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1 Korea Research Institute of Bioscience and Biotechnology, 52 Oeundong, Yusong, Daejeon 305-333, Republic of Korea
2 Division of Applied Life Science, EB-NCRC, PMBBRC, Gyeongsang National University, Jinju 660-701, Republic of Korea
Correspondence
Chang-Jin Kim
changjin{at}kribb.re.kr
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ABSTRACT |
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The GenBank/EMBL/DDBJ accession number for the 16S rRNA gene sequence of strain BH030017T is AY603978.
A transmission electron micrograph of a cell of strain BH030017T and a thin-layer chromatogram of the polar lipids of the novel strain are available as supplementary figures in IJSEM Online.
These authors contributed equally to this work. 
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TOPABSTRACTMAIN TEXTREFERENCES |
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; Nielsen et al., 1994; Ventosa et al., 1989; Yoon et al., 2004a). Among them, aerobic spore-forming, halophilic, Gram-positive rods that were originally assigned to the genus Bacillus are also taxonomically very diverse. Therefore, they have been reclassified as members of novel genera or transferred to other genera. The genus Bacillus contains six phylogenetically distinct groups on the basis of molecular analyses of 16S rRNA gene sequences (Ash et al., 1991; Stackebrandt & Liesack, 1993; Spring et al., 1996; Wainø et al., 1999; Schlesner et al., 2001) and they are attracting interest because this group of bacteria has great biotechnological potential for the production of compatible solutes or hydrolytic enzymes (Margesin & Schinner, 2001). In the course of screening of the surface sediment of a solar saltern in the Tae-An area of Korea, in order to isolate halophilic bacteria, an aerobic, Gram-positive, moderately halophilic bacterium, strain BH030017T, was isolated and subjected to taxonomic characterization.
Strain BH030017T was isolated from surface soil from a solar saltern. The soil samples were diluted serially using saline solution (0.9 %, w/v), spread on marine agar 2216 (MA; Difco) with the addition of 5 % (w/v) NaCl (final concentration 6.94 % NaCl, w/v) and incubated for 2 days at 35 °C. NaCl tolerance was investigated using nutrient broth (NB) (3.0 g beef extract and 5.0 g peptone l–1; Difco) medium containing artificial seawater (ASW) (l–1: 5.94 g MgSO4.7H2O, 4.53 g MgCl2.6H2O, 0.64 g KCl and 1.3 g CaCl2,) and various concentrations of NaCl (0–25 %, w/v). The optimum temperature for growth was tested in the range 4–55 °C on MA and at different pH values (pH 5.0–11.0) in NB supplemented with ASW containing 3 % (w/v) NaCl. Media with different pH values were prepared using the appropriate biological buffers, Na2HPO4/NaH2PO4 (below pH 8.0), Na2CO3/NaHCO3 (pH 8.0–10.0) and Na2HPO4/NaOH (pH 11.0), as described previously (Gomori, 1955). Anaerobic growth was determined by incubation in an anaerobic chamber at 35 °C for 5 days on MA.
Gram staining was determined using the bioMérieux Gram-stain kit according to the manufacturer's instructions. Catalase activity was determined by means of bubble production in a 3 % (v/v) aqueous hydrogen peroxide solution. Oxidase activity was tested using a strip containing tetramethyl-p-phenylenediamine (Merck). Nitrate reduction and the hydrolysis of starch, tyrosine, aesculin, urea, casein, Tween 80, hypoxanthine, gelatin, glycerol and xanthine were determined according to methods described previously (Lanyi, 1987; Gerhardt et al., 1994). Acid production from carbohydrates was determined as described by Leifson (1963); all suspension media were supplemented with ASW containing 3 % (w/v) NaCl. Cell morphology was studied using light microscopy (HFX-DX Labophot-2; Nikon) and transmission electron microscopy (JEM-1010; JEOL). Each agar-coated wet mount used for motility observations was prepared as described by Lee et al. (2005). For visualization of flagella, cells were mounted on Formvar-coated copper grids (Electron Microscopy Science), negatively stained with 2 % (w/v) uranyl acetate for 15 s and then subjected to transmission electron microscopy at 60 kV. Endospores were stained using the Schaeffer–Fulton method (Smibert & Krieg, 1981).
