联苯代谢对微生物的生长胁迫及分裂抑制
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摘要
以联苯/多氯联苯降解菌株R04(Rhodococcus sp. R04)和几种模式微生物为研究对象,利用高效液相色谱、荧光显微镜、扫描电子显微镜等分析微生物在联苯及其代谢物培养条件下细胞分裂和形态的变化.结果表明联苯及其代谢产物对红球菌R04和几种模式微生物细胞的分裂有抑制作用,并对部分微生物形态有影响.与前体-联苯及其代谢产物2-羟基-6-酮基-6-苯基-2,4-己二烯酸相比, 2,3-二羟基联苯对G~+、G~-细菌,或是酵母细胞分裂都有较强的抑制和形态的改变. 2,3-二羟基联苯导致R. R04和缺陷型R. R04细胞形成不完整隔膜的比例增加;造成96.4%的大肠杆菌BL21细胞表面凹陷,胞质内容物流失,菌体体积缩小;导致枯草芽孢杆菌89.6%的细胞体积明显缩小;导致金黄色葡萄球菌基本没有细胞能形成完整的分裂隔膜;导致红酵母细胞能进行出芽生殖的比例从64.2%降低到19.3%,但对其细胞形态无明显改变.联苯代谢物2,3-二羟基联苯对红球菌R04及其它微生物细胞分裂和增殖的抑制作用比其前体-联苯强,建议在研究环境化合物与微生物互作时,应考虑环境化合物代谢的毒性效应.
The biphenyl/polychlorinated biphenyl degrading strain R04(Rhodococcus sp. R04) and several model microorganisms were used as the research objects. Cell division and morphological changes of biphenyl/polychlorinated biphenyl degrading strain R04 were analyzed by high performance liquid chromatography, fluorescence microscopy and scanning electron microscopy under the conditions of biphenyl and its metabolites culture. The results showed that the division of Rhodococcus sp. R04 and several model microbial cells were inhibited under biphenyl and its metabolites culture conditions, and some microbial cells morphology were affected. Compared with the precursor-biphenyl and its metabolite 2-hydroxy-6-keto-6-phenyl-2, 4-hexadienoic acid, 2,3-dihydroxybenzene has stronger inhibition and morphological change on G~+, G~- bacteria or Rhodotorula cell division. The proportion of R.R04 cells and defective R.R04 cells forming incomplete septum was increased under the condition of 2,3-dihydroxybenzene culture. It caused that 96.4% of E. coli BL21 cells surface was dented, cytoplasmic content was lost and bacterial body volume was shrunk. It caused that 89.6% of Bacillus subtilis cells was shrunk significantly. The phenomenon that Staphylococcus aureus has almost no cells to form a complete septum was caused. The percentage of Rhodotorula cells that could germinate and reproduce was decreased from 64.2% to 19.3%, but there was no significant change in cell morphology. Biphenyl metabolite 2,3-dihydroxybiphenyl has strongly inhibitory effect on cell division and proliferation of Rhodococcus sp. R04 and other microorganisms than its precursor-biphenyl, and it is suggested that the toxic effect of environmental compounds metabolism should be considered when the interaction between environmental compounds and microorganisms.
The biphenyl/polychlorinated biphenyl degrading strain R04(Rhodococcus sp. R04) and several model microorganisms were used as the research objects. Cell division and morphological changes of biphenyl/polychlorinated biphenyl degrading strain R04 were analyzed by high performance liquid chromatography, fluorescence microscopy and scanning electron microscopy under the conditions of biphenyl and its metabolites culture. The results showed that the division of Rhodococcus sp. R04 and several model microbial cells were inhibited under biphenyl and its metabolites culture conditions, and some microbial cells morphology were affected. Compared with the precursor-biphenyl and its metabolite 2-hydroxy-6-keto-6-phenyl-2, 4-hexadienoic acid, 2,3-dihydroxybenzene has stronger inhibition and morphological change on G~+, G~- bacteria or Rhodotorula cell division. The proportion of R.R04 cells and defective R.R04 cells forming incomplete septum was increased under the condition of 2,3-dihydroxybenzene culture. It caused that 96.4% of E. coli BL21 cells surface was dented, cytoplasmic content was lost and bacterial body volume was shrunk. It caused that 89.6% of Bacillus subtilis cells was shrunk significantly. The phenomenon that Staphylococcus aureus has almost no cells to form a complete septum was caused. The percentage of Rhodotorula cells that could germinate and reproduce was decreased from 64.2% to 19.3%, but there was no significant change in cell morphology. Biphenyl metabolite 2,3-dihydroxybiphenyl has strongly inhibitory effect on cell division and proliferation of Rhodococcus sp. R04 and other microorganisms than its precursor-biphenyl, and it is suggested that the toxic effect of environmental compounds metabolism should be considered when the interaction between environmental compounds and microorganisms.
