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1 Department of Biological Sciences, Korea Advanced Institute of Science and Technology, 373-1, Guseong-dong, Yuseong-gu, Daejeon 305-701, Republic of Korea
2 Department of BioEnvironmental Chemistry, College of Agriculture and Life Sciences, Chungnam National University, Gung-dong 22, Yuseong-gu, Daejeon 305-764, Republic of Korea
Correspondence
Min-Ho Yoon
mhyoon{at}cnu.ac.kr
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ABSTRACT |
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A comparison of the fatty acid profile of strain Gsoil 616T and related type strains is available as supplementary material with the online version of this paper.
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MAIN TEXT |
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and, at the time of writing, it comprises 20 recognized species, including the recently described species Nocardioides furvisabuli (Lee, 2007) and Nocardioides insulae (Yoon et al., 2007). Strain Gsoil 616T was isolated from soil of a ginseng field in Pocheon province, South Korea, during the study of bacterial populations and was tentatively identified as Nocardioides sp. (Im et al., 2005). The isolation procedure and medium were described by Cui et al. (2007). Strain Gsoil 616T was routinely cultured on R2A agar (Difco) or half-strength R2A agar at 30 °C and maintained in glycerol suspension (20 %, w/v) at –70 °C.
The Gram reaction was performed by the non-staining method as described by Buck (1982). Cell morphology was observed at x1000 magnification with a light microscope (Nikon) using cells grown for 3 days at 30 °C on R2A agar. Motility was tested using the hanging drop technique. Catalase activity was determined by bubble production in 3 % (v/v) H2O2 and oxidase activity was determined using 1 % (w/v) tetramethyl p-phenylenediamine. Carbon-source utilization and enzyme activities were tested by using the API 20 NE, API ID 32 GN, API 50 CH and API ZYM test kits (bioMérieux). Anaerobic growth was tested in serum bottles with sodium thioglycolate (1 g l–1) added to R2A broth and replacing the upper air layer with nitrogen gas. Tests for the degradation of DNA [with DNase agar (Scharlau); performed by flooding plates with 1 M HCl], casein, chitin, starch (Atlas, 1993), xylan and cellulose (Ten et al., 2004) were evaluated after 2 weeks. Growth at different temperatures (4, 15, 20, 25, 30, 37 and 42 °C) and pH values (pH 5.0–10.0 at intervals of 0.5 pH units) was assessed after 5 days incubation. Salt tolerance was tested on R2A agar supplemented with 1–10 % (w/v) NaCl after 5 days incubation. Growth on nutrient agar, trypticase soy agar (TSA) and MacConkey agar was also evaluated, at 30 °C.
Cells of strain Gsoil 616T were strictly aerobic, Gram-positive, non-motile, non-spore-forming and rod- or coccoid-shaped. Colonies grown on R2A agar plates for 3 days were smooth, circular, white and 1–3 mm in diameter. The isolate grew well on nutrient agar and TSA, whereas it could not grow on MacConkey agar. Other physiological characteristics of strain Gsoil 616T are summarized in the species description. Phenotypic characteristics that differentiate the isolate from its closest phylogenetic relatives are listed in Table 1.
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). The G+C content of genomic DNA of strain Gsoil 616T was 73 mol%. Cellular fatty acids were determined in a culture grown on R2A agar for 6 days. The fatty acid profile of strain Gsoil 616T (compared with those of related type strains in Supplementary Table S1 in IJSEM Online) was composed of branched-chain fatty acids iso-C16 : 0 (45.5 %), iso-C15 : 0 (3.8 %), iso-C14 : 0 (2.9 %), iso-C15 : 0 3-OH (1.5 %), iso-C17 : 0 (0.9 %), anteiso-C15 : 0 (0.8 %), anteiso-C17 : 0 (0.8 %) and iso-C18 : 0 (0.5 %), unsaturated fatty acids C17 : 1
6c (7.5 %), C17 : 18c (3.6 %), C18 : 19c (2.8 %) and C18 : 36c (6,9,12) (2.1 %), saturated fatty acids C16 : 0 (6.4 %), C17 : 0 (5.9 %), C18 : 0 (4.4 %), C15 : 0 (1.6 %) and C14 : 0 (0.9 %), 10-methyl fatty acids C17 : 0 (2.8 %), C16 : 0 (1.0 %) and C18 : 0 (0.9 %) and summed feature 7 (any combination of C18 : 17c, C18 : 19t and C18 : 112t) (1.4 %), summed feature 3 (iso-C15 : 0 2-OH and/or C16 : 17c) (1.1 %) and summed feature 6 (C18 : 26,9c and/or anteiso-C18 : 0) (0.4 %). This fatty acid profile, including the major fatty acid iso-C16 : 0 and three 10-methyl fatty acids, is similar to those of Nocardioides species, although there are differences in the proportions of some fatty acids, perhaps because of differences in the extraction and cultivation conditions (Yoon et al., 1997, 2004; Yi & Chun, 2004a, b). LL-Diaminopimelic acid and the predominant menaquinone MK-8(H4) were also typical of members of the genus Nocardioides (Yoon et al., 1997, 2004, Yi & Chun, 2004a, b).
Extraction of genomic DNA, PCR-mediated amplification of the 16S rRNA gene and sequencing of the purified PCR product were carried out according to Kim et al. (2005). The full sequence of the 16S rRNA gene was compiled using SeqMan software (DNASTAR). The 16S rRNA gene sequences of related taxa were obtained from GenBank. Multiple alignments were performed by using the CLUSTAL_X program (Thompson et al., 1997) and gaps were edited in the BioEdit program (Hall, 1999). Evolutionary distances were calculated using the Kimura two-parameter model (Kimura, 1983) and phylogenetic trees were constructed by using the neighbour-joining method (Saitou & Nei, 1987) using the MEGA3 program (Kumar et al., 2004) with bootstrap values based on 1000 replications (Felsenstein, 1985).