For quantitative analysis of the whole-cell fatty acids, strain BH030017T was cultivated on MA and on MA with the addition of 3 % (w/v) NaCl, for 2 days at 35 °C. GC analysis of the fatty acid methyl esters was performed according to the instructions supplied with the Microbial Identification System (MIDI; Microbial ID). Analyses of the peptidoglycan and the isoprenoid quinones were carried out using the methods described by Komagata & Suzuki (1987). Polar lipid analyses were carried out by the Identification Service and Dr B. J. Tindall of the Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH (Braunschweig, Germany). The DNA G+C content of strain BH030017T was determined by reversed-phase HPLC (series II 1090; Hewlett Packard) using the method of Tamaoka & Komagata (1984). DNA–DNA hybridization was carried out to evaluate the genomic DNA relatedness between the isolate and closely related species. The cells were grown aerobically on MA for 2 days at 35 °C and the genomic DNA was isolated and purified according to the method described by Yoon et al. (1996). Serial dilution of the purified genomic DNA, DNA–DNA hybridization and the detection of hybrids by enzyme immunoassay were performed according to the methods described by Lim et al. (2005b).
PCR amplification, sequencing and assembly of the 16S rRNA gene were carried out as described previously (DeLong, 1992). To determine the approximate phylogenetic affiliation of strain BH030017T, the 16S rRNA gene sequence (1370 nt) was compared with those available from GenBank, using the BLAST program in GenBank (National Center for Biotechnology Information; http://www.ncbi.nlm.nih.gov/blast/), and was aligned with closely related 16S rRNA gene sequences by using CLUSTAL W software (Thompson et al., 1994). Unmatched regions of the 5'- and 3'-ends from the alignment, which were caused by the different lengths of the 16S rRNA gene sequence data, were deleted. Phylogenetic trees were constructed using three different methods, the neighbour-joining, maximum-likelihood and maximum-parsimony algorithms available in PHYLIP, version 3.6 (Felsenstein, 2002). Evolutionary distance matrices were calculated according to the algorithm of the Kimura two-parameter model (Kimura, 1980) for the neighbour-joining method. Similarity calculations were obtained from comparisons of the 16S rRNA gene sequences of the novel strain and members of the family Bacillaceae, performed using the FASTA3 program (http://www.ebi.co.uk/fasta33). To evaluate the stability of the phylogenetic tree, a bootstrap analysis (1000 replications) was performed using the SEQBOOT, DNADIST, NEIGHBOR and CONSENSE programs in the PHYLIP package.
The cells of the isolate were strictly aerobic, motile, short rods (0.5–1.2 µm wide and 1.2–1.9 µm long) bearing single polar flagella after 2 days incubation at 35 °C on MA (see Supplementary Fig. S1 available in IJSEM Online). The strain grew in NB with the addition of 0–12 % (w/v) NaCl and showed optimum growth in NB containing 2–5 % (w/v) NaCl. The cells produced ellipsoidal terminal endospores within swollen sporangia, like the phylogenetically closely related relatives in the family Bacillaceae. Strain BH030017T showed obvious Gram-, catalase- and oxidase-positive reactions and reduced nitrate to nitrite.
Analysis of the cell-wall peptidoglycan showed that strain BH030017T possessed the A1
type, i.e. with meso-diaminopimelic acid as the diagnostic diamino acid, in common with the great majority of endospore-forming, Gram-positive, rod-shaped bacteria. The major isoprenoid quinone was MK-7 and the G+C content of the genomic DNA was 36 mol%. The cellular fatty acid profile of strain BH030017T was characterized as containing branched fatty acids such as anteiso-C15 : 0 (
32.2 %), iso-C15 : 0 (29.4 %) and iso-C14 : 0 (10.0 %) as the major fatty acids on MA; these branched fatty acids (members of the 14- to 17-carbon iso and anteiso series) are typical of the fatty acids found in the cell membranes of Bacillus species (Table 1) (Albert et al., 2005). The strain contained phosphatidylglycerol (PG), diphosphatidylglycerol (DPG) and phosphatidylethanolamine (PE) as the major polar lipids (see Supplementary Fig. S2 available in IJSEM Online). The major fatty acid profile, the major lipoquinone and the major polar lipids of strain BH030017T were typical of those found in members of the genus Bacillus (Priest et al., 1988; Heyrman et al., 2004, 2005; Wieser et al., 2005; Lim et al., 2006). The typical phenotypic and chemotaxonomic properties of strain BH030017T are compared with those of phylogenetically related relatives in Table 2.
