LAUSR

Hydrangeas (Hydrangea macrophylla) are recognized for their abundant, showy inflorescences which possess sepal colors (Schreiber et al., 2011). In recent years, the shrubs are grown as potted plants and also for landscaping in China. During the spring of 2021 in seedling greenhouse located in Tonghai County, Yunnan province, China (102°73' N, 24°12' E), a severe bacterial disease was observed on 12-month-old potted plants of cv. Enziandom. Symptoms on Hydrangeas initially appeared as thick liquid produced on the surface of the infected petiole base and water soaking in the infected stems. With the development of the infection, the basal stem color was dark brown and the stem base rapidly turned to wet rot. Severely infected leaves exhibited soft rot symptoms, and the presence of soft rot on mature plants decreased ornamental and market value. This disease was found in 40% of 15 surveyed greenhouses, and the disease incidence ranged from 30 to 50% in different regions. To identify the causal agent, disease-health tissues from infected plants were surface sterilized using 75% ethanol for 30 sec, then soaked in sterile distilled water for three times. The exudate from sheared tissue was streaked onto nutrient agar plates, and plates were incubated at 28°C for 24 hours (Koike et al., 2002). Four representative bacterial isolates BXH21032401-BXH21032404 were selected for further analysis. Colonies on NA were white, small, round and translucent with smooth edges. All bacterial isolates were gram-negative, rod-shaped, with 2-4 circumferential flagella and no entrapment membrane. Physiological and biochemical test and 96 phenotypes screened using the BIOLOG GENIII microplate system (Biolog, Hayward, CA, USA), the test results of four isolated strains are consistent with those of the reported Pectobacterium versatile (Portier et al. 2019). Total genomic DNA was extracted from bacterial suspensions with TIANamp Bacteria DNA Kit (TIANGEN, China). PCR amplification assays were conducted with specially designed primers using extracted genomic DNA as a template. All sequences of the PCR products were deposited in GenBank with accession numbers MZ852237-MZ852240 (16S rRNA), MZ921938-MZ921941 (pmrA), MZ926722-MZ926725 (acnA), MZ926726-MZ926729 (icdA), MZ926730-MZ926733 (gapA), MZ852233-MZ852236 (mdh), MZ926718-MZ926721 (mtlD), MZ921934-MZ921937 (pgi), MZ926734-MZ926737 (proA) and MZ921942-MZ921945 (rpoS). Maximum Likelihood phylogenetic trees based on 16S rRNA, pmrA sequences, and multilocus sequence analysis of housekeeping genes (acnA, icdA, gapA, mdh, mtlD, pgi, proA and rpoS) (Xie et al., 2018), all clustered BXH21032401-BXH21032404 into a clade containing other confirmed strains of P. versatile (Kravchenko et al., 2021). Pathogenicity tests were performed by spraying leaves of healthy 6-month-old potted H. macrophylla (cv. Enziandom) plants with bacterial suspension (20 mL/plant, 108 CFU ml-1). Plants were inoculated with sterile distilled water as control. Soft rot symptoms appeared on inoculated petioles in 5 days after incubation in growth chamber at 28°C showed similar symptoms as observed initially in the greenhouse, and strains were re-isolated successfully from symptomatic hydrangea to complete Koch's postulates. The pathogenicity test was carried out twice. To our knowledge, this is the first time that P. versatile is reported to cause hydrangea soft rot disease in China.