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Glycomyces Halotolerans Sp. Nov., a Novel Actinomycete Isolated from a Hypersaline Habitat in Xinjiang, China

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Glycomyces Halotolerans Sp. Nov., a Novel Actinomycete Isolated from a Hypersaline Habitat in Xinjiang, China Antonie van Leeuwenhoek (2011) 100:137–143 DOI 10.1007/s10482-011-9574-1 ORIGINAL PAPER Glycomyces halotolerans sp. nov., a novel actinomycete isolated from a hypersaline habitat in Xinjiang, China Tong-Wei Guan • Zhan-Feng Xia • Jing Xiao • Nan Wu • Zheng-Jun Chen • Li-Li Zhang • Xiao-Ping Zhang Received: 12 December 2010 / Accepted: 4 March 2011 / Published online: 12 March 2011 Ó Springer Science+Business Media B.V. 2011 Abstract A novel actinomycete strain, designated all previously described representatives of the genus TRM 40137T, was isolated from a hypersaline habitat in Glycomyces. The whole-cell sugar pattern consisted of Xinjiang Province, north-west China, and subjected to a xylose and galactose. The predominant menaquinone polyphasic taxonomic study. The strain was aerobic, was MK-10(H2) and the major fatty acids were anteiso- Gram-positive and the optimum NaCl concentration for C15:0 and anteiso-C17:0. The phospholipid pattern growth was 4–5% (w/v). Phylogenetic analysis showed consists of phosphatidylglycerol, diphosphatidylglyc- that strain TRM 40137T has a 16S rRNA gene sequence erol, three unknown aminophospholipids and two similarity of 95.02% with the described species Glyc- unknown phospholipids. The G?C content of the omyces sambucus E71T and can be distinguished from genomic DNA was 68.8 mol%. A novel species Glycomyces halotolerans sp. nov. is proposed, with T T T strain TRM 40137 (=CCTCC AA 2010013 = KCTC The 16S rRNA gene sequence of strain TRM 40137 has been T deposited in GenBank under the accession number HQ651156. 19988 ) as the type strain of G. halotolerans. Electronic supplementary material The online version of Keywords Glycomyces halotolerans sp. nov. Á this article (doi:10.1007/s10482-011-9574-1) contains Polyphasic taxonomy Á 16S rRNA supplementary material, which is available to authorized users. T.-W. Guan (&) Á X.-P. Zhang (&) J. Xiao Faculty of Resource and Environmental Sciences, Key Laboratory of Marine Biological Resources, Sichuan Agricultural, University, 625000 Yaan, Third Institute of Oceanography, State Oceanic People’s Republic of China Administration, Xiamen, Fujian 361005, e-mail: [email protected] People’s Republic of China X.-P. Zhang N. Wu e-mail: [email protected] Key Laboratory of Biogeography and Bioresource in Arid Land, Xinjiang Institute of Ecology and Geography, T.-W. Guan Á Z.-F. Xia Á Z.-J. Chen Á L.-L. Zhang Chinese Academy of Sciences, 830011 Urumqi, Key Laboratory of Protection and Utilization People’s Republic of China of Biological Resources in Tarim, Basin of Xinjiang Production & Construction Corps, Tarim University, 843300 Alar, Xinjiang, People’s Republic of China T.-W. Guan State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, 100101 Beijing, People’s Republic of China 123 138 Antonie van Leeuwenhoek (2011) 100:137–143 Introduction medium. SN medium contained (g l-1): sucrose, 30.0 g; NaNO3, 2.0 g; K2HPO4, 1.0 g; MgSO4, 0.5 g; The genus Glycomyces, which belongs to the family KCl, 0.5 g; NaCl, 50 g; agar, 18.0 g; pH 7.0–7.5. The Glycomycetaceae of the suborder Glycomycineae range of growth temperature was tested at 5–60°Con (Stackebrandt et al. 1997), was originally established ISP medium 4 containing 5% (w/v) NaCl. For NaCl by Labeda et al. (1985) and the description was tolerance experiments, ISP medium 4 was used as the later emended by Labeda and Kroppenstedt (2004). basal medium and salt concentrations ranging from The genus Glycomyces consists of 10 validly pub- 0–20% (w/v) at intervals of 1% were evaluated. The lished species, namely G. algeriensis, G. arizonensis, pH range of growth was investigated between G. harbinensis, G. lechevalierae, G. rutgersensis, 4.0–12.0 at intervals of 1 pH unit using the following G. tenuis, G. sambucus, G. mayteni, G. endophyticus buffer systems: pH 4.0–5.0: 0.1 M citric acid/0.1 M and G. scopariae. sodium citrate; pH 6.0–8.0: 0.1 M KH2PO4/0.1 M (Labeda et al. 1985; Evtushenko et al. 1981; NaOH; pH 9.0–12.0: 0.1 M NaHCO3/0.1 M Na2CO3. Labeda and Kroppenstedt 2004; Gu et al. 2007; Qin Media and procedures used for determination of et al. 2008; Qin et al. 2009) and it is the type genus of physiological features and carbon source utilization the family Glycomycetaceae. Here, we report a novel were those described by Gordon et al. (1974). Glycomyces-like strain isolated from a hypersaline Antibiotic susceptibility was determined by the habitat in Xinjiang Province, north-west China, method of Williams (1967). designated TRM 40137T. The aim of the study was to determine the exact taxonomic position of the Chemotaxonomy strain by means of a polyphasic characterization. Biomass for chemotaxonomic studies was obtained after