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1 Institut für Angewandte Mikrobiologie, Justus-Liebig Universität Giessen, IFZ – Heinrich-Buff-Ring 26–32, D-35392 Giessen, Germany
2 College of Agriculture and Natural Resources, Department of Soil and Environmental Sciences, National Chung Hsing University, Taichung 402, Taiwan, ROC
3 Grup d'Oceanografia Interdisciplinari, Institut Mediterrani d'Estudis Avançats (CSIC-UIB), Esporles, Mallorca, Spain
Correspondence
Peter Kämpfer
peter.kaempfer{at}agrar.uni-giessen.de
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. Phenotypic tests were performed according to Kämpfer et al. (1991), and further physiological tests [API ZYM enzyme profiles (bioMérieux), API 20E (bioMérieux), Biolog GN2 system] were done according to the instructions of the relevant manufacturer. Flexirubin-like pigments were observed by flooding the plates with 20 % (w/v) potassium hydroxide (Fautz & Reichenbach, 1980). Fluorescence was tested 48 h after plating on King's B medium. DNA used for the determination of G+C content was isolated by using the UltraClean microbial DNA isolation kit according to the instructions of the manufacturer (MOBIO). The DNA G+C content was determined as described previously (Peña et al., 2005). Cell morphology was observed under a Zeiss light microscope at x1000 magnification, using cells that had been grown for 3 days at 30 °C on NA; the results are given in the species description. The 16S rRNA gene sequence was analysed as described previously (Kämpfer et al., 2003; Young et al., 2005).
Analysis of the sequence data was performed by using the software package MEGA, version 2.1 (Kumar et al., 2001), after multiple alignment of the data by CLUSTAL X (Thompson et al., 1997). A distance-matrix method (distance options according to the Kimura two-parameter model), including clustering by the neighbour-joining method (Fig. 1) and a discrete character-based maximum-parsimony method, was used. In each case, bootstrap values were calculated from 1000 replications. The 16S rRNA gene sequence of strain CC-BB4T was a continuous stretch of 1386 bp. Sequence-similarity calculations indicated that strain CC-BB4T showed the greatest degree of similarity with Labrys monachus VKM B-1479T (91.3 %), Labrys methylaminiphilus JLW10T (91.3 %) and Bradyrhizobium japonicum DSM 30131T (91.3 %). Lower sequence similarities (<91 %) were found with respect to members of all other genera shown in Fig. 1.
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, and the fatty acid profile of strain CC-BB4T (given in the species description) was found to be similar to those of the genera listed in Table 1, although significant differences were also found.
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; Kämpfer, 1990), are given in the species description and in Table 1
. The utilization of methanol, methylamine, formaldehyde and formamide (1 and 5 mM in each case) was tested in the medium described by Widdel & Bak (1992). The presence of the mxaF gene (McDonald & Murrell, 1997), which encodes the 
-subunit of the methanol dehydrogenase present in all methylotrophs (Hanson & Hanson, 1996; McDonald et al., 1995), was tested with the PCR assay described by McDonald & Murrell (1997); DNA extracted from Methylosinus trichosporium OB3bT was used as the positive control. No positive PCR amplification products were obtained with this test. In conclusion, strain CC-BB4T is not a methylotrophic bacterium. No flexirubin-like or fluorescent pigments were observed. Strain CC-BB4T was unable to produce acid from various carbohydrates, but tests with organic acids as the carbon substrates gave positive results. In the API 20E test, strain CC-BB4T was positive only for the Voges–Proskauer (acetoin) reaction; in the API ZYM enzyme profiling test, CC-BB4T was positive for esterase, esterase lipase, lipase (weakly), leucine arylamidase and naphthol-AS-BI-phosphohydrolase. On the basis of these results, we propose that strain CC-BB4T should be classified within a novel genus, Pseudolabrys gen. nov., as the type strain of Pseudolabrys taiwanensis sp. nov.
Description of Pseudolabrys gen. nov.
Pseudolabrys (Pseu.do.la'brys. Gr. adj. pseudes false; N.L. masc. n. Labrys a bacterial genus name; N.L. masc. n. Pseudolabrys the false Labrys).
Cells are Gram-negative, aerobic, non-motile and short rod-shaped. Reproduction of cells is by division and not by budding. Methanol, methylamine, formaldehyde and formamide are not used as sole sources of carbon. Growth occurs after 96 h incubation on TSA and NA at 30 °C. The major fatty acids are C16 : 0, C18 : 1
7c and C19 : 0 cyclo 8c. The DNA G+C content is 67 mol%. The type species is Pseudolabrys taiwanensis.
Description of Pseudolabrys taiwanensis sp. nov.
Pseudolabrys taiwanensis (tai.wan.en'sis. N.L. masc. adj. taiwanensis of Taiwan, where the type strain was isolated).
