Ken Pernezny

An outbreak of bacterial spot of lettuce in Florida caused by Xanthomonas campestris pv. vitians

A widespread and damaging outbreak of a leaf spot disease of lettuce occurred in the 1992-93 winter vegetable season in southern Florida. Individual leaf lesions were dark brown to black, water-soaked, and greasy in appearance. A yellow-pigmented bacterium was consistently isolated. All 1992-93 lettuce strains and reference strains produced symptoms in greenhouse test plants that were identical to those seen in the field. Disease reactions generally were more severe in cos and butterhead lettuce than in crisphead. Strains were gram-negative rods, and negative for glucose fermentation, nitrate reduction, urease production, and utilization of asparagine as a sole source of carbon an dnitrogen. Aesculin was hydrolyzed, gelatin was liquefied, and proteolysis occurred in litmus milk. Cellular fatty acid profiles matched well to library database strains of Xanthomonas campestris pv. vitians

Bacterial diseases of tomato in the Czech and Slovak Republics and lack of streptomycin resistance among copper-tolerant bacterial strains

Abstract Surveys of foliar and fruit bacterial diseases of tomato were conducted in 1992 and 1993 in the Czech and Slovak Republics. Pseudomonas syringae pv. tomato , causal agent of bacterial speck, was isolated most frequently. Pseudomonas syringae pv. tomato accounted for 86% of the strains recovered from clinic samples at the Research Institute for Crop Production in Prague in 1992. All of the strains recovered from diseased samples collected in Moravia, Czech Republic in 1993 and 90% of those from Slovakia in 1993 were also P. s. pv. tomato . The bacterial spot pathogen Xanthomonas campestris pv. vesicatoria was recovered infrequently in both years. No strains of P. s. pv. tomato or X. c. pv. vesicatoria were resistant to 200 μg ml −1 streptomycin. Copper tolerance was prevalent; frequency of copper tolerance of P. s. pv. tomato strains was greater than 75% in both 1992 and 1993.

Losses in fresh-market tomato production in Florida due to target spot and bacterial spot and the benefits of protectant fungicides

Three large-scale field tests were conducted under commercial growing conditions in south-western Florida to quantify yield losses associated with foliar diseases of fresh-market tomatoes, document the benefits of protectant fungicides, and establish a protocol to be used in making yield-loss assessments for fresh-market tomatoes. The benefits of protectant fungicides were amply demonstrated in two of three experiments. Marketable and extra-large fruit yields were reduced 30 and 43% in these tests, respectively, when no fungicides were used. Net returns on investment were

Bacterial Leaf Spot of Lettuce: Relationship of Temperature to Infection and Potential Host Range of Xanthomonas campestris pv. vitians

7,800 to

Investigation of Seed Treatments for Management of Bacterial Leaf Spot of Lettuce

14,800/ha greater in plots treated with chlorothalonil than in control plots. Much of the loss in marketable yield was due to direct damage to fruit by the target spot fungus. As much as two-thirds of the fruit had to be discarded in no-spray plots because of blemishes from target spot. Tank-mix sprays of copper-mancozeb provided good early season control of bacterial spot, but there was no correlation between bacterial spot damage levels and yield or monetary losses. Little return on investment in protectant fungicides was recorded where target spot was not a factor in the third experiment, even though low to moderate levels of bacterial spot were present.

A Serious Outbreak of Race 6 of Xanthomonas campestris pv. vesicatoria on Pepper in Southern Florida