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; Schlesner et al., 2001). A phylogenetic tree constructed using the neighbour-joining method showed that strain BH030017T formed a cluster with Bacillus clarkii DSM 8720T and Bacillus agaradhaerens DSM 8721T within the family Bacillaceae with a relatively low bootstrap value (51 %), meaning that the isolate can also form a cluster with other relatives in the Bacillaceae, such as members of the genus Pontibacillus (Fig. 1). The overall topology of the neighbour-joining tree was supported by the trees constructed using the maximum-likelihood and maximum-parsimony algorithms (data not shown). Strain BH030017T shared 16S rRNA gene nucleotide similarities of less than 95.1 % with respect to the closely related species shown in Fig. 1. DNA–DNA hybridization was carried out to evaluate the phylogenetic position of strain BH030017T within the family Bacillaceae; hybridization was carried out with the reference strains B. clarkii DSM 8720T (13 % hybridization), Bacillus humi DSM 16318T (16 %), Bacillus algicola KMM 3737T (14 %), Bacillus subtilis NCDO 1769T (7 %), Bacillus hwajinpoensis KCCM 41641T (12 %), Virgibacillus halodenitrificans DSM 10037T (6 %) and Pontibacillus chungwhensis KCTC 3890T (7 %). Thus, the phylogenetic position of the novel strain within the family Bacillaceae could not be determined definitively by DNA–DNA hybridization. However, on the basis of the phenotypic and phylogenetic properties, strain BH030017T represents a novel species within the genus Bacillus, for which the name Bacillus taeanensis sp. nov. is proposed.
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Cells are Gram-positive, spore-forming, strictly aerobic rods. Cells are approximately 0.5–1.2 µm wide and 1.2–1.9 µm long and motile by means of single polar flagella. Colonies are cream, smooth, slightly raised and circular on MA. Catalase-positive and oxidase-positive. Nitrate is reduced to nitrite. An ellipsoidal endospore is formed terminally in a swollen sporangium. Growth occurs at 15–50 °C (optimum, 35 °C), pH 5.5–9.0 (optimum, pH 7.5) and 0–12 % (w/v) NaCl (optimum, 2–5 %). The peptidoglycan type is A1, with meso-diaminopimelic acid as the diagnostic diamino acid. The major isoprenoid quinone is MK-7. Starch, tyrosine, aesculin and urea are hydrolysed. Hydrolysis of casein, L-hypoxanthine, Tween 80, xanthine and gelatin is not observed. Acids are produced from D-glucose, sucrose, D-melibiose, trehalose, maltose, D-raffinose, D-fructose, D-xylose, D-mannitol, D-mannose and inositol, but not from D-ribose, glycerol, L-arabinose, L-rhamnose, 
-D-lactose or adonitol. The predominant cellular fatty acids are anteiso-C15 : 0 (32.2 %), iso-C15 : 0 (29.4 %) and iso-C14 : 0 (10.0 %) on MA. Contains PG, DPG and PE as the major polar lipids. The DNA G+C content is 36 mol% (HPLC).
The type strain, BH030017T (=KCTC 3918T=DSM 16466T), was isolated from a solar saltern in the Tae-An area of Korea.
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ACKNOWLEDGEMENTS |
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REFERENCES |
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TOPABSTRACTMAIN TEXTREFERENCES |
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Arahal, D. R., Marquez, M. C., Volcani, B. E., Schleifer, K. H. & Ventosa, A. (1999). Bacillus marismortui sp. nov., a new moderately halophilic species from the Dead Sea. Int J Syst Bacteriol 49, 521–530.
Ash, C., Farrow, J. A. E., Wallbanks, S. & Collins, M. D. (1991). Phylogenetic heterogeneity of the genus Bacillus as revealed by comparative analysis of small-subunit ribosomal-RNA sequences. Lett Appl Microbiol 13, 202–206.
DeLong, E. F. (1992). Archaea in coastal marine environments. Proc Natl Acad Sci U S A 89, 5685–5689.
Felsenstein, J. (2002). PHYLIP (phylogeny inference package), version 3.6a. Distributed by the author. Department of Genome Sciences, University of Washington, Seattle, USA.
Gerhardt, P., Murray, R. G. M., Wood, W. A. & Krieg, N. R. (1994). Phenotypic characterization. In Methods for General and Molecular Bacteriology, pp. 607–654. Edited by P. Gerhardt. Washington, DC: American Society for Microbiology.