Shows the following properties in addition to those given in the genus description. Oxidase- and catalase-positive. The fatty acid profile of the type strain comprises iso-C15 : 0 (0.8 %), C16 : 0 (4.8 %), iso-C15 : 0 3-OH (2.0 %), iso-C17 : 19c (0.8 %), iso-C17 : 0 (6.1 %), C17 : 0 (2.9 %), C18 : 17c (24.1 %), C18 : 0 (2.9 %), 11-methyl C18 : 07c (1.2 %), C19 : 0 cyclo 8c (51.2 %), C18 : 1 2-OH (1.0 %) and C20 : 26,9c (0.8 %). Aesculin is not hydrolysed. Flexirubin-like pigments and fluorescence are absent. Acid production is negative from glucose, lactose, sucrose, D-mannitol, dulcitol, salicin, adonitol, inositol, rhamnose, maltose, trehalose, cellobiose, D-arabitol, D-mannose, sorbitol, L-arabinose, raffinose, D-xylose, methyl D-glucoside, erythritol and melibiose. The following carbon sources are utilized (method according to Kämpfer et al., 1991 and Biolog GN2): acetate, cis- and trans-aconitate, fumarate, DL-3-hydroxybutyrate, DL-lactate, L-malate (weakly), mesaconate (weakly), 3-hydroxybenzoate, succinic acid, bromosuccinic acid, succinamic acid, formic acid, -ketoglutaric acid, pyruvic acid methyl ester and succinic acid monomethyl ester. The following tests are negative (Biolog GN2): -cyclodextrin, dextrin, glycogen, Tweens 40 and 80, N-acetyl-D-galactosamine, N-acetyl-D-glucosamine, adonitol, L-arabinose, D-arabitol, D-cellobiose, i-erythritol, D-fructose, L-fucose, D-galactose, gentiobiose, -D-glucose, myo-inositol, -D-lactose, lactulose, maltose, D-mannitol, D-mannose, D-melibiose, methyl 
-D-glucoside, D-psicose, D-raffinose, L-rhamnose, D-sorbitol, sucrose, D-trehalose, turanose, xylitol, citric acid, D-galactonic acid lactone, D-galacturonic acid, D-gluconic acid, D-glucosaminic acid, D-glucuronic acid, - and 
-hydroxybutyric acids, p-hydroxyphenylacetic acid, itaconate, -ketobutyric acid, -ketovaleric acid, quinic acid, D-saccharic acid, sebacic acid, glucuronamide, L-alaninamide, D- and L-alanine, L-alanyl glycine, L-asparagine, L-aspartic acid, L-glutamic acid, glycyl L-aspartic acid, glycyl L-glutamic acid, L-histidine, hydroxy-L-proline, L-leucine, L-ornithine, L-phenylalanine, L-proline, L-pyroglutamic acid, D- and L-serine, L-threonine, DL-carnitine, -aminobutyric acid, urocanic acid, inosine, uridine, thymidine, phenylethylamine, putrescine, 2-aminoethanol, 2,3-butanediol, glycerol, DL--glycerol phosphate, -D-glucose 1-phosphate, D-glucose 6-phosphate, salicin, p-arbutin, D-xylose, maltitol, adipate, pyruvate, azelate, suberate, L-tryptophan, 4-hydroxybenzoate and phenylacetate. Positive for hydrolysis of L-alanine p-nitroanilide (pNA). Negative for hydrolysis of p-nitrophenyl (pNP) -D-glucopyranoside, o-nitrophenyl D-galactopyranoside (ONPG), pNP -D-glucuronide, pNP -D-glucopyranoside, pNP -D-xylopyranoside, bis-pNP phosphate, pNP phenylphosphonate, pNP phosphorylcholine, 2-deoxythymidine-5'-pNP phosphate, L-glutamate--3-carboxy pna and L-proline pNA (method according to Kämpfer et al., 1991). Positive results (API ZYM) are seen for activities of esterase, esterase lipase, lipase (weak), leucine arylamidase and naphthol-AS-BI-phosphohydrolase, while results are negative for alkaline phosphatase, valine arylamidase, cystine arylamidase, trypsin, -chymotrypsin, acid phosphatase, -galactosidase, -galactosidase, -glucuronidase, -glucosidase, -glucosidase, N-acetyl--glucosaminidase, -mannosidase and -fucosidase. Additional physiological features are given in Table 1.
The type strain, CC-BB4T (=CCUG 51779T=CIP 108932T), was isolated from soil from Sinshe in Taichung County, Taiwan.
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ACKNOWLEDGEMENTS |
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REFERENCES |
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TOPABSTRACTMAIN TEXTREFERENCES |
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Young, C.-C., Kämpfer, P., Shen, F.-T., Lai, W.-A. & Arun, A. B. (2005). Chryseobacterium formosense sp. nov., isolated from the rhizosphere of Lactuca sativa L. (garden lettuce). Int J Syst Evol Microbiol 55, 423–426.
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