引文
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[14]李叶,张爽,李玉灵,等.多氯联苯暴露对绦柳初生根及显微结构的影响[J].北方园艺, 2016,(24):55-60.Li Y, Zhang S, Li Y L, et al. Effects of Polychlorinated Biphenyls(PCBs)on the Initial Roots and Microstructure of Salix matsudana f.pendula Schneid[J]. Northern Horticulture, 2016,(24):55-60.
[15]杨秀清,郑媛,李鹏丽,等.红球菌-R04生物降解多卤代联苯的影响因素研究[J].中国环境科学, 2010,30(5):694-698.Yang X Q, Zheng Y, Li PL, et al. Influencing factor for the biodegradation of polyhalogenated biphenyls by Rhodococcus sp. R04[J]. China Environmental Science, 2010,30(5):694-698.
[16]杨秀清,商慧慧.红球菌R04细胞的不对称分裂及其在联苯胁迫下的分裂抑制[J].微生物学报, 2018,58(5):893-906.Yang X Q, Shang H H. Asymmetric cell division of Rhodococcus sp.R04 and its division inhibition under biphenyl stress[J]. Acta Microbiologica Sinica, 2018,58(5):893-906.
[17]曹星星,杨秀清.联苯/多氯联苯对红球菌R04细胞形态及隔膜的影响[J].山西大学学报(自然科学版), 2016,39(2):287-294.Cao X X, Yang X Q. The effect of biphenyl/polychlorinated biphenyl on cell morphology and septum of Rhodococcus sp. R04[J]. Journal of Shanxi University(Natural Science Edition), 2016,39(2):287-294.
[18] Hayteas D L, Duffield D A. The determination by HPLC of PCB and p,p′-DDE residues in marine mammals stranded on the Oregon Coast[J]. Marine Pollution Bulletin, 1997,34(10):844-848.
[19] Yamada T, Shimomura Y, Hiraoka Y, et al. Oxidative stress by biphenyl metabolites induces inhibition of bacterial cell separation[J].Applied Microbiology&Biotechnology, 2006,73(2):452-457.
[20] Mizukami-Murata S, Fujita K, Nakano T. Effect of lower chlorinated hydroxylated-polychlorobiphenyls on development of PC12cells[J].Environmental Science&Pollution Research, 2017,25(17):1-12.
[21] Subramanian S, Schnoor J L, Aken B V. Effects of Polychlorinated Biphenyls(PCBs)and Their Hydroxylated Metabolites(OH-PCBs)on Arabidopsis thaliana[J]. Environmental Science&Technology, 2017,51(12):7263-7270.
[22] Wang L, Li F. Study on the Quantitative Structure-activity Relationship of Acute Toxicities of Substituted Aromatic Compounds to Daphnia magna[J]. Journal of Anhui Agricultural Sciences, 2010,38(23):12542-12544.
[23]陈景文,廖宜勇,赵元慧,等.取代氮杂环类化合物对大型蚤(Daphnia magna Straus)的定量结构-活性关系研究[J].农药, 1996,35(5):21-24.Chen J W, Liao Y Y, Zhao Y H, et al. Study on quantitative structureactivity relationship of substituted nitrogen heterocyclic compounds to Daphnia magna Straus[J]. Pesticide, 1996,35(5):21-24.
[24] Struck S, Schmidt U, Gruening B, et al. Toxicity versus potency:elucidation of toxicity properties discriminating between toxins, drugs,and natural compounds[J]. Genome informatics, 2008,20(20):231-242.
[25] Albert J O. Nitrogenremoval in constructed wetlands treating nitrified meat processing effluent[J]. Water Science and Technology, 1995,32(3):137-147.
[26]徐文玉.细菌细胞壁的教学方案[J].微生物学通报, 1985,12(5):44-48.Xu W Y. Teaching program of bacterial cell wall[J]. Acta Mcrobiologica Sinica, 1985,12(5):44-48.
[27]滕菲,郭桂萍,赵勇,等.革兰氏阳性菌和阴性菌对山梨酸钾的耐受差异性[J].食品与生物技术学报, 2012,31(4):417-422.Teng F, Guo G P, Zhao Y, et al. Study of the Tolerance Difference between Gram Positive and Gram Negative Bacteria to Potassium Sorbate[J]. Food Science and Biotechnology, 2012,31(4):417-422.
[28] Rundegren J, T Sj?din, Petersson L, et al. Delmopinol interactions with cell walls of gram-negative and gram-positive oral bacteria[J].Oral Microbiology&Immunology, 1995,10(2):102-109.
