570 微生物学通报 2009, Vol.36, No.4,, [], [2],,,,, [3],,, 3 XJSS-2XJSS-39 XJSS-58 6S rrna, XJSS-2 XJSS-58 Streptomyces humidus subsp. Antitumoris Streptomyc

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1 微生物学通报 APR 20, 2009, 36(4): 569~574 Microbiology 2009 by Institute of Microbiology, CAS 研究报告三株沙漠放线菌 XJSS-2 XJSS-39 和魏艳红 XJSS-58 的多相分类及特性研究 2 阿不都克里木热依木熊鹰许尚营邓子厚袁永泽 ( ) ( ) * 刘德立摘要 : 从新疆吐鲁番鄯善县典型沙漠地区采集的样品中分离获得 3 株放线菌菌株 XJSS-2 XJSS-39 和 XJSS-58, 对其形态特征生理生化特性反硝化能力细胞壁化学组分以及 6S rrna 基因序列等进行了多相分类研究, 发现 3 株沙漠放线菌均能产生纤维素酶, 淀粉酶和蛋白酶, 且菌株 XJSS-2 具有很强的反硝化能力, XJSS-58 属于轻度耐盐菌初步确定 XJSS-2 XJSS-39 和 XJSS-58 可能分别属于 Streptomyces humidus subsp. antitumoris Nocardiopsis dassonvillei 和 Streptomyces mutabilis 关键词 : 沙漠放线菌, 链霉菌属, 拟诺卡氏菌属, 多相分类, 特性 Polyphasic Taxonomy and Characteristics of Three Desert Actinomycosis XJSS-2, XJSS-39 and XJSS-58 WEI Yan-Hong ABDUKERIM Rehim 2 XIONG Ying XU Shang-Ying DENG Zi-Hou YUAN Yong-Ze LIU De-Li * (. College of Life Sciences Laboratory of Genetic Regulation and Integrative Biology, Huazhong Normal University, Wuhan, Hubei , China) (2. College of Life Science and Chemistry, Xinjiang Normal University, Urumqi, Xinjiang , China) Abstract: Three actinomycete strains XJSS-2, XJSS-39 and XJSS-58 were isolated from soil sample collected from typical desert areas in Shanshan County, Turpan City, Xinjiang Province. Based on the polyphasic studies, including their morphology, physiological and biochemical characteristics, denitrifying capability, chemotaxonomy and 6S rrna gene sequence analysis, we found that three desert actinomycetes are able to produce cellulase, amylase and protease, the strain XJSS-2 has a strong capacity to denitrification, XJSS-58 belongs to the mild salt-tolerant bacteria. XJSS-2, XJSS-39 and XJSS-58 were primarily identified as Streptomyces humidus subsp. antitumoris, Nocardiopsis dassonvillei and Streptomyces mutabilis. Keywords: Desert actinomycosis, Streptomycetes, Nocardia, Polyphasic taxonomy, Characteristics, 基金项目 :(No. 2007AA30C26, 2007AA20C50); (No ) * 通讯作者 :Tel: ; : deliliu2002@yahoo.com.cn 收稿日期 : ; 接受日期 :

