The taxonomic position of a Gram-positive, non-motile, non-spore-forming coryneform, isolated from activated sludge and designated strain CAU 212 T , was investigated using a polyphasic approach. Cellular morphology, biochemical tests and chemotaxonomic investigations revealed that strain CAU 212 T had the characteristics of the genus Corynebacterium. Comparative 16SrRNA gene sequence analysis showed that the organism formed a hitherto-unknown subline within the genus Corynebacterium. Sequence divergence values of more than 4.3 % from recognized Corynebacterium species, together with phenotypic differences, showed that the bacterium represents a previously unrecognized member of the genus Corynebacterium, for which the name Corynebacterium doosanense sp. nov. is proposed. The type strain is CAU 212The genus Corynebacterium was established by Lehmann & Neumann (1896) and represents one of the largest groups within the Actinobacteria. At the time of writing, Corynebacterium comprised 95 recognized taxa. In recent years, many of the recognized species within this genus originated from clinical specimens of humans (Hall et al., 2003;Renaud et al., 2007;Yassin, 2007;Yassin & Siering, 2008), animals (Goyache et al., 2003a, b; Fernández-Garayzábal et al., 2003Collins et al., 2004;Vela et al., 2006) and environments (Yassin et al., 2003; Chen et al., 2004;Feurer et al., 2004). Until now, however, there have been relatively few reports of corynebacteria from activated sludge. During the course of routine screening of bacteria for industrial purposes, a Corynebacterium-like strain, designated CAU 212 T , was isolated from activated sludge from the wastewater treatment plant in Yeongdeuk-gun, Republic of Korea.The isolation procedure for strain CAU 212 T followed that of Gordon & Mihm (1962) by using glucose-yeast extract agar (GYEA; comprising per litre: 10 g yeast extract, 10 g glucose and 15 g agar) supplemented with 50 mg cycloheximide l 21 and 20 mg nalidixic acid l
21. An activated sludge sample was diluted with sterilized distilled water and appropriate dilutions were spread onto the GYEA medium and incubated aerobically for 3 days at 30 u C. An opaque, yellow, low-convex bacterial colony with a diameter of 1-2 mm was subcultured on sheep blood agar plates (Asan Pharm Co.). The pure culture of CAU 212 T was preserved in 25 % (v/v) glycerol at 270 u C. The cell morphology of colonies was observed at different incubation times, every 24 h for 3 days, with a microscope following Gram staining.
Strain CAU 212T was characterized biochemically using the API Coryne, API 20 Strep and API ZYM kits (bioMérieux) according to the manufacturer's instructions. The ChristieAtkins-Munch-Petersen (CAMP) test with Staphylococcus aureus was performed as described by von Graevenitz & Funke (1996). Cell-wall murein was prepared by mechanical disruption of cells and complete acid hydrolysates were analysed as described by Schleifer & Kandler (1972). Cellular fatty acid methyl esters were extracted by acid methanolysis (Minnikin et al.,...
