一株高效降解中长链烷烃菌株的分离鉴定及其降解特性研究
Isolation and Characterization of a Novel Medium-Long Alkane-Degrading Strain

作者: 唐赟 , 谭洪 :西华师范大学生命科学学院;

关键词: 石油污染烷烃降解分离鉴定不动杆菌Petroleum Pollution Hydrocarbon-Degrading Bacteria Isolation and Identification Acinetobacter

摘要: 从南充市炼油厂被石油污染土壤中分离到一株中长链烷烃降解菌TY12。采用形态学观察、生理生化指标测定、抗生素抗性实验、16S rRNA基因序列同源性分析等多种方法对该菌株进行鉴定。菌株为Acinetobacter beijerinckii,革兰氏阴性,短杆状,无荚膜,无芽孢,接触酶阳性,氧化酶阴性,菌体大小为0.9~1.6 μm × 1.5~2.5 μm;对新霉素、氯霉素、卡拉霉素等14种抗生素敏感,而对四环素、阿莫西林、氨苄西林耐药;在GenBank中与其16S rRNA基因序列相似度最高的模式株分别为Acinetobacter beijerinckii CCM7266 (AJ626712)、Acinetobacter beijerinckii LMH6214 (AJ303013),相似性为100%。研究表明,以工业乙醇为唯一碳源的无机盐培养基培养种子最适宜;其最适温度和pH分别为30℃和7.0,无水乙醇最适加入量为0.7%,最适酵母粉浓度为0.025 g/L。在初始正十二烷浓度为1%(W/V)培养基中接入1%种子液,于30℃、180 r/min震荡培养36 h,正十二烷降解率可达92%;菌株对正构烷烃的降解范围较宽,能以C12~C32的正构烷烃为唯一碳源和能源生长。该菌是一株能够降解中长链烷烃的不动杆菌新种,具有应用到石油污染修复的潜力。

Abstract: A strain of medium-long chain n-alkanes degrading strain TY12 was screened from petroleum-contaminated soils of Nanchong oil refinery. We adopted several phenotypic and genotypical methods, such as morphological, physical and biochemical characteristics, antibiotic susceptivities, 16S rRNA gene sequences based phylogenetic analysis, to outline the basic biological characteristics and determinate the phylogenetic position. The Gram-negative isolate TY12 was a member of the genus Acinetobacter, short rods, no capsules, no endospores, catalase-positive, oxidase-negative, 0.9 - 1.6 μm in diameter and 1.5 - 2.5 μm in length; TY12 was susceptible to 14 kinds of tested antibiotics but resistant to Tetracycline, Amoxicillin, Ampicillin; The similarity between its 16S rRNA gene and that of its most closely related type strain in GenBank Acinetobacter beijerinckii CCM7266 (AJ626712), Acinetobacter beijerinckii LΜH6214 (AJ303013) were 100%. The optimal temperature and pH for the strain utilizing industrial ethanol were 30˚C and 7.0, and the optimal concentration of ethanol and concentration of yeast extract were 0.7% and 0.025 g/L, respectively. The n-dodecane degradation was 92% after the strain was growing on in hydrocarbon degradation medium with 1% (W/V) of n-dodecane at 30˚C and 180 r/min for 36 h. The strain could degrade a large range of n-alkanes with chain length C12 - C32. It has potential in bioremediation of oil contaminated environment.

文章引用: 唐赟 , 谭洪 (2013) 一株高效降解中长链烷烃菌株的分离鉴定及其降解特性研究。 生物过程, 3, 9-15. doi: 10.12677/BP.2013.31002

参考文献

[1] 陆健, 黄潇, 伍贤明等. 高效烷烃降解菌xcz的分离鉴定及降解特性[J]. 土壤, 2008, 40(3): 460-464.

[2] V. Mishra, R. Lal and Srinivasan. Enzymes and operons mediating xenobiotic degradation in bacteria. Critical Reviews in Microbiology, 2001, 27(2): 133-166.

[3] N. Das, P. Chandran. Microbial degradation of petroleum hydrocarbon contaminants: An overview. Biotechnology Research International, 2011, 2011: Article ID 941810.

[4] S. Harayama, Y. Kasai and A. Hara. Microbial communities in oil contaminated seawater. Environmental Biotechnology, 2004, 15(3): 205-214.

