Microsoft Word - 20周丽英.doc

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1 Chinese Journal of Eco-Agriculture, Aug. 202, 20(8): DOI: /SP.J * 周丽英林素兰李叶仁杰艺林文雄林淑婷刘,2 林瑞余,2** 杰肖清铁 ( ; ) 为探明前期分离得到的 3 株水稻根际耐镉细菌碳源代谢特性, 在进行生理生化鉴定的基础上, 采用 BIOLOG ECO 微平板法, 分析了 3 株细菌 ( 菌株 A 菌株 B 和菌株 C) 的碳源代谢功能生理生化试验结果表明, 3 株菌均属于革兰氏阴性细菌, 且均具有极生鞭毛, 菌株 C 的生理生化鉴定为铜绿假单胞菌, 菌株 A 与菌株 B 为假单胞菌属的其他种, 结果与之前的分子生物学鉴定结果一致 BIOLOG 分析结果表明, 3 株菌株在培养 72 h 的 AWCD( 平均颜色变化率 ) 均接近最大值, 在培养 24 h 时的 AWCD 存在显著差异, 这种差异主要体现在对氨基酸的利用上在对提供的 3 种单一碳源利用上, 3 株菌对 N- 乙酰 -D- 葡萄糖胺 L- 精氨酸 L- 天冬酰胺甲叉丁二酸腐胺 4- 羟基苯甲酸丙酮酸甲酯吐温 -40 和吐温 -80 的利用率均较高, 且对 L- 精氨酸甲叉丁二酸丙酮酸甲酯的利用率存在显著差异本研究结果对深入了解耐镉菌株增殖所需要的碳源以及将来优化耐镉细菌最适培养基提供了理论依据, 也为构建相应的基因工程菌提供了微生物资源镉耐镉菌株生理生化特性 BIOLOG ECO 微平板法碳源利用功能分析 AWCD : Q93-33 : A : (202) Analysis of carbon source metabolism functions of cadmium-tolerant bacteria separated from rice (Oryza satiua) rhizosphere ZHOU Li-Ying, LIN Su-Lan, LI Yi, LIN Shu-Ting, LIU Jie, XIAO Qing-Tie, YE Ren-Jie, LIN Wen-Xiong,2, LIN Rui-Yu,2 (. School of Life Sciences, Fujian Agriculture and Forestry University, Fuzhou , China; 2. Institute of Agroecology, Fujian Agriculture and Forestry University, Fuzhou , China) Abstract To understand metabolic abilities of carbon sources in three cadmium-tolerant bacteria (strain A, strain B and strain C) isolated from rice rhizosphere soils, basic characteristics of bacteria strains were determined by physiological and biochemical reactions and metabolic functions of carbon sources analyzed via the BIOLOG ECO plate method. The main principle was based on the fact that color-reduction reaction showed differences in carbon source utilization. The physiological and biochemical characteristics showed that all three bacteria strains behaved as Gram-negative with born flagellum. Strain C was identified as Pseudomonas aeruginosa, while strains A and B both belonged to different Pseudomonas species. The conclusions on physiological and biochemical characteristics were the same as in previous molecular identifications. BIOLOG ECO analysis showed that average well color development (AWCD) nearly approached the maximum after culturing the strains for 72 hours. Significant differences were noted in AWCD among cultured strains for 24 hours, dominated by amino acid utilization efficiency. Among the thirty-one single carbon sources used, higher utilization rates were noted in nine compounds. The compounds included N-Acetyl-Dglucosamine, L-Arginine, L-Asparagine, Itaconic acid, Putrescicine, 4-Hydroxy benzoic acid, Pyruvic acid methyl ester, Tween 40 and Tween 80. Also significant differences were detected among the three bacteria strains regarding utilization rates of L-Arginine, Itaconic acid and Pyruvic acid methyl ester. On the basis of carbon source utilization, more information was gathered on strain functions. Thus this study expanded our knowledge on needed carbon sources for cadmium-tolerant strain growth. The study also taught us optimization strategy of cultured media which was beneficial for future research work. Furthermore, it provided precious * ( , ) (2009J0056) (JA09084) ** : (968 ),,,, lrylin2004@63.com (986 ),,, feixuezly@yeah.net : :

2 microorganism resources for constructing corresponding bacteria gene engineering. Key words Cadmium, Cadmium-resistant bacteria, Physio-biochemical characteristics, BIOLOG ECO plate method, Carbon source utilization, Functional analysis, AWCD (Received Jan. 4, 202; accepted Apr. 0, 202) 20 50,,,, [],, (Pseudomonas aeruginosa) 000 mg L [2], (P. fluorescens) (P. putida) (Aeromonas hydrophila) [3 4], (Pseudomonas sp.),,, [4] 3, 700 mg L, [5],,, BIOLOG ECO, 材料与方法. 3 ( A B C) [5] 20% 80, 00 μl 300 mg L, 37 8~24 h,, 24 h, BIOLOG ECO, BIOLOG ECO Biolog.2.2. [6], M. R V-P.2.2 BIOLOG ECO [7], 0 ml, l ml 99 ml,, 20 r min 2 h, 5 ml, 45 ml, 3, 000, ELSIA BIOLOG ECO 25, 50 μl,, 28, 0 h 24 h 48 h 72 h 96 h 20 h 44 h, ELSIA 590 nm, 3 [8 9].2.3 ELSIA AWCD(average well color development), AWCD=[ (C R)]/n, C 3, R, n 24 h AWCD 2 结果与分析 2., A B C,,, M. R 3 B, A, V-P C [5], C, B Pseudomonas sp., B C, A, C, C, B C V-P, A B C, A B 2.2 (AWCD), 3, 72 h,

