抗耐药细菌药用植物内生菌的筛选与鉴定

doi:10.13560/j.cnki.biotech.bull.1985.2015.04.022

摘要/Abstract

摘要： 耐甲氧西林金黄色葡萄球菌(MRSA)、耐头孢霉素大肠杆菌和耐亚胺培南铜绿假单胞菌在医院临床治疗中引起的严重感染，极大地危害着人类身体健康。旨在从植物组织中分离和筛选高活性的拮抗上述耐药细菌的内生菌株。从分离自22种药用植物组织的197株内生菌中，经过两步筛选获得18株对MRSA有拮抗作用的菌株，未筛选到对耐头孢霉素大肠杆菌和耐亚胺培南铜绿假单胞菌有拮抗作用的菌株；经16S rRNA序列分析，其中8株为链霉菌属，6株为芽孢杆菌属，4株为假单胞菌属；对抑菌效果较强的5株菌进行发酵培养，检测发酵液对MRSA的抑菌效果，其中QN1和CF2的发酵液对MRSA有很好的抑制效果。

关键词: 植物内生菌, 抑菌活性, 耐药细菌, 耐甲氧西林金黄色葡萄球菌

Abstract: Methicillin-resistant Staphylococcus aureus(MRSA), Cephalosporin-resistant Escherichia coli and Imipenem-resistant Pseudomonas aeruginosa have caused severe infection in clinical therapy and greatly endanger human health. The aim of this study is to isolate and screen highly active antagonistic endophytes against drug-resistant bacteria from medicinal plant. After two-step screening, 18 strains have antagonistic effect on MRSA were obtained from 197 strains isolated from 22 medicinal plants, none of the strains has antagonistic effect on Cephalosporin-resistant Escherichia coli and Imipenem-resistant Pseudomonas aeruginosa；16S rRNA sequence analysis showed that eight strains belong to Streptomyces, six strains belong to Bacillus, and four strains belong to Pseudomonas. Five strains which have strong anti bacterial activity were chosen for fermentation, and the fermented liquid of QN1 have strong inhibition effect on MRSA.

Key words: endophytes, antibacterial activity, drug-resistant bacteria, Methicillin-resistant Staphylococcus aureus

刘晓瑜, 马玉超^{. 抗耐药细菌药用植物内生菌的筛选与鉴定[J]. 生物技术通报, 2015, 31(3): 154-160.}

Liu Xiaoyu, Ma Yuchao. Screening and Identification of Antagonistic Endophytes Against Drug-resistant Bacteria from Medicinal Plants[J]. Biotechnology Bulletin, 2015, 31(3): 154-160.