Gomori, G. (1955). Preparation of buffers for use in enzyme studies. Methods Enzymol 1, 138–146.
Heyrman, J., Balcaen, A., Rodriguez-Diaz, M., Logan, N. A., Swings, J. & De Vos, P. (2003). Bacillus decolorationis sp. nov., isolated from biodeteriorated parts of the mural paintings at the Servilia tomb (Roman necropolis of Carmona, Spain) and the Saint-Catherine chapel (Castle Herberstein, Austria). Int J Syst Evol Microbiol 53, 459–463.
Heyrman, J., Vanparys, B., Logan, N. A., Balcaen, A., Rodríguez-Díaz, M., Felske, A. & De Vos, P. (2004). Bacillus novalis sp. nov., Bacillus vireti sp. nov., Bacillus soli sp. nov., Bacillus bataviensis sp. nov. and Bacillus drentensis sp. nov., from the Drentse A grasslands. Int J Syst Evol Microbiol 54, 47–57.
Heyrman, J., Rodríguez-Díaz, M., Devos, J., Felske, A., Logan, N. A. & De Vos, P. (2005). Bacillus arenosi sp. nov., Bacillus arvi sp. nov. and Bacillus humi sp. nov., isolated from soil. Int J Syst Evol Microbiol 55, 111–117.
Ivanova, E. P., Alexeeva, Y. A., Zhukova, N. V., Gorshkova, N. M., Buljan, V., Nicolau, D. V., Mikhailov, V. V. & Christen, R. (2004). Bacillus algicola sp. nov., a novel filamentous organism isolated from brown alga Fucus evanescens. Syst Appl Microbiol 27, 301–307.[CrossRef][Medline]
Kimura, M. (1980). A simple method for estimating evolutionary rates of base substitutions through comparative studies of nucleotide sequences. J Mol Evol 16, 111–120.[CrossRef][Medline]
Komagata, K. & Suzuki, K. (1987). Lipid and cell-wall analysis in bacterial systematics. Methods Microbiol 19, 161–208.
Lanyi, B. (1987). Classical and rapid identification methods for medically important bacteria. Methods Microbiol 19, 1–67.
Lee, E. M., Jeon, C. O., Choi, I., Chang, K. S. & Kim, C. J. (2005). Silanimonas lenta gen. nov., sp. nov., a slightly thermophilic and alkaliphilic gammaproteobacterium isolated from a hot spring. Int J Syst Evol Microbiol 55, 385–389.
Leifson, E. (1963). Determination of carbohydrate metabolism of marine bacteria. J Bacteriol 85, 1183–1184.
Lim, J.-M., Jeon, C. O., Song, S. M. & Kim, C.-J. (2005a). Pontibacillus chungwhensis gen. nov., sp. nov., a moderately halophilic Gram-positive bacterium from a solar saltern in Korea. Int J Syst Evol Microbiol 55, 165–170.
Lim, J.-M., Jeon, C. O., Park, D. J., Kim, H. R., Yoon, B. J. & Kim, C.-J. (2005b). Pontibacillus marinus sp. nov., a moderately halophilic bacterium from a solar saltern, and emended description of the genus Pontibacillus. Int J Syst Evol Microbiol 55, 1027–1031.
Lim, J.-M., Jeon, C. O., Lee, J. C., Ju, Y. J., Park, D. J. & Kim, C.-J. (2006). Bacillus koreensis sp. nov., a spore-forming bacterium, isolated from the rhizosphere of willow roots in Korea. Int J Syst Evol Microbiol 56, 59–63.[CrossRef][Medline]
Margesin, R. & Schinner, F. (2001). Potential of halotolerant and halophilic microorganisms for biotechnology. Extremophiles 5, 73–83.[CrossRef][Medline]
Nielsen, P., Rainey, F. A., Outtrup, H., Priest, F. G. & Fritze, D. (1994). Comparative 16S rDNA sequence analysis of some alkaliphilic bacilli and the establishment of a sixth rRNA group within the genus Bacillus. FEMS Microbiol Lett 117, 61–66.[CrossRef]
Nielsen, P., Fritze, D. & Priest, F. G. (1995). Phenetic diversity of alkaliphilic Bacillus strains: proposal for nine new species. Microbiology 141, 1745–1761.
Priest, F. G., Goodfellow, M. & Todd, C. (1988). A numerical classification of the genus Bacillus. J Gen Microbiol 134, 1847–1882.
