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Bacterial Leaf Spot of Tomato (Solanum lycopersicum) in Louisiana is Caused by Xanthomonas perforans, Tomato Race 4

    Authors and Affiliations
    • M. L. Lewis Ivey , Department of Plant Pathology & Crop Physiology, LSU AgCenter, Baton Rouge, LA 70803
    • A. Strayer , Department of Plant Pathology, University of Florida, Gainesville 32611
    • J. K. Sidhu , Department of Plant Pathology & Crop Physiology, LSU AgCenter, Baton Rouge, LA 70803
    • G. V. Minsavage , Department of Plant Pathology, University of Florida, Gainesville, FL 32611.

      In fall 2013 and spring 2014, bacterial spot characterized by necrotic leaf spots with chlorosis was observed on commercially produced indeterminate tomato plants in three parishes (Livingston, East Baton Rouge, and Tangipohoa) in southern Louisiana. Disease incidence was 100% and foliar disease severity ranged from 20 to 80%, on a scale of 0 to 100%. No symptoms were observed on the fruit at the time of observation. Diseased leaves were collected from each location, and lesions were excised, surface sterilized by dipping the leaf tissue in 70% ethanol, air dried, and macerated in 10 mM potassium phosphate buffer (KPB), pH 7.4. Ten-fold serial dilutions in KPB were plated on yeast dextrose calcium carbonate medium and four yellow, mucoid colonies were selected and purified. All isolates were classified as Gram negative according to the potassium hydroxide (KOH, 3%) test, oxidase negative, catalase positive, and induced a hypersensitive response in tobacco (Nicotiana tabacum) plants 48 h after inoculation with a 108 CFU/ml bacterial suspension. All isolates (MLI2-13, MLI26-14, MLI28-14, and MLI38-14) were identified as Xanthomonas spp. based on the production of an ∼420-bp product using a PCR assay with the genus-specific primers RST65/69 (Obradovic et al. 2004). All strains were identified as Xanthomonas perforans race 4 using real-time multiplex Taqman PCR with X. perforans probe, which targets hrpB2, and primers F1 and R1 (Strayer et al. 2014) and HR test results on tomato differential genotypes [FL216, Hawaii 7998, and a near isogenic line of FL8000 containing RxopJ4 (formerly Xv4)] (Sharlach et al. 2013; Stall et al. 2009). Pathogenicity tests were performed on 7-week-old tomato seedlings (cv. Roma). Three seedlings per test strain were sprayed until runoff with a suspension (∼108 CFU/ml) of X. perforans strain MLI2-13, ML26-14, ML28-14, or MLI38-14. Two additional seedlings were sprayed with water and served as negative controls. All plants were grown in the greenhouse under high relative humidity (∼80%) at day/night temperatures of 33/28°C for 14 days. Seedlings inoculated with the Xanthomonas strains exhibited water-soaked lesions within 10 days after inoculation. Seedlings sprayed with water did not develop symptoms. Isolations were performed as described above and the recovered isolates were similar in morphology to the original strains and produced an ∼420-bp amplicon using a PCR assay with the genus-specific primers RST65/69. Bacterial spot of tomato is widespread in Louisiana; however, this is the first report that identifies the species and race of Xanthomonas causing this disease. This new knowledge will allow commercial producers and homeowners to select tomato varieties with corresponding genetic resistance and will allow researchers to monitor shifts in Xanthomonas spp. populations causing bacterial spot of tomato in Louisiana.