2 570 微生物学通报 2009, Vol.36, No.4,, [], [2],,,,, [3],,, 3 XJSS-2XJSS-39 XJSS-58 6S rrna, XJSS-2 XJSS-58 Streptomyces humidus subsp. Antitumoris Streptomyces mutabilis, XJSS-39 Nocardiopsis dassonvillei 材料和方法. 材料.. 菌株来源 :,, 50 mg/l HV [4], 30 C 5 d~7 d, 3, XJSS-2XJSS-39 XJSS 主要仪器和试剂 :PCR ( )(Bio-Rad)(Nicon); DNA dntppmd8-t EcoR Hind (TaKaRa ).2 形态和培养特征观察, 30 C 4 d,,, 30 C 2 d,.3 生理生化特性 ShirlingGottlieb [5] NaClISP5 [5], NaCl0%2%4%6%8%0% 2%,, 30 C 2 d 3 d~5 d, 2 30 d, 2 d~3 d, 2 ph ISP5, ph ph C 2 d, 3 d~5 d, 2.4 反硝化能力测定 2 [6], 3 0%SC [7], 30 C, 2 h4 h6 h8 h0 h 2 h, NO 3 NO 2, Brucine NO 3 [8], NO [9] 2.5 细胞壁化学组成分析 [0] (TLC).6 6S rrna 基因序列确定与分析.6. 总 DNA 的提取及 6S rrna 基因扩增 : DNAKutchma AJ [] 6S rrna PCRP5 -AGAGTTTG ATCCTGGCTCAG-3 ; P25 -GGTTACCTTGTTAC GACTT-3 95 C 30 s; 54 C 30 s, 72 C min 30 s, 30 ; 72 C 5 minpcr %.6.2 6S rrna 基因序列测定及分析 : PCR, pmd8-t, 0 µl (T µl, PCR 5 µl, 4 µl), 6 C 0 µl E. coli DH5α, LB, 37 C

3 : XJSS-2XJSS-39 XJSS-58 57, EcoR/Hind EcoR,, Blast GenBank 6S rrna, MEGA 2 结果 2. 形态和培养特征 XJSS-2XJSS-39 XJSS-58,, XJSS-58 3 NaCl XJSS-2 XJSS-39 8%, XJSS-58 0%, 3 37 C XJSS-2 XJSS-39 ph 7.0, XJSS-58 ph 8.0 XJSS-2,,, ( A), XJSS-39,,, ( B),,, XJSS-58,,,, ( C),, XJSS-2XJSS-39 XJSS-58 图三株沙漠放线菌菌株在燕麦片琼脂培养基上培养的光镜照片 ( 000) Fig. Morphological characteristics of three desert actinomycosis on Oatmeal agar medium( 000) Note: A: XJSS-2; B: XJSS-39; C: XJSS-58. Table 表三株沙漠放线菌菌株的培养特征 Cultural characteristics of three desert actinomycosis Culture medium Oatmeal Agar Czapek Dox Agar Nutrient Agar Gause s No. Agar Potato Juice Agar Starch Inorganic Salt Agar Aerial hyphae Substrate mycelium Dissoluble pigment XJSS-2 XJSS-39 XJSS-58 XJSS-2 XJSS-39 XJSS-58 XJSS-2 XJSS-39 XJSS-58 Fawn Offwhite Light yellow Tawny Light brown Green gray Dark gray Off white Light yellow Light gray Light yellow Deep yellow Tan Sage green Dark gray Tan

4 572 微生物学通报 2009, Vol.36, No 生理生化特性 2 XJSS-2,,,,, Table 2 表 2 三株沙漠放线菌菌株部分生理生化特性 Physiological and biochemical properties of three desert actinomycosis Characteristic XJSS-2 XJSS-39 XJSS-58 Characteristic XJSS-2 XJSS-39 XJSS-58 Gelatin liquefiying Glucose Milk freezing + + Cellobiose Milk solidification + + Rhamnose + Hydrogen sulfide + + Arabinose Nitrate reduction Melibiose Amylolysis Raffinose Cellulase Maltose + + Urease + Glycerin Lecithin enzyme Inositol Oil hydrolysis + Sorbitol Melanin production + + Red Xianchun Rifampicin endurance (50 µg/ml) Xylitol Penicillin endurance (6 µg/ml) Mannitol Antibiotics production Aesculin D- D-mannose Degradation of xanthine D- D-fructose Degradation of tyrosine D- D-xylose / C The optimum growth temperature 37 C 37 C 37 C D- D-galactose + + ph The optimum growth ph Sugar NaCl NaCl tolerance 8% 8% 0% D- D-galactose + + Sodium lodohippurate + + +: ; :. Note: +: Positive reaction; : Negative reaction.

