaviridae, genus Tospovirus]), a potentially devastating disease of onion vectored by onion thrips (Thrips tabaci Lindeman), has been reported from most states in the western United States where significant onion production occurs, with the most recent report from Texas (1). In June 2006, volunteer onion (Allium cepa) plants in Orleans County, New York (Elba muckland) were found to have symptoms indicative of IYSV infection. The scapes (seed stalks) of the volunteer onions found at the edge of a cull pile from a 2005 onion crop exhibited diamond-shaped lesions, each with a distinct green center and a double yellow border. Approximately 25 of 100 plants of red and yellow onion cultivars exhibited characteristic IYSV lesions. The cull pile was composed primarily of locally grown onions, although a few of the bulbs were grown from imported bare-root transplants imported from Arizona. Symptomatic plants tested positive for IYSV using IYSV-specific antiserum from Agdia Inc. (Elkhart, IN) in a double-antibody sandwich-ELISA. The presence of IYSV was verified by reverse transcription (RT)-PCR using primers derived from the small RNA of IYSV (S-RNA). The primers flanked the IYSV nucleocapsid (N) gene (5'-TAA AAC AAA CAT TCA AAC AA-3' and 5'-CTC TTA AAC ACA TTT AAC AAG CAC-3' (3). RT-PCR assays produced a PCR amplicon of expected size (approximately 1.2 kb) and the product was cloned and sequenced. Nucleotide sequence analysis confirmed the identity of the amplicon as that of the IYSV S-RNA. Sequence comparisons showed 95 to 98% identity with known IYSV N gene sequences available in GenBank. The virus is poorly transmitted to onion by mechanical inoculation and we did not have access to a noninfested colony of the onion thrips vector to transfer the virus from these samples to noninfected onions. No asymptomatic plants were tested. Among the onion-growing states in the eastern United States, IYSV has previously only been reported from Georgia (2). To our knowledge, this is the first report of IYSV in New York and the greater northeastern United States. The finding of this disease in New York confirms further spread of the virus within North America and the need for research to develop more effective management options to reduce the impact of IYSV on onion crops. References: (1) M. Miller et al. Plant Dis. 90:1359, 2006. (2) S. W. Mullis et al. Plant Dis. 90:377, 2006. (3) H. R. Pappu et al. Arch. Virol. 151:1015, 2006.

First Report of Iris yellow spot virus in Commercial Leek (Allium porrum) in the United States.

Iris yellow spot virus (IYSV; family Bunyaviridae, genus Tospovirus) has a wide host range, with onion (Allium cepa L.) being one of the most economically important hosts. IYSV has been widely reported from this species throughout most onion-production regions of the United States and many areas of the world in recent years. A relative of onion, leek (Allium porrum L.), has been reported to be a host of IYSV in countries such as the Netherlands, Reunion Island, and Australia (1,4). A related tospovirus, Tomato spotted wilt virus (TSWV), was recently reported causing necrotic lesions and extended bleaching of leaf tips of leek in Georgia (2). In September of 2006, disease symptoms suspected to be caused by IYSV were observed on central and outer leaves of plants in a 2.6-ha section of commercial leeks being grown from seed (cvs. Tadorna and King Richard). The leek plants were adjacent to a 3.1-ha section of seeded onion (cv. Exacta) that had been harvested 2 weeks earlier. Twenty-five to thirty percent of unharvested onion plants next to the leek section also exhibited IYSV-type disease symptoms generally on the central leaves. Both Allium spp. were seeded 5 months earlier and grown under certified organic, pivot-irrigated conditions in Larimer County in northern Colorado. Disease symptoms on leek and onion leaves appeared as dry, white-to-straw-colored, spindle- or diamond-shaped lesions that ranged in size from 5 to 10 × 25 to 50 mm or larger depending on lesion age. Lesion centers, especially on leek, often had green centers with concentric rings of alternating green and straw-colored tissue. Green tissue near necrotic lesions of a single symptomatic leaf from 10 plants each of leek and onion was sampled and analyzed using a double-antibody sandwich (DAS)-ELISA (Agdia, Inc., Elkhart, IN). Five of ten leek and nine of ten onion samples were positive for IYSV. Using reverse transcription (RT)-PCR and primers specific to the small RNA of IYSV (5'-TAA AAC AAA CAT TCA AAC AA-3' and 5'-CTC TTA AAC ACA TTT AAC AAG CAC-3'), the complete nucleocapsid (N) gene was amplified from symptomatic leek plants and then sequenced (3). Comparisons with IYSV N gene sequences available in the GenBank confirmed the identity of the virus as IYSV. Leek samples were negative for TSWV when tested by RT-PCR with TSWV-specific primers. In addition, three specimens of the presumed thrips vector recovered from five IYSV-infected leek plants were identified as Thrips tabaci (L. A. Mahaffey and W. S. Cranshaw, personal communication). Earlier in the season, T. tabaci was observed in the nearby planting of onion that also exhibited IYSV in September. To our knowledge, this is the first report of natural infection of commercial leek with IYSV in the United States. The incidence of plants (25 to 30%) with foliar lesions on multiple leaves and stunting of 5% of infected plants in both leek cultivars suggests that IYSV could seriously reduce leek stem development and marketability. References: (1) I. Cortes et al. Phytopathology 88:1276, 1998. (2) C. Nischwitz et al. Plant Dis. 90:525, 2006. (3) H. R. Pappu et al. Arch. Virol. 151:1015, 2006. (4) T. N. Smith et al. Plant Dis. 90:729, 2006.