incubation at 37°C for 7 d in shake flasks of Materials and methods trypticase soy liquid medium with 5% (w/v) NaCl. Strain TRM 40137T was investigated as described Organism previously (Lee 2006) to determine the type of diamino acid in cell-wall hydrolysates (Staneck and Strain TRM 40137T was isolated from a soil sample Roberts 1974). Polar lipids were extracted, examined collected from the Lop Nur salt lake, Xinjiang by two dimensional TLC and identified using the Province, north-west China (GPS coordinates for 0 00 0 00 procedures of Minnikin et al. (1984). Procedures for the sampling site are 39°41 79 N89°54 50 E), by identification of the cell-wall sugars in whole-cell using SSC medium. The lake environment was hydrolysates were performed by using method as described previously (Guan et al. 2010). SSC medium described by Hasegawa et al. (1983). Menaquinones contains (g l-1): starch, 1.0 g; sucrose, 5.0 g; casein were extracted using the method of Collins et al. hydrolysate acid, 0.1 g; KNO , 0.5 g; CaCO , 0.1 g; 3 3 (1977) and analysed by HPLC (Groth et al. 1997). K HPO , 1.0 g; MgCl , 0.5 g; NaCl, 100 g; agar, 2 4 2 Cellular fatty acid composition was determined as 18.0 g; pH 7.0–7.5. The organism was grown and described by Sasser (1990) using the Microbial maintained on ISP 4 medium (Shirling and Gottlieb Identification System (MIDI, Inc.). 1966) containing 5% (w/v) NaCl. Phenotypic characteristics Molecular analysis Cultural characteristics were observed on the media Amplification and sequencing of the 16S rRNA gene of Shirling and Gottlieb (1966) amended with 5% sequence was performed as described according to NaCl after incubation at 37°C for 1–2 weeks. The Cui et al. (2001) and PCR products were purified colony colour was determined using the ISCC-NBS using a PCR purification kit (Sangon, Shanghai, colour charts (Kelly 1964) and morphological char- China). The almost-complete 16S rRNA gene acteristics of mycelia were examined by scanning sequence of strain TRM 40137T (1431 nucleotides) electron microscopy of 15 day cultures grown on SN was determined and deposited in the GenBank 123 Antonie van Leeuwenhoek (2011) 100:137–143 139 database as HQ651156. Multiple alignments with Results and discussion sequences of most closely related Glycomyces and calculations of levels of sequence similarity were A novel actinomycete strain, designated TRM 40137T, carried out on the EzTaxon server 2.0 (Chun et al. was isolated from a hypersaline habitat in Xinjiang 2007). The sequence of strain TRM 40137T was Province, north-west China, and subjected to a poly- manually aligned with the 16S rRNA gene sequences phasic taxonomic study. The strain was aerobic, Gram- of all recognized species in the family Glycomycet- stain positive and filamentous (Supplementary Fig. 1). aceae. Phylogenetic analysis was performed using The sequence analysis of the 16S rRNA gene indicated the software package MEGA version 4.0 (Tamura that strain TRM 40137T formed a distinct lineage et al. 2007). Phylogenetic trees were constructed within the family Glycomycetaceae and always had the using the neighbour-joining (Saitou and Nei 1987) closest phylogenetic affinity to the genus Glycomyces and maximum-parsimony (Fitch 1971) methods. with high levels of bootstrap support (Fig. 1; (Supple- Evolutionary distance was generated as described mentary Fig. 2). The 16S rRNA gene sequence of by Kimura (1980). The topology of the phylogenetic strain TRM 40137T showed 95.02% similarity to tree was evaluated by the bootstrap resampling G. sambucus and 93.34–94.98% to the other represen- method of Felsenstein (1985) with 1,000 replicates. tative strains of the genus Glycomyces. The strain had The genomic DNA of strain TRM 40137T for the lower 16S rRNA gene sequence similarities with determination of G?C content was prepared accord- Haloglycomyces albus (92.05%) and the described ing to the method of Marmur (1961). The DNA G?C Stackebrandtia species (90.22–90.92%). The G?C content of strain TRM 40137T was determined by content of the genomic DNA is 68.8 mol% which is using the HPLC method of Mesbah et al. (1989). consistent with the values for the genus Glycomyces. 71 Glycomyces lechevalierae NRRL B-16149T (AY462041) 99 Glycomyces rutgersensis IFO 14488T (D85484) 80 Glycomyces algeriensis NRRL B-16327T (AY462044) 100 Glycomyces endophyticus YIM 56134T (EU200681) 99 Glycomyces harbinensis IFO 14487T (D85483) Glycomyces mayteni YIM 61331T (EU814511) 55 98 Glycomyces sambucus DSM 45047T (DQ460469) 100 T Glycomyces scopariae YIM 56256 (EU200682) 62 Glycomyces arizonensis NRRL B-16153T (AY462042) 97 T 100 Glycomyces tenuis IFO 15904 (D85482) TRM 40137 T (HQ651156) Haloglycomyces albus YIM 92370T (EU660053) Stackebrandtia albiflava YIM 45751T (DQ985165) 86 T Stackebrandtia nassauensis DSM 44728 (AY650268) Pimelobacter simplex DSM 20130T (Z78212) 0.01 Fig. 1 Phylogenetic tree of strain TRM 40137T and its near the neighbour-joining method of Saitou and Nei (1987).
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