机油高效降解菌的筛选鉴定及降解特性的初步研究

doi:10.13560/j.cnki.biotech.bull.1985.2017-1028

摘要/Abstract

摘要： 为丰富机油降解微生物菌种库，筛选适应性更强的机油高效降解菌株。以20#机油和真空泵油为唯一碳源，筛选出机油高效降解菌，经细菌形态学、生理生化及16S rRNA序列分析对机油高效降解菌株进行鉴定，采用紫外分光光度法和气相色谱质谱联用（GC-MS）研究菌株降解特性。从初筛的22株机油降解菌中筛选出4株机油高效降解菌株，分别为JZ6、JZ18、JZ41和JZ50，经鉴定4株菌株分别为Massilia sp、Pseudomona sp、Sphingobacterium sp和Shinella zoogloeoides sp，在含机油培养基中30℃培养7 d后，机油降解率分别为42.62%、33.67%、33.36%和40.52%。在温度20-40℃，pH5-9条件下菌株都具有降解机油的能力，4株菌株均能以十二烷、十六烷、十八烷和苯和萘为唯一碳源生长，JZ6和JZ50还能在含芘和菲的培养基中生长。GC-MS分析发现菌株JZ6、JZ18、JZ41和JZ50对总烷烃的降解率分别达到68.66%、52.69%、49.37%和61.40%，对直链烷烃的降解率分别为86.89%、55.98%、58.42%和89.13%，其中菌株JZ6和JZ50对除烷烃的其他芳香烃类物质也具有一定的降解作用。筛选出的4株机油降解菌具有较强机油降解能力，适应性强，可用于机油污染环境的生物修复。

关键词: 机油降解菌, 16S rRNA, 紫外分光光度法, GC-MS

Abstract: We aimed to isolate and identify highly efficient and highly-adapted engine oil-degrading bacteria for enriching the engine oil-degrading microbial strain library. Efficiently oil-degrading bacteria were obtained using the media individually supplemented with No.20 oil and vacuum pump oil as the sole carbon source，and were identified by morphology，physiological and biochemical properties，and 16S rRNA gene phylogenetics. Degradation characteristics of the strains were analyzed using UV spectrophotometry and gas chromatography mass spectrometry（GC-MS）. The most efficient oil-degrading strains，designed as JZ6，JZ18，JZ41，and JZ50，from 22 oil-degrading strains were classified as Massilia sp，Pseudomona sp，Sphingobacterium sp，and Shinella zoogloeoides sp，respectively. The degradation rate of oil reached 42.62%，33.67%，33.36% and 40.52%，when these strains were cultured in oil-containing media at 30℃ for 7 d. The strains were capable of degrading oil under the condition of pH5-9 and 20-40℃. The four strains grew on the media respectively added by dodecane，hexadecane，octadecane，benzene and naphthalene as the sole carbon source，and the strain JZ6 and JZ50 also grew well in the media containing pyrene and phenanthrene. GC-MS analysis showed that the degradation rate of strain JZ6，JZ18，JZ41 and JZ50 to total alkane was 68.66%，52.69%，49.37%，and 61.4%，and that to straight chain was 86.89%，55.98%，58.42%，and 89.13%，respectively. The strain JZ6 and JZ50 also degraded other aromatic hydrocarbon ta varied degree. The four strains of efficient oil-degrading bacteria were obtained，and had strong adaptability，thus these isolates can be used in the biological remediation of oil-polluted environment.

Key words: engine oil-degrading strain, 16SrRNA, UV spectrophotometry, GC-MS

黄曼曼 ,邓百万 ,王梦姣 ,陈文强 ,刘开辉 ,尹璐. 机油高效降解菌的筛选鉴定及降解特性的初步研究[J]. 生物技术通报, 2018, 34(6): 155-163.

HUANG Man-man ,DENG Bai-wan ,WANG Meng-jiao ,CHEN Wen-qiang ,LIU Kai-hui YIN Lu. Isolation and Identification of Bacteria Efficiently Involved in Engine Oil Degradation and Their Biodegradable Property[J]. Biotechnology Bulletin, 2018, 34(6): 155-163.

