First Report of Pseudomonas cichorii Associated with Leaf Spot on Soybean in South Korea.

Unusual symptoms were observed on 5% of the soybean (Glycine max (L.) Merill) plants in commercial fields in South Korea in September 2008 and 2009. The lesions were at first water soaked, then enlarged and turned dark brown or black, often with concentric white rings and sometimes surrounded by a bright yellow halo. Most lesions were roughly circular to irregular. Six bacteria were isolated on tryptic soy agar (TSA) media from plants of soybean cv. Daewon. The isolate JBC1 formed colonies that were whitish to greenish, circular with convex elevation, and unable to grow anaerobically or at 37°C. Cells of the isolate were rod shaped with polar multitrichous flagella. The isolate was positive for the following characteristics: production of fluorescent and diffusible pigment on King's medium B, production of oxidase, ability to rot potato, and utilization of l-arabinose, d-aspartate, citrate, galactose, glucose, meso-inositol, mannitol, and meso-tartrate. However, the isolate was negative in the following tests: formation of yellow colonies on peptone sucrose agar and yeast extract dextrose calcium carbonate agar media, levan formation, production of nondiffusible pigment, urease and arginine dihydrolase, hydrolysis of starch, nitrate reduction, and utilization of d-arabinose, benzoate, geraniol, l-rhamnose, d-sorbitol, sucrose, and trehalose (1). The isolates elicited a clear hypersensitive reaction in tobacco leaves. The carbon source oxidation pattern analyzed by Biolog (Hayward, CA) GN database indicated that the isolate belonged to Pseudomonas cichorii (Swingle 1925) Stapp 1928. DNA was isolated with a commercial genomic DNA extraction kit (Solgent, Daejeon, South Korea) and the 16S rDNA was amplified using universal 27F and 1492R primers. The 1,367-bp amplicon was sequenced (GenBank Accession No. JF951725) and had 100% sequence identity with P. cichorii Accession No. EF101976.1. Phylogenetic analysis based on 1,367 bp of the 16S rDNA sequence also revealed that isolate JBC1 was closely related to P. cichorii. Healthy soybean plants of cv. Jangyeop were spray inoculated with 20 ml of a 10 suspension prepared from a 2-day-old culture grown on TSA. Healthy plants sprayed with just water served as noninoculated checks. Typical disease symptoms that were observed in the field developed on leaves of soybean plants 3 days after spray inoculation, while the check plants remained symptomless. The bacteria reisolated from inoculated plants were confirmed to be identical to the original strain by 16S rDNA analysis and the Biolog test. Inoculation on tomato, watermelon, melon, and oriental melon plants resulted in dark brown or black lesions forming on leaf margins and tips, which is different from those on soybean plants (2,3). With paprika and eggplant, necrotic spots with concentric white rings developed on the leaves of each plant. We propose leaf spot as the name for this disease on soybean. To our knowledge, this is the first report of leaf spot on soybean caused by P. cichorii, which is becoming a more important pathogen in subtropical regions and greenhouses (4). References: (1) B. Cottyn et al. Syst. Appl. Microbiol. 32:211, 2009. (2) A. Obradovic et al. Plant Dis. 86:443, 2002. (3) E. Pauwelyn et al. J. Phytopathol. 159:298, 2011. (4) S. M. Walkil et al. Nigeria J. Appl. Biosci. 38:2540, 2011.