First Report of Bacterial Fruit Blotch of Watermelon Caused by Acidovorax avenae subsp. citrulli in Hungary.

Typical bacterial fruit blotch (BFB) symptoms were observed on cvs. Crisby, Suzy, Top Gun, and Lady watermelon (Citrullus lanatus (Thunb.) Matsum. & Nakai.) fruits in southeastern Hungary (Medgyesegyháza) in July 2007 when the mean maximum daytime temperature was greater than 32°C. Approximately 20 to 30 ha of watermelons were observed to be affected by the disease. Source of the infection was not determined, however, grafted watermelon transplants that were grown in this area had been imported from Turkey where the pathogen is present (2). Disease symptoms started with irregularly shaped, water-soaked lesions on the surface of the fruits. The lesions enlarged and the epidermis became brown and cracked. BFB symptoms were not readily visible on mature foliage. Colonies of the BFB pathogen were creamy white on nutrient agar (Difco, Detroit, MI). Strains were gram negative, oxidase positive, and produced acid from glucose aerobically. A cell suspension (50 μl of ~1 × 10⁷ CFU/ml) from a 24-h nutrient plate culture was infiltrated with a hypodermic syringe into the intercellular spaces of fully developed intact tobacco (Nicotiana tabacum L. cv. White Burley) leaves to determine the hypersensitive reaction (HR) (1). A typical HR developed 20 h after leaf infiltration. Bacterial pathogenicity was tested on surface-sterilized, mature fruits of different plant species by injecting cell suspensions into the fruit tissues as previously described (each fruit was injected in five places; the negative control (sterile water) as well). Fruits were incubated for 7 days at 25°C and then observed for symptom development. Necrosis was observed at each point of inoculation with the pathogen for watermelon and green pepper (Capsicum annuum L.). Necrosis was also observed for cucumber (Cucumis sativus L.), zucchini (Cucurbita pepo L. convar. giromontiina Greb.), squash (C. pepo L.), and patisson (C. pepo L. convar. patissoniana Greb.). Necrosis was not observed when the pathogen was inoculated onto fruit of melon (Cucumis melo L.), tomato (Lycopersicon esculentum Mill.), and eggplant (Solanum melongena L.). Additionally, symptoms were not observed at the points inoculated with sterile water (negative control) for any of the fruits tested. To identify the pathogen, PCR was used with Acidovorax avenae subsp. citrulli-specific primers WFB1/2 (4). The 16s rDNA region amplified with a general bacterial primer pair (63f forward and 1389r reverse) (3) was cloned into a pBSK+ vector (Stratagene, La Jolla, CA) and sequenced by M13 forward and reverse primers (GenBank Accession No. AM850114). On the basis of the symptoms, biochemical tests (API 20NE; Biomérieux, Marcy l'Etoile, France), fatty acid methyl ester analysis (74.5 to 83.6% similarity), and 16SrDNA sequence homology (100% sequence similarity with AAC00-1), the pathogen was identified as A. avenae subsp. citrulli. To our knowledge, this is the first report of BFB of watermelon in Hungary. References: (1) Z. Klement. Nature 199:299, 1963. (2) M. Mirik et al. Plant Dis. 6:829, 2006. (3) A. M. Osborn et al. Environ. Microbiol. 2:39, 2000. (4) R. R. Walcott and R. D. Gitaitis. Plant Dis. 84:470, 2000.