3 8 : 085 表菌株的生理生化试验结果 Table Physiological and biochemical properties of the three strains Test item A Strain A B Strain B C Strain C P. aeruginosa Gram staining Flagellum staining Mobility test Fluorescent test Pyovendin test Oxidase test Catalase test Gelatin liquefaction Amylolysis Nitrate reduction test Nitrite reduction test M. R M. R test V-P V-P test + + +, + stands for positive test result; stands for negative test result. 图不同耐镉菌株的 AWCD 随时间的变化 Fig. Dynamic changes in averages of well color development (AWCD) for carbon sources of different cadmium-resistant bacterium AWCD, 20 h AWCD, C A B AWCD 24~72 h, 24 h, 3 AWCD 2.3 (AWCD) 2,, A B 24 h C, 3, 24 h, A B, C; 96 h, A B C, A, 72 h B C, A B, 0 h 72 h B C, B C, A 24 h B C, 3, A B C 2.4 (AWCD) 3 24 h 3 ( 3), 3 9, : N- -D- L- L A B N- - D- L-, C; 3 L-, A> B> C; A 4- B C ; A B C -40 ; -80 B A C, A C 3 讨论与结论, C, 3 A C, B ; A, B C ; V-P, C,, A B, A B C [5]

4 Fig. 2 图 2 不同菌株的各类碳源的 AWCD 值随时间变化 Dynamic changes in averages of well color development (AWCD) for different carbon sources in different cadmium-resistant bacterium 图 3 培养 24 h 时不同耐镉菌株 3 种碳源的 AWCD Fig. 3 Averages of well color development (AWCD) for 3 kinds of carbon sources in different cadmium-resistant bacterium after culture for 24 h : D- D-Glucosaminic acid; 2: D- D-Galacturonic acid; 3: D- D-xylose; 4: D- D-Glucuronic acid; 5: D- D-Cellobiose; 6: N- -D- N-Acetyl-D-glucosamine; 7: α-d- α-d-lactose; 8: α- α-cyclodextrin; 9: β- -D- β-methyl-d-glucose; 0: Glycogen; : -- Glucose--Phosphate; 2: L- L-Arginine; 3: L- L-serine; 4: L- L-threonine; 5: L- L-Asparagine; 6: -L- Glycyl-L-Glutamic acid; 7: D- D-Malic acid; 8: α- α-ketobutyric acid; 9: γ- γ-hydroxybutyric acid; 20: Itaconic acid; 2: L- L-Phenylalanin; 22: Phenylethylamine; 23: Putrescicine; 24: 2-2-Hydroxy benzoic acid; 25: 4-4-Hydroxy benzoic acid; 26: D- D-Mannitol; 27: α- D,L-α-Glycetol phosphate; 28: Pyruvic acid methyl ester; 29: -Erythritol; 30: -40 Tween-40; 3: -80 Tween-80.

5 8 : 087 BIOLOG ECO, 3, C, 3 72 h AWCD, 3 24 h AWCD, ; 3, 3 N- -D- L- L , ;, 3 L-, BIOLOG ECO [0 2], [3 5],,,,, BIOLOG ECO,, 参考文献 [],. [J]., 2009, 37(7): [2],,,. [J]., 200, 35(0): [3]. [D]. :, 2007: 3 7 [4],,,. [J]., 20, 42(): 5 55 [5],,,. [J]., 202, 20(5): [6],. [M]. :, 200: [7],,,. [J]., 200, 30(4): [8] Lin R Y, Rong H, Zhou J J, et al. Impact of allelopathic rice seedlings on rhizospheric microbial populations and their functional diversities[j]. Acta Ecologica Sinica, 2007, 27(9): [9] Kong W D, Zhu Y G, Fu B J, et al. The veterinary antibiotic oxytetracycline and Cu influence functional diversity of the soil microbial community[j]. Environ Pollut, 2006, 43(): [0] Garland J L. Analysis and interpretation of community-level physiological profiles in microbial ecology[j]. FEMS Microbiol Ecol, 997, 24(4): [],,,. [J]., 2004, 5(): [2] Graham M H, Haynes R J. Catabolic diversity of soil microbial communities under sugarcane and other land uses estimated by Biolog and substrate-induced respiration methods[j]. Applied Soil Ecology, 2005, 29(2): [3] Zheng H, Ouyang Z Y, Wang X K, et al. Effects of regenerating forest cover on soil microbial communities: A case study in hilly red soil region, Southern China[J]. Forest Ecology and Management, 2005, 27(2/3): [4],,. [J]., 2007, 40(8): [5] He X Y, Wang K L, Xu L L, et al. Soil microbial metabolic diversity and its seasonal variations along a vegetation succession in a karst area: A case study in southwest China[J]. Acta Scientiae Circumstantiae, 2008, 28(2):

Microsoft Word - 793-797 tb20150504赵宏宇s-高校教改纵横.doc

Microsoft Word - 793-797 tb20150504赵宏宇s-高校教改纵横.doc 微生物学通报 Microbiology China tongbao@im.ac.cn Apr. 20, 2016, 43(4): 793 797 http://journals.im.ac.cn/wswxtbcn DOI: 10.13344/j.microbiol.china.150504 高校教改纵横生物工程专业发酵课程群建设探索 * 赵宏宇