Epidemiological aspects, including optimum temperature for infection and host range of Xanthomonas campestris pv. vitians, causal organism of bacterial leaf spot (BLS) of lettuce, were investigated. The optimum temperature for infection was determined to be 22.7°C based on growth chamber studies. Internal populations were monitored over time in lettuce, tomato, pepper, parsley, cilantro, and beet. Each plant species was infiltrated with the bacterium at 105CFU/ml. Highest populations developed in lettuce (108CFU/cm2) followed by pepper with 106CFU/cm2, whereas the other plant species harbored much lower populations (105 to 103CFU/cm2). Infectivity titration endpoints were similar in pepper and lettuce (103 to 104CFU/ml). For other plant species tested, infectivity titration endpoints were 106 to 107 CFU/ml. Electrolyte leakage data and corresponding internal population data support the conclusion that fresh-market tomato is not a host of X. campestris pv. vitians but, instead, interacts in an incompatible response. Electrolyte leakage from cells of tomato plants inoculated with X. campestris pv. vitians or a pepper strain of X. axonopodis pv. vesicatoria peaked at 48 h, suggesting that tomato is not a host for the BLS pathogen. Both electrolyte leakage and population dynamics results point to pepper as a potential host of X. campestris pv. vitians.

Multilocus Sequence Typing of Strains of Bacterial Spot of Lettuce Collected in the United States

Chemical seed treatments were evaluated for efficacy of disinfestation of lettuce seed that had been inoculated with Xanthomonas campestris pv. vitians. Three concentrations of each chemical were evaluated by treating seed lots for 5 or 15 min. In addition, the effects of each seed treatment on seed germination and early plant growth were examined by observing seed germination rates. Bacteria were not detected when seed were treated with 3 or 5% hydrogen peroxide for 5 or 15 min. Treatment of seed with 0.52% sodium hypochlorite was relatively ineffective at 5 and 15 min. When sodium hypochlorite was used at a 1% concentration for 15 min, the level of bacterial infestation was reduced to 2%. Suspensions of copper hydroxide plus mancozeb also reduced seedborne inoculum to ≤2%. Treatment of seed with copper hydroxide alone, benzoyl peroxide, or calcium peroxide did not reduce seed infestation levels significantly. Seed germination rates were 90% or greater for the majority of seed treatments tested in laboratory assays. Hydrogen peroxide treatments at a concentration of 5% reduced seed germination up to 28% compared with controls. However, no significant differences in germination were observed among control treatments (noninoculated, nontreated seed and inoculated, nontreated seed) and any of the chemical seed treatments when seed were sown in a pasteurized soil mix in the greenhouse.

An Outbreak of a Leaf Spot Disease of Cabbage in Southern Florida Caused by Xanthomonas campestris pv. armoraciae

In the 1997-1998 winter vegetable season, a widespread and serious outbreak of bacterial spot occurred on pepper in commercial fields throughout southern Florida. This was the first serious epidemic of bacterial spot on pepper in southern Florida since the 1993-1994 season. Cultivars affected included those with resistance to races 1-3 of X. campestris pv. vesicatoria, commonly found in previous Florida surveys (1). Field sampling designs, isolation methods, and pathogen identification were carried out as previously described (1). Twenty-seven fields were sampled in Palm Beach, St. Lucie, Martin, Broward, and Collier counties. The race of each strain was determined by infiltration of 3.0 × 108 CFU per ml of sterile tap water suspension into Early Calwonder and three near-isogenic lines of Early Calwonder with genes for vertical resistance to specific races of the pathogen. Test plants were maintained in the greenhouse and observed for hypersensitive and disease reactions over several days. Of a total of 244 X. campestris pv. vesicatoria strains collected, 73.4% were identified as race 6 and 10.2% as race 4. Irrespective of the presence of resistance genes in cultivars sampled, race 6 (eastern Florida) and race 4 (southwestern Florida) predominated in this study. Copper tolerance was widespread among X. campestris pv. vesicatoria strains of all races, with 91% of the strains showing tolerance to 250 g per ml of CuSO4 · 5H2O incorporated into 0.5% (wt/vol) glucose-nutrient agar. Selection pressure resulting from widespread planting of race 1, 2, 3-resistant cultivars may have contributed to this outbreak. High disease pressure also may be related to the unusually wet winter. Rainfall accumulation from December 1997 through March 1998 was 25.6 cm above normal. Reference: (1) K. Pohronezny et al. Plant Dis. 76:118, 1992.