Sequence diversity of the nucleoprotein gene of iris yellow spot virus (genus Tospovirus, family Bunyaviridae) isolates from the western region of the United States.

Iris yellow spot virus (IYSV), a tentative virus species in the genus Tospovirus and family Bunyaviridae, is considered a rapidly emerging threat to onion production in the western United States (US). The present study was undertaken to determine the sequence diversity of IYSV isolates from infected onion plants grown in California, Colorado, Idaho, Oregon, Utah and Washington. Using primers derived from the small RNA of IYSV, the complete sequence of the nucleoprotein (NP) gene of each isolate was determined and the sequences compared. In addition, a shallot isolate of IYSV from Washington was included in the study. The US isolates of IYSV shared a high degree of sequence identity (95 to 99%) with one another and to previously reported isolates. Phylogenetic analyses showed that with the exception of one isolate from central Oregon and one isolate from California, all the onion and shallot isolates from the western US clustered together. This cluster also included onion and lisianthus isolates from Japan. A second distinct cluster consisted of isolates from Australia (onion), Brazil (onion), Israel (lisianthus), Japan (alstroemeria), The Netherlands (iris) and Slovenia (leek). The IYSV isolates evaluated in this study appear to represent two distinct groups, one of which largely represents isolates from the western US. Understanding of the population structure of IYSV would potentially provide insights into the molecular epidemiology of this virus.

New Outbreaks of Bacterial Wilt of Dry Bean in Nebraska Observed from Field Infections.

Bacterial wilt caused by Curtobacterium flaccumfaciens pv. flaccumfaciens was one of the more problematic diseases of dry bean (Phaseolus vulgaris L.) throughout the irrigated High Plains (Colorado, Nebraska, and Wyoming) in the 1960s and early 1970s, but has not been observed since that time. However, in August of 2003, plants exhibiting wilting and irregular, interveinal necrotic foliar lesions surrounded by a bright yellow border were found in three dry bean fields (market class Great Northern) in Scotts Bluff County, Nebraska. During 2004, plants exhibiting identical symptoms were additionally found occurring in more than 40 dry bean fields in western Nebraska. Affected fields were planted with dry bean from multiple market classes and seed sources, including yellow bean, Great Northern bean, and pinto bean, and incidence varied from trace levels to 80 to 90%. Isolations were made from leaf and stem tissues and seeds collected after harvest from infected plants, and all yielded slow-growing, creamy yellow or orange, fluidal colonies on nutrient broth-yeast extract medium. The bacterium was identified as C. flaccumfaciens pv. flaccumfaciens based on cell morphology (coryneformshaped motile rods), positive Gram stain and KOH reactions, fatty acid profiles, and BIOLOG (Hayward, CA) identifications. Great Northern (cv. Orion) plants were inoculated by bacterial suspensions (5 × 107 CFU/ml) injected into leaf axils adjacent to the first fully expanded trifoliolate and were incubated in the greenhouse under ambient conditions fluctuating between 24 and 35°C. Wilting symptoms developed 7 days after inoculation with foliar necrosis and yellowing symptoms appearing after 24 days. Identical bacterial colonies were reisolated from inoculated tissues, completing Koch's postulates. Although recent reports of wilt have been made in North Dakota (2) and western Canada (1) in 1995 and 2002, respectively, they were based only on the presence of discolored seeds observed in dockage from processing plants after harvest. To our knowledge, this report represents the first widespread observations of bacterial wilt from field infections in Nebraska in more than 30 years. References: (1) J. R. Venette et al. Plant Dis. 79:966, 1995. (2) T. F. Hsieh et al. Plant Dis: 86:1275, 2002.