参考文献

[1] 冯晋阳. 石油烃优良降解菌的筛选分离及其降解性能的研究[D] . 西安：西安建筑科技大学, 2004.
[2] Maragkidou A, Arar S, Al-Hunaiti A, et al. Occupational health risk assessment and exposure to floor dust PAHs inside an educational building[J] . Sci Total Environ, 2016, （16）1492-1499.
[3] 李颖. 石油污染土壤的生物修复[D] . 北京：北京化工大学, 2008.
[4] 冯俊生, 张俏晨. 土壤原位修复技术研究与应用进展[J] . 生态环境学报2014, 23（11）：1861-1867.
[5] Montenegro IP, Mucha AP, Reis I, et al. Effect of petroleum hydrocarbons in copper phytoremediation by a salt marsh plant（Juncus maritimus）and the role of autochthonous bioaugmentation[J] . Environ Sci Pollut Res Int, 2016, 23（19）：19471-1980.
[6] Ma J, Yang Y, Dai X, et al. Effects of adding bulking agent, inorganic nutrient and microbial inocula on biopile treatment for oil-field drilling waste[J] . Chemosphere, 2016, 150：17-23.
[7] 唐金花, 于春光, 张寒冰. 石油污染土壤微生物修复的研究进展[J] . 湖北农业科学, 2011, 50（20）：4125-4128.
[8] 李宝明. 石油污染土壤微生物修复的研究[D] . 北京：中国农业科学院, 2007.
[9] 阮志勇. 石油降解菌株的筛选、鉴定及其石油降解特性的初步研究[D] . 北京：中国农业科学院, 2006.
[10] 崔丽虹. 石油烃降解菌的筛选、鉴定及复合菌群降解效果的研究[D] . 北京：中国农业科学院, 2009.
[11] 秦芳玲, 曹丽娟, 燕永利, 等. 几株机油降解菌及其处理含油废水的效果[J] . 油田化学, 2007, 24（3）：269-271.
[12] 辛蕴甜. 石油降解菌的降解性能、固定化及降解动力学研究[D] . 上海：东华大学, 2013.
[13] Chen M, Xu P, Zeng G, et al. Bioremediation of soils contaminated with polycyclic aromatic hydrocarbons, petroleum, pesticides, chlorophenols and heavy metals by composting：Applications, microbes and future research needs[J] . Biotechnol Adv, 2015, 33（6）：45-55.
[14] Ameen F, Hadi S, Moslem M, et al. Biodegradation of engine oil by fungi from mangrove habitat[J] . J Gen Appl Microbiol, 2015, 61（5）：85-92.
[15] 邓绍云, 徐学义, 邱清华. 我国石油污染土壤修复研究现状与展望[J] . 北方园艺, 2012（4）：184-190.
[16] 赵硕伟, 沈嘉澍, 沈标. 复合菌群的构建及其对石油污染土壤修复的研究[J] . 农业环境科学学报, 2011, 30（8）：1567-1572.
[17] 张秀霞, 徐娜娜, 秦丽姣. 固定化微生物降解石油的影响因素研究[J] . 安全与环境学报, 2011, 11（5）：77-81.
[18] Vigneron A, Alsop EB, Chambers B. Complementary microorganisms in highly corrosive biofilms from an offshore oil production facility[J] . Appl Environ Microbiol, 2016, 82（8）：45-54.
[19] Cui C, Li Z, Qian J. Complete genome of Martelella sp. AD-3, a moderately halophilic polycyclic aromatic hydrocarbons-degrading bacterium[J] . Biotechnol, 2016, 225：29-30.
[20] Lambo AJ, Patel TR. Isolation and characterization of a biphenyl-utilizing psychrotrophic bacterium, Hydrogenophaga taeniospiralis IA3-A, that cometabolize dichlorobiphenyls and polychlorinated biphenyl congeners in Aroclor 1221[J] . Journal of Basic Microbiology, 2006, 46（2）：94-107.
[21] 吴玉新. 紫外分光光度法测定污水中油含量的研究[J] . 环境保护, 1998.
[22] 展惠英. 紫外分光光度法测定废水中油的含量[J] . 甘肃联合大学学报, 2007, 21（1）：65-67.
[23] 韩寒冰, 刘杰凤. 机油降解菌的分离及其降解特性研究[J] . 安徽农业科学, 2009, 37（21）：9883-9884.
[24] 徐冯楠, 冯贵颖, 马雯等. 高效石油降解菌的筛选及其降解性能研究[J] . 生物技术通报, 2010（7）：221-223.
[25] 罗小艳. Massilia sp. WFl对菲的降解特性研究[D] . 杭州：浙江大学, 2015.
[26] 伍凤姬. 一株芘高效降解菌的筛选及其降解途径研究[D] . 上海, 华南理工大学, 2015. 
[27] 杨智, 陈吉祥, 秦波等. 3株石油降解红球菌（Rhodococcus spp.）特性及相关基因分析[J] . 应用与环境生物学报, 2015, 21（5）：805-812.
[28] Kuiper I, Lagendijke EL, Bloemberg GV, et al. Rhizoremedi-ation：abeneficialplant-microbe interaction[J] . Molecular Plant-Microbe Interactions, 2004, 17（1）：6-14.
[29] Cappello S, Santisi S, Calogero R, et al. Characterisation of oil-degrading bacteria isolatedWater[J] . Air, & Soil Pollution, 2012, 223（6）：3219-3226.
[30] Plotnikova EG, Altyntseva OV, Kosheleva IA, et al. Bacteria—degraders of polycyclic aromatic hydrocarbons, isolated from soil and bottom sediments in salt-mining areas[J] . Mikrobiologiia, 2000, 70（1）：61-69.
[31] Quatrini P, Scaglione G, De Pasquale C, et al. Isolation of Gram-positive n-alkane degraders from a hydrocarbon-contaminated Mediterranean shoreline[J] . J Appl Microbiol, 2008, 104（1）：251-259.
[32] Soudi MR, Kolahchi N. Bioremediation potential of a phenol degrading bacterium, Rhodococcus erythropolis SKO-1[J] . Prog Biol Sci, 2011, 1（1）：31-40.
[33] 孙庆华, 柏耀辉, 赵翠等. Shinella zoogloeoides BC026对吡啶的降解特性研究[J] . 环境科学, 2008, 29（10）：2939-2943.
[34] 苏莹, 陈莉, 汪辉, 刘兆普. 海洋石油降解菌的筛选与降解特性[J] . 应用与环境生物学报, 2008, 14（4）：518-522.
[35] Lin X, Yang B, Shen J, et al. Biodegradation of crude oil by an Arctic psychrotrophic bacterium Seudoalteromomas sp. P29[J] . Curr Microbiol, 2009, 5（3）：341-345.