Detection of Xanthomonas campestris pv. carotae in carrot seed.

was plated in triplicate onto MD5 agar Kuan, T.-L., Minsavage, G. V., and Gabrielson, R. L. 1985. Detection of Xanthomonas campestris medium. pv. carotae in carrot seed. Plant Disease 69:758-760. Seed rinsing was compared with seed A method has been developed to isolate Xanthomonas campestris pv. carotae (X. c. carotae) in soaking as a means of extracting the carrot seed lots. The method consists of a low-temperature (5 C) stationary aqueous soak of carrot bacteria. Twenty grams of carrot seed seed in darkness for 18 hr followed by a vigorous shaking of the seed in water containing Tween 20, were shaken vigorously in 100 ml of cold concentration of bacterial cells by centrifugation (7,796 g for 15 min), and plating of a dilution sterile saline for 30 sec; the suspension series of these cells onto a modified Kado and Heskett's D5 medium. The method yielded was then filtered through sterile cheeserepeatable and consistent results in replicated testing of seed lots. The bacterium was detected in 34 cloth and collected in another sterile of 35 seed samples from infected fields and zero of six seed samples from healthy fields.. One flask. This was then centrifuged. The naturally infected seed in 10,000 seeds can be detected by this method. This is the first report of pellet was resuspended, diluted, and direct isolation of X. c. carotae from carrot seed. aliquots plated as described before. To provide comparison seed-soak data, 100 Bacterial blight of carrots, caused by infected seed fields were used to test this ml of fresh sterile saline was poured onto Xanthomonas campestris (Pam.) Dows. method. Tests of several seed lots were the rinsed seeds, extracted, and plated as pv. carotae Kendr. (X. c. carotae), was replicated at different times to confirm before. first reported in California in 1934 (9). the repeatability of the test method. Evaluation and identification of X. c. The disease has since been reported in Isolation media. Kado and Heskett's carotae. The pathogen and saprophytic Idaho (2,10,17), Arizona, New Mexico (8) D5 medium was modified (MD5) by bacteria were evaluated after inoculated (4), New York (13), Wisconsin (14), substituting filter-sterilization (0.22 tgm) plates were incubated in the dark at 30 C Australia (16), Canada (16), Italy (3), and for heat-sterilization of D (+) cellobiose for 7-10 days. On the MD5 medium, Japan (12). It is of increasing concern in (Sigma Chemical Company, St. Louis, colonies of X. c. carotae were typically carrot production fields in Florida (J. 0. MO) and by addition of cycloheximide straw yellow, glistening, round, smooth, Strandberg, personal communication). (final concentration 200 mg/ L) (Acti- convex with entire margins, and 3-5 mm Seed transmission of the disease has been dione; Upjohn Company, Kalamazoo, in diameter. Colonies typical of X. c. demonstrated by Kendrick (9) with MI). These were added after the carotae were purified by two successive artificially infested seed and by Ark and autoclaved basal medium cooled to 60 C. streak transfers from single colonies to Gardner (1) with naturally infected seed. Media-filled plates were dried for 3 days plates of YNA before further testing. Several investigators (1,2,9,10,17) have at room temperature to eliminate surface All suspected isolates of X. c. carotae observed bacterial blight in carrot seed moisture before use. Recovery of bacteria were tested for pathogenicity on seedlings umbels. An important means of control on MD5 medium was compared by of Daucus carota L. var. sativus Hoffm. will be the use of clean carrot seed. dilution plating with recovery on yeast- 'Chancellor' at the second- to fourth-trueEffective eradicant seed treatments and extract nutrient agar (YNA) (7) using leaf stage by atomizing the leaves to control procedures in carrot seed fields three isolates of X. c. carotae obtained runoff with bacterial suspensions of 1 X are vital to the production and availability from J. 0. Strandberg (Agricultural 106 colony-forming units (cfu) per of clean carrot seed. Research and Extension Center, P.O. milliliter. At least five plants were An accurate method is needed to detect Box 909, Sanford, FL) and seven inoculated with each bacterial isolate the pathogen in and on seed, to evaluate pathogenic isolates obtained from plant tested. The inoculated plants were disease control programs in seed fields, to samples.