Sexual Stage Development of Uromyces appendiculatus and Its Potential Use for Disease Resistance Screening of Phaseolus vulgaris.

In June 1989, pycnia and aecia of the bean rust pathogen were observed in eastern Colorado on volunteer plants of pinto bean cvs. UI 114 and UI 126 that grew from seed that remained in the field after harvest the previous season. Harvested aeciospores were viable and produced typical reddish-brown uredinia on unifoliolate leaves of UI 114 seedlings in the greenhouse. Evidence of bean rust overwintering has been confirmed in 10 years from 1989 to 2002 in eastern Colorado and the surrounding region. Overwintering conditions were reproduced at Fort Collins, CO in fall 1992 and spring 1993. Debris treatments had significantly higher disease incidence on stems, total number of lesions on stems, disease incidence on leaves, and total number of lesions on leaves of plants of pinto cvs. UI 114 and Olathe than plants without debris. Bean leaves of both cultivars had significantly higher disease incidence than stems. There also was an increased incidence of aecial infection for UI 114 seedlings that germinated through leaf debris with rust compared with stem debris under greenhouse conditions with abundant moisture. Bean leaf debris smaller than 0.36 mm in diameter resulted in significantly more aecial lesions on UI 114 than larger leaf debris and stem debris. Additional greenhouse experiments demonstrated that diverse collections of naturally occurring, overwintered, rust-infested bean debris from eastern Colorado produced different levels of pycnial and aecial infection on pinto cvs. UI 114, Olathe, and Chase. Chase, currently resistant to the uredinial stage of the prevalent bean rust races in Colorado and surrounding states, had a low incidence of aecial lesions on seedling stems when exposed to pycnial and aecial stages in the greenhouse. These observations indicate that selection of bean rust resistance genes should rely on multiple sources of resistant germ plasm to counter the potential increase in new races that could derive from sexual recombination in bean-production regions.

First Report of Albugo tragopogonis on Cultivated Sunflower in North America.

White rust, caused by Albugo tragopogonis (Pers.) S.F. Gray, was observed on a few plants of both oilseed and confection sunflowers (Helianthus annuus L.) in northwestern Kansas (Cheyenne County) in 1992. The disease was observed again from 1993 to1995 in nine counties in western Kansas, with incidence per field ranging up to 35%. White rust was found only on late-planted fields in 1996 and 1997 and was not found at all from 1998 to 2001. White rust was also observed on cultivated and wild sunflower (H. annuus) for the first time in eastern Colorado (Kit Carson and Yuma counties) from 1994 to 1997, but was absent from 1998 to 2001. Leaf pustules on both cultivated and wild sunflowers were similar in appearance. Pustules were convex, chlorotic on the upper side of the leaf, and concave and dull white on the under side of the leaf. Pustules on cultivated sunflower were generally limited to three to six leaves in the middle of the plant and affected 10 to 40% of the leaf area. Sporangial dimensions fell within the reported dimensions for A. tragopogonis (2). In 1997, water-soaked lesions 1 to 2 cm long containing oospores of A. tragopogonia were observed on the lower to middle portions of stems of cultivated sunflower in western Kansas and the adjacent area of Colorado. Stem lesions were observed much less frequently than foliar lesions and only in 1997. Sporangia were not observed in stem lesions, nor were any other fungi isolated from these lesions. To our knowledge, this is the first report of white rust occurring on cultivated sunflower in any production area of North America; the disease has not been observed in the major U.S. sunflower production area of North Dakota, South Dakota, and Minnesota. Foliar white rust lesions generally have little economic impact on sunflower, but the presence of stem lesions is significant because stem lesions may lead to lodging (3). Lodging due to A. tragopogonia was not observed in either Kansas or Colorado. White rust has previously only been reported on wild H. annuus in Wisconsin and on perennial Helianthus spp. in Missouri and Illinois (1). References: (1) D. F. Farr et al. Fungi on Plants and Plant Products in the United States. The American Phytopathological Society, St. Paul, MN 1989. (2) K. G. Mukeri. Description of Pathogenic Fungi and Bacteria. CMI, Kew, Surrey, UK 1976. (2) P. S. van Wyk et al. Helia 22:83, 1995.