参考文献

[1] Leekha S, Terrell CL, Edson RS. General principles of antimicrobial therapy[C]. Mayo Clinic Proceedings. Elsevier, 2011, 86(2)：156-167.
[2] Walsh C. Antibiotics：actions, origins, resistance[M]. American Society for Microbiology(ASM), 2003.
[3] Stewart PS, William Costerton J. Antibiotic resistance of bacteria in biofilms[J]. The Lancet, 2001, 358(9276)：135-138.
[4] Spellberg B, Bartlett JG, Gilbert DN. The future of antibiotics and resistance[J]. New Engl J Med, 2013, 368(4)：299-302.
[5] Berkelman RL, Bryan RT, Osterholm MT, et al. Infectious disease surveillance：a crumbling foundation[J]. Science-AAAS-Weekly Paper Edition-including Guide to Scientific Information, 1994, 264(5157)：368-370.
[6] 张小江, 陈雨, 朱任媛, 等. 北京协和医院内科住院患者细菌耐药性监测[J]. 协和医学杂志, 2013, 4(4)：417-424.
[7] 朱景倩, 赵建平, 涂军伟, 等. 呼吸科病房病原菌分布及耐药性研究[J]. 中华医院感染学杂志, 2013, 23(8)：1943-1945.
[8] 申桂娟, 陆军, 祝进. 耐甲氧西林金黄色葡萄球菌的临床分布与耐药性分析[J]. 中华医院感染学杂志, 2014, 24(2)：282-284.
[9] 于旭红, 王冬, 王睿. 耐甲氧西林金黄色葡萄球菌的研究进展[J]. 中国临床药理学杂志, 2011, 27(4)：306-310.
[10] Qin S, Xing K, Jiang JH, et al. Biodiversity, bioactive natural products and biotechnological potential of plant-associated endophytic actinobacteria[J]. Appl Microbiol Biotechnol, 2011, 89(3)：457-473.
[11] 魏宝阳, 曹亮, 李顺祥, 等. 内生菌与药用植物的关系及对次生代谢产物的影响[J]. 中国农学通报, 2011, 27(19)：83-88.
[12] Crawford DL, Lynch JM, Whipps JM, et al. Isolation and characterization of actinomycete antagonists of a fungal root pathogen[J]. Appl Environ Microbiol, 1993, 59(11)：3899-3905.
[13] Shirling EB, Gottlieb D. Methods for characterization of Streptomyces species[J]. Int J Syst Bacteriol, 1996, 16(3)：313-340.
[14] Qin S, Li J, Chen HH, et al. Isolation, diversity and antimicrobial activity of rare actinobacteria from medicinal plants of tropical rain forests in Xishuangbanna, China[J]. Appl Environ Microbiol, 2009, 75(19)：6176-6186.
[15] Todorova S, Kozhuharova L. Characteristics and antimicrobial activity of Bacillus subtilis strains isolated from soil[J]. World J Microbiol Biotechnol, 2010, 26(7)：1207-1216.
[16] 窦桂铭, 刘晓瑜, 张银东, 等. 大豆菌核病拮抗内生细菌的筛选与鉴定[J]. 广东农业科学, 2013, 40(7)：78-82.
[17] Ma Y, Yu H, Pan W, et al. Identification of nitrile hydratase-producing Rhodococcus ruber TH and characterization of an amiE negative mutant[J]. Bioresour Technol, 2010, 101(1)：285-291.
[18] Kim OS, Cho YJ, Lee K, et al. Introducing EzTaxon-e：a prokaryotic 16S rRNA gene sequence database with phylotypes that represent uncultured species[J]. Int J Syst Evol Microbiol, 2012, 62(Pt 3)：716-721.
[19] Tamura K, Peterson D, Peterson N, et al. MEGA5：molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods[J]. Mol Biol Evol, 2011, 28(10)：2731-2739.
[20] Saitou N, Nei M. The neighbor-joining method：a new method for reconstructing phylogenetic trees[J]. Mol Biol Evol, 1987, 4(4)：406-425.
[21] Monaghan RL, Barrett JF. Antibacterial drug discovery—then, now and the genomics future[J]. Biochem Pharmacol, 2006, 71(7)：901-909.
[22] Hiramatsu K, Aritaka N, Hanaki H, et al. Dissemination in Japanese hospitals of strains of Staphylococcus aureus heterogeneously resistant to vancomycin[J]. The Lancet, 1997, 350(9092)：1670-1673.
[23] Pasberg-Gauhl C. A need for new generation antibiotics against MRSA resistant bacteria[J]. Drug Discovery Today：Technolo-gies, 2014, 11：109-116.
[24] Zhang J, Chen YP, Miller KP, et al. Antimicrobial metallopolymers and their bioconjugates with conventional antibiotics against multidrug-resistant bacteria[J]. J Am Chem Soc, 2014, 136(13)：4873-4876.
[25] 张守村, 韩松, 黄晓艳, 等. 一株抗 MRSA 内生细菌的鉴定及其活性物质[J]. 微生物学通报, 2011, 38(6)：860-864.
[26] Liu G, Chater KF, Chandra G, et al. Molecular regulation of antibiotic biosynthesis in Streptomyces[J]. Microbiol Mol Biol R, 2013, 77(1)：112-143.
[27] Van Wezel GP, McDowall KJ. The regulation of the secondary met-abolism of Streptomyces：new links and experimental advances[J]. Nat Prod Rep, 2011, 28(7)：1311-1333.