Isolation and identification of the causal agents of blackleg disease of potato occurred in Nagasaki Prefecture, Japan

Recently, blackleg of potatoes occurred in the fields of Minamishimabara, Shimabara, and Unzen City in Nagasaki Prefecture. Fourteen bacteria were isolated from the potatoes showing blackleg symptoms. When the isolates were inoculated to the stems of potatoes, the original symptoms were reproduced. Moreover, the colonies of reisolated bacteria from the inoculated potatoes showed exactly the same morphological characteristics as the inoculated bacteria. These isolates were analyzed for bacterial and biochemical prop-erties including gram reaction, anaerobic growth, yellow pigment production on yeast dextrose carbonate medium, growth in 5% NaCl, erythromycin sensitivity, acid production from maltose and lactose, utilization of malonate growth in nutrient agar at 37 degrees C, and reduction of sucrose. According to the properties, 12 iso-lates responded similarly to Pectobacterium brasiliense and two isolates did to P. parmentieri. The 16S ribosomal RNA, recA, and dnaX genes were amplified by polymerase chain reaction, and the amplicons were sequenced. Homology search results and phylogenetic analyses based on the nucleotide sequences of the genes supported the identification by the bacterial and biochemical properties. It was concluded that the causal agents of blackleg of potato occurred in Nagasaki Prefecture were identified as P. brasiliense and P. parmentieri. Since no subsequent outbreaks of the disease have been observed in Nagasaki Prefecture, it is unlikely that the disease was caused by indigenous pathogens. Diversity analysis using various strains worldwide and further epidemiological studies are also required to identify the origin of the pathogens.