Iris yellow spot virus on Onion in Colorado.

A new disease was found in September 2001 on greenhouse-produced onion transplants of cv. Colorado 6 grown in a field in Larimer County in northern Colorado. Symptoms included straw-colored, dry, tan, spindle- or diamond-shaped lesions on the leaves and scapes of onion plants. Infected plants were scattered (less than 5% incidence) throughout the outer perimeter of the sprinkler-irrigated field. Iris yellow spot virus (IYSV) in two collections each with 4 to 6 symptomatic onion plants was confirmed with western blot assays by James Moyer of North Carolina State University. Western blot showed a faint band from protein extracts of infected Nicotiana benethimiana. Western blot assay is the most definitive method of identification. IYSV can be mechanically transmitted to N. benethimiana, but it cannot be recovered and transmitted back to onion, and it is difficult to detect in infected onion plants. IYSV is a tospovirus that is transmitted by various species of thrips, including onion thrips and western flower thrips (1). The host range for this disease includes onion, leek, and iris. IYSV has been reported previously on onion in Israel, Brazil, and Idaho (2). There are no reports that this disease affects bulb quality or marketability; however, heavy losses of onion bulb production are reported (1). University and industry personnel in other onion-growing areas of the country are encouraged to monitor onion and other host fields for evidence and distribution of IYSV. References: (1) A. Kritzman et al. Plant Dis. 85:838, 2001. (2) L. Pozzer et al. Plant Dis. 83:345, 1999.

Vegetative Compatibility Groups of Fusarium oxysporum f. sp. cepae from Onion in Colorado.

Nineteen isolates of Fusarium oxysporum f. sp. cepae recovered from diseased onions growing in the western, southern, and northern regions of Colorado were placed into vegetative compatibility groups (VCGs) based on pairing of complementary mutants. Pathogenic isolates from these regions were cultured on variations of potassium chlorate (1.5 or 3.0%) mutation media, potato dextrose agar (PDA), and minimal medium (MM) supplemented with L-asparagine and L-threonine. Chlorate PDA and 3% chlorate MM with L-threonine did not generate the nitrate nonutilizing (nit) mutants required, while MM with L-asparagine (1.5 and 3% chlorate) and MM with L-threonine (1.5% chlorate) generated complementary nit mutants required for compatibility pairings. Five VCGs of F. oxysporum cepae were identified. One VCG was present in all three regions of Colorado examined. Four VCGs were restricted to either western or eastern Colorado. Additional sampling and evaluation of a more diverse collection of F. oxysporum cepae isolates from other regions of onion production is needed to determine the diversity of this pathogen. Such information could assist in breeding for resistance to F. oxysporum cepae.

First Report of a Bulb Decay of Onion by Enterobacter cloacae in Colorado.

Sweet Spanish onion cultivars (Allium cepa L.) in northern Colorado displayed symptoms of postharvest bulb rot during September to November of 1999. This disease appears identical to that reported from infected onions in California in 1988 and is presumably associated with high temperature stress (1). Mature, firm bulbs harvested from scattered fields in Weld County exhibited a brownish discoloration and breakdown of inner scales. Gram negative, rod-shaped, cream-colored bacteria were consistently recovered from infected bulb tissue on nutrient agar. Physiological tests showed that the bacteria utilized glucose in an oxidative and fermentative manner and were catalase positive and oxidase negative. A representative strain was identified by Microbe Inotech Laboratories (St. Louis, MO) as Enterobacter cloacae (Jordan) Hormaeche & Edwards (2) using Biolog analysis, with a similarity index of 0.81. To confirm pathogenicity, a 0.5- to 1.0-ml suspension of bacteria (108 CFU/ml sdw) was injected into firm onion bulbs (7.5 to 10.0 cm diameter). After incubation for 14 days at 22°C in closed plastic bags in the dark, bulbs were cut in half and evaluated. Tan to brown discoloration and initial dry rot, similar to that observed postharvest, was observed in inoculated bulbs. The pathogen was reisolated from six of eight bulbs inoculated with the representative strain. No discoloration or disease developed on eight control bulbs injected with water. To our knowledge, this is the first report of E. cloacae from onion grown in Colorado. References: (1) A. L. Bishop and R. M. Davis. Plant Dis. 74:692, 1990. (2) Hormaeche and Edwards. Int. J. System. Bacteriol. 30:293, 1960.

