The purple coneflower (Echinacea purpurea cv. Taichung No. 1) (Chen et al. , 2019), belonging to the Asteraceae family, has been cultivated on a total area of approximately 5 hectares in Yunlin and Hualien counties, primarily for processing into functional food supplements and flower tea products. In March 2022 and April 2023, soft rot symptoms were observed on the roots of purple coneflower plants. Aboveground tissues became desiccated due to water deficiency, and black rot developed at later stages. Disease incidence reached approximately 5-10% of the field. Five diseased plants were sampled in 2022 and 2023. The macerated stem base tissues were soaked in water and examined under a light microscope (400x), revealing motile, rod-shaped bacteria. Bacterial suspensions were streaked onto nutrient agar (NA), and incubated at 28°C for 1-2 days, producing round, gray-white, shiny and slimy colonies (0. 7-0. 9 mm in diameter). Strains TCPcf01, TCPcf05, and TCPcf07 from 2022, as well as TCPcf09 and TCPcf10 from 2023, were isolated. All strains produced pits on crystal violet pectate (CVP) medium, exhibited catalase and phosphatase activities and produced indole. The strains also showed facultative anaerobic characteristics based on oxidative and fermentation tests (Schaad et al. , 2001). Additionally, a 438-bp fragment was amplified using the Dickeya-specific primer pair 5A/5B (Chao et al. , 2006), and the corresponding sequences were deposited in GenBank (PX673856-PX673860). Further identification was conducted by sequencing the partial 16S rDNA (using primers 27F and 1492R) (Lane, 1991) and the housekeeping gene recN (Marrero et al. , 2013). The 16S rDNA and recN fragments were deposited in GenBank under accession numbers PV802530-PV802534 and PQ872932-PQ872936, respectively. Strains TCPcf01, TCPcf05, and TCPcf07 shared identical sequences for both genes, as did TCPcf09 and TCPcf10. The 16S rDNA sequences of TCPcf01 and TCPcf10 shared 99. 5% identity. Compared with the fully sequenced reference strain Dickeya chrysanthemi Ech1591 (CP001655), 16S rDNA identities were 99. 5% and 100%, respectively. The identities were 100% and 99. 5% when compared with the type strain D. chrysanthemi LMG2804ᵀ (= CFBP2048ᵀ; NR₁19368. 1). For recN gene, TCPcf01 and TCPcf10 (98. 7% identical) exhibited 98. 3% and 99. 6% identity to Ech1591, and 99. 5% and 98. 2% identity to CFBP2048ᵀ. To satisfy Koch’s postulate, puncture inoculation was performed using sterile toothpicks on the stem bases of 30-day-old purple coneflower plants with three plants per strain (TCPcf01 or TCPcf10). Each toothpick was dipped in a bacterial suspension (approximately 10⁶ CFU/ml), and sterile toothpicks served as controls. The inoculated plants were sealed in plastic bags for 2 days and then kept at 28°C. All inoculated plants developed soft rot symptoms at the stem base and roots, leading to water deficiency in the aboveground parts. Reisolated bacteria matched the recN sequences of TCPcf01 or TCPcf10. Prior to this study, no bacterial diseases had been documented on purple coneflower in Taiwan. Based on previous reports of the host range of Dickeya chrysanthemi (Samson et al, 2005), this study represents the first identification of Dickeya chrysanthemi pv. chrysanthemi causing soft rot of Echinacea purpurea (Asteraceae) in Taiwan.
Kuo et al. (Thu,) studied this question.