The nearly complete 16S rRNA gene sequence (1457 bp) of strain Gsoil 616T was determined and subjected to phylogenetic analysis, which indicated that strain Gsoil 616T belongs to the genus Nocardioides (Fig. 1). Strain Gsoil 616T showed highest 16S rRNA gene sequence similarity to Nocardioides luteus KCTC 9575T and Nocardioides aquiterrae GW-9T (96.1 %), followed by Nocardioides pyridinolyticus KCTC 0074BPT (95.5 %) and other species of Nocardioides (<95 %). These values (<97 %) were low enough to place strain Gsoil 616T within a novel species of the genus Nocardioides according to Stackebrandt & Goebel (1994).
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Description of Nocardioides panacihumi sp. nov.
Nocardioides panacihumi (pa.na.ci.hu'mi. N.L. n. Panax scientific name of ginseng; N.L. gen. n. panacis of ginseng; L. n. humus soil; N.L. gen. n. panacihumi of soil of a ginseng field).
Cells are Gram-positive, strictly aerobic and non-motile, 0.3–0.5 µm in diameter and 0.7–1.2 µm long after 3 days on R2A agar. Cells show rod-to-coccus morphogenesis from the early exponential phase to the stationary phase. Colonies grown on R2A agar (Difco) for 3 days are smooth, circular, convex, opaque and white. Neither substrate mycelium nor aerial mycelium is formed. Good growth occurs at 30 °C and at pH 7.0. Growth is observed at 15–30 °C, pH 5.0–8.0 and in 0–1 % (w/v) NaCl. Nitrate is reduced weakly under aerobic conditions. Anaerobic growth does not occur. DNA and casein are degraded but xylan, cellulose, starch and chitin are not. Substrate utilization, enzyme production and other physiological characteristics are indicated in Table 1
. In addition to the properties listed in Table 1, acetate, L-alanine, gluconate, glycogen, 3-hydroxybenzoate, 4-hydroxybenzoate, 3-hydroxybutyrate, maltose, phenylacetate, L-proline, propionate, L-rhamnose, salicin and valerate are utilized as sole carbon sources. N-Acetyl-
-glucosamine, adipate, L-arabinose, caprate, citrate, L-fucose, L-histidine, inositol, itaconate, 2-ketogluconate, 5-ketogluconate, L-lactate, malate, malonate, mannitol, D-melibiose, D-ribose, L-serine, D-sorbitol and suberate are not utilized as sole carbon sources. In API 50 CH tests, acid is produced from amygdalin, L-arabinose, aesculin ferric citrate, D-glucose, D-fructose, D-lyxose, D-raffinose, L-rhamnose, sucrose and D-turanose, but acid is not produced from N-acetylglucosamine, L-arabinose, DL-arabitol, arbutin, adonitol, starch, D-cellobiose, dulcitol, erythritol, DL-fucose, D-galactose, gentiobiose, glycerol, glycogen, methyl 
-glucopyranoside, gluconate, inositol, inulin, 2- or 5-ketogluconate, D-lactose, D-lyxose, maltose, D-mannitol, D-mannose, methyl -D-mannoside, melezitose, melibiose, ribose, salicin, D-sorbitol, L-sorbose, D-tagatose, trehalose, xylitol, L-xylose or methyl -D-xyloside. According to the API ZYM gallery, the type strain is positive for N-acetyl--glucosaminidase, esterase lipase (C8) and -fucosidase and negative for -glucuronidase and naphthol-AS-BI-phosphohydrolase. According to the API 20NE gallery, the type strain is positive for -glucosidase but negative for arginine dihydrolase, -galactosidase, tryptophan degradation and glucose fermentation. The diagnostic diamino acid in the cell-wall peptidoglycan is LL-2,6-diaminopimelic acid. The predominant menaquinone is MK-8(H4). The major fatty acid (>45 % of total fatty acids) is iso-C16 : 0. The G+C content of genomic DNA of the type strain is 73 mol%. Other phenotypic characteristics are given in Table 1.
The type strain, Gsoil 616T (=KCTC 19187T =DSM 18660T), was isolated from soil of a ginseng field of Pocheon province, South Korea.
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ACKNOWLEDGEMENTS |
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Yi, H. & Chun, J. (2004a). Nocardioides ganghwensis sp. nov., isolated from tidal flat sediment. Int J Syst Evol Microbiol 54, 1295–1299.
Yi, H. & Chun, J. (2004b). Nocardioides aestuarii sp. nov., isolated from tidal flat sediment. Int J Syst Evol Microbiol 54, 2151–2154.
Yoon, J.-H., Rhee, S.-K., Lee, J.-S., Park, Y.-H. & Lee, S. T. (1997). Nocardioides pyridinolyticus sp. nov., a pyridine-degrading bacterium isolated from the oxic zone of an oil shale column. Int J Syst Bacteriol 47, 933–938.
Yoon, J.-H., Kim, I.-G., Kang, K. H., Oh, T.-K. & Park, Y.-H. (2004). Nocardioides aquiterrae sp. nov., isolated from groundwater in Korea. Int J Syst Evol Microbiol 54, 71–75.
Yoon, J.-H., Lee, C.-H. & Oh, T.-K. (2006). Nocardioides lentus sp. nov., isolated from an alkaline soil. Int J Syst Evol Microbiol 56, 271–275.
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