First Report of a Leaf Blight and Bulb Decay of Onion by Pantoea ananitas in Colorado.

Sweet Spanish onion (Allium cepa L.) cultivars grown in southern Colorado displayed symptoms of foliar blight and bulb rotting after bulb initiation in early July of 1997, 1998, and 1999. This disease appears identical to that reported from infected onions in Georgia in 1997 (1). Leaf blighting began as whitish to tan lesions, which rapidly coalesced, causing a general wilt, discoloration, and death of affected foliage. A yellow-cream to light orange discoloration progressed into bulbs, resulting in the rotting of neck tissue and between scales. Infection of more than 70% of onion plants exposed to heavy rain and storm damage after bulb initiation occurred in scattered fields in Otero County. Gram negative, rod-shaped, yellow-colored bacteria were consistently recovered from infected foliar and bulb tissues on nutrient agar during this 3-year period. Physiological tests showed that the bacteria utilized glucose in an oxidative and fermentative manner and were catalase positive and oxidase negative. Two strains recovered during 1997 were identified by Microbe Inotech Laboratories (St. Louis, MO) as Pantoea ananas by gas-chromatography fatty acid methyl ester analysis, with similarity indices of 0.70 and 0.79. A literature search determined that the accepted classification is now Pantoea ananatis Serrano (2). To confirm pathogenicity, a 0.5- to 1.0-ml suspension of bacteria (108 CFU/ml sdw) of one of the strains was injected into firm onion bulbs (7.5 to 10.0 cm diameter). After incubation for 14 days at 22°C in enclosed plastic bags in the dark, bulbs were cut in half and scored for visual evidence of yellow to tan discoloration and initial dry rotting prior to reisolation of the pathogen from five of eight inoculated bulbs. No discoloration or disease developed on eight control bulbs injected with water. To our knowledge, this is the first report of P. ananatis from onion grown in Colorado and the western United States. References: (1) R. D. Gitaitis and J. D. Gay. Plant Dis. 81:1096, 1997. (2) H. G. Truper and L. de Clari. Int. J. System. Bacteriol. 47:908, 1997.

First Report of a Leaf Blight of Onion Caused by Xanthomonas campestris in Colorado.

Sweet Spanish onion (Allium cepa L.) cultivars in southern Colorado (Otero and Prowers counties) have been found with symptoms of a foliar blight since 1996, and the same symptoms have been observed in northern Colorado (Weld County) since 1997. This onion disease appears to be identical to that reported from Barbados in 1971, Hawaii in 1978, and Texas in 1998 (1). Leaf blighting in scattered fields began as linear, tan to brown, water-soaked lesions that rapidly coalesced and were often surrounded by chlorotic areas, causing a general discoloration and tip die-back of affected foliage. The disease occurs generally after periods of heavy rainfall or storms. Bulb size may be reduced, and 10 to 15% yield losses have been recorded from control plots in copper-based bactericide screening nurseries naturally infected primarily by this pathogen at Rocky Ford during 1996 to 1998. Disease progression into bulbs has not been observed. Gram negative, rod-shaped, yellow bacteria were consistently recovered from infected foliar and bulb tissues on nutrient agar. The bacterial isolates utilized glucose in an oxidative manner, were catalase positive, oxidase negative, and negative for the tetrazolium salt tolerance test (0.1 and 0.02%). Two strains, one recovered during 1996 and the other in 1997, were identified by Microbe Inotech Laboratories (St. Louis, MO) as Xanthomonas campestris based on fatty acid methyl ester analysis (similarity indices of 0.81 and 0.82). A literature search indicated that classification to pathovar is lacking (2). To prove pathogenicity, a suspension (108 CFU/ml sdw) of one of the strains was sprayed to runoff onto a flat of approximately 100 8-week-old plants. Inoculated plants were placed in a dew chamber for 24 h, and then transferred to a bench and maintained at 25 to 28°C with a 12-h photoperiod and misting period for 14 days. A sample of 10 randomly selected plants with symptoms of water-soaking and discoloration was collected from which X. campestris was reisolated. No symptoms developed on seedlings sprayed with water only. This is the first report of X. campestris from onions grown in Colorado and the western continental United States. References: (1) T. Isakeit et al. Plant Dis. 84:201, 2000. (2) M. Van Den Mooter and J. Swings. Int. J. Syst. Bacteriol. 40:348-369, 1990.