The etiology of recent pepper anthracnose outbreaks in Florida

Studies on genetic diversity and recombination in bacterial pathogens are providing a better understanding of the mechanisms shaping bacterial diversity, which can affect disease control. Xanthomonas campestris pv. vitians, causal agent of bacterial leaf spot of lettuce, is a threat to the worldwide lettuce industry. We examined the genetic variation within a sample of 83 strains from California, Florida, and Ohio using multilocus sequence typing of six housekeeping genes, totaling 2.7 kb. Additionally, polymorphism in two virulence-related genes, hrpB2 and a putative glycosyl hydrolase, were examined. Based on housekeeping genes, we found three genetic groups of strains that were all able to induce the disease. These included strains collected from weeds and irrigation water that had haplotypes identical to strains from diseased lettuce. High linkage disequilibrium across the sequenced loci indicates that the pathogen is predominantly clonal but recombination has contributed to the observed sequence variation. Although there was significant genetic variation in X. campestris pv. vitians within and among sampled states, identical haplotypes were observed across all three states. This finding suggests that seedborne inoculum may contribute to the diversity of X. campestris pv. vitians in the United States. Knowledge of the genetic structure of the pathogen may be used for developing resistant lettuce varieties.

Comparison of two culture media for determination of the copper resistance of Xanthomonas strains and their usefulness for prediction of control with copper bactericides

From October to December 2001, a leaf spot disease was observed in numerous commercial fields of red and green cabbage (Brassica oleracea var. capitata L.) in the Everglades Agricultural Area, south and east of Lake Okeechobee and in the environs of Immokalee in southwestern Florida. Discrete water-soaked to greasy appearing spots were observed in the leaf blades with no evidence of marginal V-shaped lesions characteristic of black rot caused by Xanthomonas campestris pv. campestris. Profuse bacterial streaming was observed when cut leaf sections were examined microscopically. A bacterium that formed yellow colonies on nutrient agar was consistently isolated from these lesions. Ten bacteria were isolated, purified, and characterized. All strains were aerobic, gram-negative rods. Strains were positive for esculin hydrolysis, proteolysis in litmus milk, and gelatin liquefaction. Strains were negative for urease production, nitrate reduction, oxidase, and utilization of asparagine as a sole source of carbon and nitrogen. Fatty acid methyl ester analysis indicated a match with Florida library strains of X. campestris pv. raphani (similarity indices 0.605-0.738). Suspensions (2 × 107 CFU/ml in phosphate-buffered saline) of two Oklahoma strains identified as X. campestris pv. armoraciae provided by J. P. Damicone (3) and four representative Florida strains were applied to plants using a hand-held sprayer. Pathogenicity of the strains was tested on three replicate greenhouse-grown plants of the following: green cabbage cv. Market Early; red cabbage cv. Salad Delight; radish cv. Red Silk; tomato cv. Sunny; sweet bell pepper cv. Jupiter; and fresh horseradish roots purchased from a retail grocery chain. A strain of X. campestris pv. campestris originally isolated from Homestead, FL was also included in pathogenicity tests. All Florida and Oklahoma strains produced leaf spots, but no V-shaped lesions, on leaves of green cabbage, red cabbage, radish, tomato, and horseradish. Typical black rot symptoms were observed only in radish and green and red cabbage inoculated with the X. campestris pv. campestris strain. On the basis of these results, we identify the Florida strains as X. campestris pv. armoraciae (1,2,3), recognizing the precedent of X. campestris pv. armoraciae over X. campestris pv. raphani based on extensive genetic and serological data (1). Our strains appear to be more similar to those causing outbreaks on crucifers in Oklahoma (3) than those in Ohio (2), because Florida strains were pathogenic on tomato. References: (1) A. M. Alvarez et al. Phytopathology 84:1449, 1994. (2) F. Sahin and S. A. Miller. Plant Dis. 81:1334, 1997. (3) Y. Zhao et al. Plant Dis. 84:1008, 2000.