5 : XJSS-2XJSS-39 XJSS ,, XJSS-39 XJSS-58,,,,,, 2.3 反硝化能力测定 XJSS-2 SC, 8 h 0 mmol/lno 3, 00%, NO 2 ( 2) 2.5 6S rrna 基因序列分析 XJSS-2 6S rrna 494 bp, XJSS-39 6S rrna 492 bp, XJSS-58 6S rrna 499 bp 3 6S rrna GenBank, XJSS-2XJSS-39 XJSS-58 EU598255EU EU S rrna NCBI Blast, GenBank 6S rrna, MEGA Clustal W, Neighbor-joining ( 3) 3, XJSS-2, S. humidus subsp. antitumoris S. macrosporeus, XJSS-39, N. dassonvillei XJSS-58, S. mutabilis S. maritimus, 3 分析与讨论图 2 XJSS-2 反硝化能力测定 (NO 3 去除率 ) Fig. 2 Denitrification by XJSS-2 (rate of NO 3 removal) XJSS-39 XJSS-58 NO 3 SC,, NO 3, NO 细胞壁化学组成分析 XJSS-2 XJSS-58 LL-DAP,, XJSS-2 XJSS-58 XJSS-39 meso-dap, 3, XJSS-2 XJSS-58, XJSS S rrna GenBank, XJSS-2 S. humidus subsp. antitumoris S. macrosporeus, 99.73% 99.06%XJSS-58 S. mutabilis S. maritimus, 99.93% 99.3%XJSS-39 N. dassonvillei 99.65%, XJSS-2 S. humidus subsp. antitumoris, XJSS-39 N. dassonvillei, XJSS-58 S. mutabilis

6 574 微生物学通报 2009, Vol.36, No.4 图 3 基于 6S rrna 基因序列用 Neighour-joining 法构建的系统进化树 Fig. 3 Phylogenetic tree showing the relationships among type strains and experimental strains based on 6S rrna sequences with Neighour-joining method 0.0,. Note: The scale bar indicates 0.0 subtitutions per nucleotide position. Seqboot values were showed on the branches. 参考文献 [] Okami Y, Hotta K. Search and discovery of new antibiotics in actinomycetes in biotechnology (Goodfellow M, Williams ST and Mordarski M Eds). London: Academic press, 988, pp [2],.., 2004, 22(4): [3] Antranikian G, Aguilar A. First meeting on biotenchnology of extremophiles. Hamburg: Technical University Hamburg, Harburg, 993. [4] Hayakawa M, Nonomura H. Humic acid vitamin agar, a new medium for the selective isolation of soil actinomycetes. Fermemt Technol, 987, 65: [5] Shirling EB, Gottlieb D. Methods for characterization of Streptomyces species. Int J Syst Bacteriol, 966, 6: [6],,,.., 2007, 27(6): [7] Patureau D, Zumstein E, Delgenes JP. Aerobic denitrification isolation from diverse natural and managed ecosystems. Microbiol Ecol, 2000, 39: [8] APHA. Standard methods for the examination of water and wastewater. Washington DC, USA: American Public Health Association, 995. [9],,,.., 2004, 4: [0].., 986, 3(5): [] Kutchma AJ, Roberts MA, Knaebel DB, et al. Small-scale isolation of genomic DNA from Streptomyces mycelia of spores. Bio Tech, 998, 24:

Microsoft Word 李梓铭-已校对.doc

Microsoft Word 李梓铭-已校对.doc 5 4 Vol. 5 No. 4 2014 4 Journal of Food Safety and Quality Apr., 2014 1, 李梓铭 2 1,, 谢靓 2 2,, 蒋立文 3*, 李云倩 (1., 410128; 2., 410128; 3., 410128) 1 摘要 : 目的 MRS, 方法 18% NaCl, PCR 16S rdna, 结果,, 结论 16S rdna 关键词