Epiphytic Pseudomonas syringae on Dry Beans Treated with Copper-Based Bactericides.

The response of epiphytic populations of Pseudomonas syringae and other bacteria on dry bean plants to four copper-based bactericides was evaluated. The bactericides showed little difference in efficacy, but epiphytic populations on pinto bean leaflets, flowers, and pods were occasionally reduced when compared to populations on non-treated control plants, especially after repeated bactericide applications. Although there was a trend toward a relationship between epiphytic leaflet and flower populations, there were cases where P. syringae was undetected on one organ but abundant in samples from the other organ. P. syringae pv. syringae strains recovered from epiphytic populations demonstrated much greater copper resistance than did strains of P. syringae pv. phaseolicola, as measured by growth on media amended with cupric hydroxide. This difference between the pathovars may have implications for integrated pest management strategies.

Pathogenicity of Fusarium oxysporum f. sp. phaseoli Isolates from Spain to Phaseolus vulgaris.

Fusarium oxysporum Schechtend.:Fr. f. sp. phaseoli J. B. Kendrick & W. C. Snyder (FOP) is the causal agent of the common bean (Phaseolus vulgaris L.) disease known as Fusarium wilt or Fusarium yellows. FOP has been reported from the Castilla y Leon region in Spain, where it is a serious problem on most commercial bean cultivars (1). Five FOP isolates from Spain (AB-6, AB-111, AB-112, AS-1, and AS-4) obtained from J. M. Díaz-Mínguez and the isolate FOP-CO1 (ATCC 90245) from Colorado were tested for pathogenicity on two American lines: Pinto U.I. 114, considered as a universal susceptible check, and Flor de Mayo, a Mexican landrace. Seedlings were root-clip inoculated and evaluated according to the CIAT 1 to 9 severity scale, in which 1 to 3 = resistant, 3.1 to 6 = intermediate, and 6.1 to 9 = susceptible. Three inoculum concentrations were tested: 104, 105, and 106 conidia/ml. AS-1 and AS-4 produced resistant reactions in both U.I. 114 (1.5 to 2.1 and 1.2 to 1.5) and Flor de Mayo (1.9 to 2.3 and 1.4), respectively, at all inoculum concentrations. Susceptible reactions to AB-6 (7.3 to 8.9), AB-111 (7.6 to 8.9), AB-112 (7.5 to 7.9), and FOP-CO1 (8.1 to 8.6) were observed in Pinto U.I. 114, regardless of inoculum concentration, although severity ratings increased as the inoculum concentration was increased. Flor de Mayo exhibited a susceptible reaction (6.7 to 7.8) to FOP-CO1 at all inoculum concentrations tested, and intermediate reactions to AB-6 (4.9), AB-111 (5.4), and AB-112 (5.0) at the lowest inoculum concentration. Susceptible reactions (7 to 8.2 for AB-6; 6.8 to 7.0 for AB-111, and 7.3 to 7.7 for AB-112) occurred with higher inoculum concentrations, and severity ratings increased as the inoculum concentration increased. Recently, FOP isolates from Greece and Italy were recognized as each belonging to different pathogenic races (2); consequently, more research on the Spanish isolates (AB-6, AB-111, and AB-112) is needed to determine if they are similar to the races reported from the Mediterranean Basin or should be classified as a new FOP race (s). References: (1) J. M. Díaz-Mínguez et al. Plant Dis. 80:600, 1996. (2) S. L. Woo et al. Phytopathology 86:966, 1996.