Reva, O. N., Smirnov, V. V., Pettersson, B. & Priest, F. G. (2002). Bacillus endophyticus sp. nov., isolated from the inner tissues of cotton plants (Gossypium sp.). Int J Syst Evol Microbiol 52, 101–107.[Abstract]
Roberts, M. S., Nakamura, L. K. & Cohan, F. M. (1996). Bacillus vallismortis sp. nov., a close relative of Bacillus subtilis, isolated from soil in Death Valley, California. Int J Syst Bacteriol 46, 470–475.
Ruiz-García, C., Béjar, V., Martíez-Checa, F., Llamas, I. & Quesada, E. (2005). Bacillus velezensis sp. nov., a surfactant-producing bacterium isolated from the river Vélez in Málaga, southern Spain. Int J Syst Evol Microbiol 55, 191–195.
Schlesner, H., Lawson, P. A., Collins, M. D., Weiss, N., Wehmeyer, U., Völker, H. & Thomm, M. (2001). Filobacillus milensis gen. nov., sp. nov., a new halophilic spore-forming bacterium with Orn–D-Glu-type peptidoglycan. Int J Syst Evol Microbiol 51, 425–431.[Abstract]
Smibert, R. M. & Krieg, N. R. (1981). General characterization. In Manual of Methods for General Microbiology, pp. 409–443. Edited by P. Gerhardt, R. G. E. Murray, R. N. Costilow, E. W. Nester, W. A. Wood, N. R. Krieg & G. B. Phillips. Washington, DC: American Society for Microbiology.
Spring, S., Ludwig, W., Marquez, M. C., Ventosa, A. & Schleifer, K. -H. (1996). Halobacillus gen. nov., with description of Halobacillus litoralis sp. nov. and Halobacillus trueperi sp. nov., and transfer of Sporosarcina halophila to Halobacillus halophila comb. nov. Int J Syst Bacteriol 46, 492–496.
Stackebrandt, E. & Liesack, W. (1993). Nucleic acids and classification. In Handbook of New Bacterial Systematics, pp. 152–189. Edited by M. Goodfellow & A. G. O'Donnell. London: Academic Press.
Tamaoka, J. & Komagata, K. (1984). Determination of DNA base composition by reverse-phase high-performance liquid chromatography. FEMS Microbiol Lett 25, 125–128.
Thompson, J. D., Higgins, D. G. & Gibson, T. J. (1994). CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res 22, 4673–4680.
Ventosa, A., Garcia, M. T., Kamekura, M., Onishi, H. & Ruiz-Berraquero, F. (1989). Bacillus halophilus sp. nov., a moderately halophilic Bacillus species. Syst Appl Microbiol 12, 162–165.
Wainø, M., Tindall, B. J., Schumann, P. & Ingvorsen, K. (1999). Gracilibacillus gen. nov., with description of Gracilibacillus halotolerans gen. nov., sp. nov.; transfer of Bacillus dipsosauri to Gracilibacillus dipsosauri comb. nov., and Bacillus salexigens to the genus Salibacillus gen. nov., as Salibacillus salexigens comb. nov. Int J Syst Bacteriol 49, 821–831.
Wieser, M., Worliczek, H., Kämpfer, P. & Busse, H.-J. (2005). Bacillus herbersteinensis sp. nov. Int J Syst Evol Microbiol 55, 2119–2123.
Yoon, J.-H., Kim, H., Kim, S.-B., Kim, H.-J., Kim, W. Y., Lee, S. T., Goodfellow, M. & Park, Y.-H. (1996). Identification of Saccharomonospora strains by the use of genomic DNA fragments and rRNA gene probes. Int J Syst Bacteriol 46, 502–505.
Yoon, J. H., Kim, I. G., Kang, K. H., Oh, T.-K. & Park, Y. H. (2004a). Bacillus hwajinpoensis sp. nov. and an unnamed Bacillus genomospecies, novel members of Bacillus rRNA group 6 isolated from sea water of the East Sea and the Yellow Sea in Korea. Int J Syst Evol Microbiol 54, 803–808.
Yoon, J. H., Oh, T.-K. & Park, Y. H. (2004b). Transfer of Bacillus halodenitrificans Denariaz et al. 1989 to the genus Virgibacillus as Virgibacillus halodenitrificans comb. nov. Int J Syst Evol Microbiol 54, 2163–2167.
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