First Report of Bacterial Wilt Caused by Ralstonia solanacearum on Roselle in Taiwan

Roselle (Hibiscus sabdariffa L.) is a herbaceous plant belonging to the Malvaceae family. Its calyxes are rich in vitamin C and anthocyanins and are used to make roselle drink and hibiscus tea. Roselles are grown in counties of Taitung, Pingtung, and Chiayi in Taiwan. In addition to a few local cultivars, the major cultivar currently grown in Taiwan is Roselle cv. Victor. In April of 2012, a wilt disease appeared on seedlings of a cultivar, Chiada 1, at the Chungpu Township of Chiayi County. Mature plants were free from this disease. Leaves appeared weak and drooping when they were still green, followed by collapse of the whole plant a few days later. Browning of vascular and pith tissues was evident, especially at the base of the stem. A whitish mass of bacteria oozed from the cut end of diseased stems, suggesting that bacteria might be the cause of this disease. A total of 15 bacterial strains were collected. Colonies on tetrazolium chloride medium (3) were round to oval and fluidal, each with a pink or red center after incubation at 30°C for 48 h. When tobacco leaves were infiltrated with these strains, a hypersensitive reaction (HR) typical of phytopathogenic bacteria was induced. All strains produced the expected amplicon (282 bp) after PCR with the Ralstonia solanacearum-specific primer pair, AU759f and AU760r (4). Three hexose alcohols (mannitol, sorbitol, and dulcitol), rather than three disaccharides (lactose, maltose, and cellobiose), were utilized, which suggests R. solanacearum biovar 4 (2). R. solanacearum phylotype I was determined by phylotype-specific multiplex PCR (1). Pathogenicity of the strains was tested on roselle, tomato, pepper, and eggplant. Young plants of the various species were inoculated at the four- to six-leaf stage by soil drenching with 30 ml of bacterial suspension (about 10 CFU/ml). Control plants were inoculated with sterile water. Each treatment comprised eight plants with a single plant in each pot. Plants were incubated in a greenhouse at 25 to 31°C and 56 to 93% humidity. Wilting was observed 4 to 6 days after inoculation, while the control did not wilt. To find the correlation between plant growth stage and resistance to the pathogen, 2-, 3-, 4-, and 5-week-old roselle plants cv. Chiada 1 were transplanted into artificially infested soil. Eight plants in each treatment were planted with a single plant in each pot. The disease incidences for plants of different ages were 75%, 62.5%, 50%, and 12.5%, respectively. Thi