[5] H. P. Kleber, R. Claus and O. Asperger. Enzymologie der n-alkanoxidation bei acinetobacter. Acta Biotechnologica, 1983, 3(3): 251-260.

[6] Y. Sakai, J. H. Maeng, Y. Tani and N. Kato. Use of long-chain n-alkanes (C13-C44) by an isolate, Acinetobacter sp. M-1. Bios-cience, Biotechnology, and Biochemistry, 1994, 58(11): 2128- 2130.

[7] R. G. Lageveen, G. W. Huisman, H. Preusting, et al. Formation of polyesters by Pseudomonas oleovarans: Effect of substrates on formation and Composition of Poly-(R)-3-Hydroyalkanoates and Poly-(R)-3-Hydroxyalkenoates. Applied and Environmental Microbiology, 1988, 54(12): 2924-2932.

[8] 黄磊, 李丹, 孙丹, 谢玉娟等. 一株低温石油烃降解菌的分类鉴定及降解特性研究[J]. 环境科学, 2007, 28(9): 2101-2102.

[9] F. C. Neidhardt, P. L. Bloch and D. F. Smith. Culture medium for enterobacteria. J. Bacterial, 1974, 119(3): 736-747.

[10] 仇天雷, 承磊, 邓宇等. 一株近海沉积物中产甲烷菌的分离及鉴定[J]. 中国沼气, 2007, 25(2): 3-6.

[11] 东秀珠, 蔡妙英. 常见细菌系统鉴定手册[M]. 北京: 科学出版社, 2001: 353-398.

[12] R. E. 布坎南, N. E. 吉本斯, 著, 中国科学院微生物研究所《伯杰细菌鉴定手册》翻译组, 译. 伯杰鉴定手册(第八版) [M]. 北京: 科学出版社, 1995: 591-611.

[13] J. 萨姆布鲁克, D. W. 拉赛尔, 著, 黄培堂等, 译. 分子克隆实验指南(第三版). 北京: 科学出版社, 2002.

[14] W. G. Weisburg, S. M. Barna, D. A. Pelletier, et al. 16S ribosomal DNA amplification for phylogenetic study. Journal of Bacteriology, 1991, 173(2): 697-703.

[15] 陈书霞, 王晓武, 房玉林. 单菌落PCR法直接快速鉴定重组克隆[J]. 微生物学通报, 2006, 33(3): 52-56.

[16] N. Saitou, M. Nei. The neighbor-joining method: A new method for reconstructing phylogenetic trees. Molecular Biology and Evolution, 1987, 4(4): 406-425.

[17] A. Nemec, M. Musilek, M. Maixnerova, et al. Acinetobacter beijerinckii sp. nov. and Acinetobacter gyllenbergii sp. nov., haemolytic organisms isolated from humans. Journal of Systematic and Evolutionary Microbiology, 2009, 59(1): 118-124.

[18] 吴业辉, 邵宗泽. 海洋烷烃降解菌Alcanivorax sp. A-11-3的分离鉴定及其降解酶基因研究[J]. 台湾海峡, 2008, 27(4): 427-434.

[19] 王丽萍, 刘昱慧, 邵宗泽. 一株来自大西洋表层海水的烷烃降解菌Gordonia sp. S14-10的分离鉴定及其降解相关特性的分析[J]. 微生物学报, 2009, 49(12): 1634-1642.

[20] L. Li, X. Liu, W. Yang, et al. Crystal structure of long-chain alkane monooxygenase (LadA) in complex with coenzyme FMN: Unveiling the long-chain alkane hydroxylase. Journal of Molecular Biology, 2008, 376(2): 453-465.

[21] T. Maier, H. H. Foerster, O. Asperger, et al. Molecular characterization of the 56-kDa CYP153 from Acinetobacter sp. EB104. Biochemical and Biophysical Research Communications, 2001, 286(3): 652-658.

[22] M. Throne-Holst, A. Wentzel, T. E. Ellingsen, et al. Identification of novel genes involved in long-chain n-alkane degradation by Acinetobacter sp. strain DSM 17874. Applied and Environmental Microbiology, 2007, 73(10): 3327-3332.

[23] M. Throne-Holst, S. Markussen, A. Winnberg, et al. Utilization of n-alkanes by a newly isolated strain of Acinetobacter venetianus: The role of two AlkB-type alkane hydroxylases. Applied Microbiology and Biotechnology, 2006, 72(2): 353-360.

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