[2] Safe S. Polychlorinated biphenyls(PCBs), dibenzo-p-dioxins(PCDDs), dibenzofurans(PCDFs), and related compounds:environmental and mechanistic considerations which support the development of toxic equivalency factors(TEFs)[J]. Crc Critical Reviews in Toxicology, 1990,21(1):51-88.
[3] Akahane M, Matsumoto S, Kanagawa Y, et al. Long-Term Health Effects of PCBs and Related Compounds:A Comparative Analysis of Patients Suffering from Yusho and the General Population[J]. Archives of Environmental Contamination and Toxicology, 2018,74(2):203-217.
[4] Weber R, Herold C, Hollert H, et al. Life cycle of PCBs and contamination of the environment and of food products from animal origin[J].Environmental Science and Pollution Research, 2018,25(1):16325-16343.
[5]徐莉,滕应,张雪莲,等.多氯联苯污染土壤的植物-微生物联合田间原位修复[J].中国环境科学, 2008,28(7):646-650.Xu L, Teng Y, Zhang X L, et al. Combined remediation of PCBs polluted soil by plant and microorganism in a field trial[J]. China Environmental Science, 2008,28(7):646-650.
[6] Yang Xiuqing, Sun Yan, Qian Shijun. Biodegradation of sevenpolychlorinated biphenyls by a newly isolated aerobic bacterium(Rhodococcus sp. R04)[J]. J. Ind. Microbiol. Biotechnol.,2004,31(9):415-420.
[7] Hayase N, Taira K, Furukawa K. Pseudomonas putida KF715bphABCD operon encoding biphenyl and polychlorinated biphenyl degradation:cloning, analysis, and expression in soil bacteria[J]. Journal of Bacteriology, 1990,172(2):1160-1164.
[8]孙艳,钞亚鹏,钱世钧.嗜吡啶红球菌R04的联苯降解途径的研究[J].微生物学报, 2003,43(5):653-658.Sun Yan, Chao Y P, Qian S J. Study on the Degradation Pathway of Biphenyl by Rhodococcus pyridinovorans R04[J]. Acta Microbiologica Sinica, 2003,43(5):653-658.
[9] Bhowmik S, Horsman G P, Bolin J T, et al. The molecular basis for inhibition of BphD, a C-C bond hydrolase involved in polychlorinated biphenyls degradation:large 3-substituents prevent tautomerization[J].Journal of Biological Chemistry, 2007,282(50):36377-36385.
[10] Parnell J J, Park J, Denef V, et al. Coping with polychlorinated biphenyl(PCB)toxicity:physiological and genome-wide responses of Burkholderiaxenovorans LB400 to PCB-mediated stress[J]. Applied and Environmental Microbiology, 2006,72(10):6607-6614.
[11] Bourquin A W, Cassidy S. Effect of polychlorinated biphenyl formulations on the growth of estuarine bacteria[J]. Applied Microbiology, 1975,29(1):125-127.
[12] Joseph L J, Sandra W J. Intellectual Impairment in Children Exposed to Polychlorinated Biphenyls in Utero[J]. New England Journal of Medicine, 1996,335(11):783-789.
[13] Berg M V D, Craane B L H J, Sinnige T, et al. Biochemical and toxic effects of polychlorinated biphenyls(PCBs), dibenzo-P-dioxins(PCDDs)and dibenzofurans(PCDFs)in the cormorant(phalacrocorax carbo)after in ovo exposure[J]. Environmental Toxicology&Chemistry, 2010,13(5):803-816.
[14]李叶,张爽,李玉灵,等.多氯联苯暴露对绦柳初生根及显微结构的影响[J].北方园艺, 2016,(24):55-60.Li Y, Zhang S, Li Y L, et al. Effects of Polychlorinated Biphenyls(PCBs)on the Initial Roots and Microstructure of Salix matsudana f.pendula Schneid[J]. Northern Horticulture, 2016,(24):55-60.
[15]杨秀清,郑媛,李鹏丽,等.红球菌-R04生物降解多卤代联苯的影响因素研究[J].中国环境科学, 2010,30(5):694-698.Yang X Q, Zheng Y, Li PL, et al. Influencing factor for the biodegradation of polyhalogenated biphenyls by Rhodococcus sp. R04[J]. China Environmental Science, 2010,30(5):694-698.
[16]杨秀清,商慧慧.红球菌R04细胞的不对称分裂及其在联苯胁迫下的分裂抑制[J].微生物学报, 2018,58(5):893-906.Yang X Q, Shang H H. Asymmetric cell division of Rhodococcus sp.R04 and its division inhibition under biphenyl stress[J]. Acta Microbiologica Sinica, 2018,58(5):893-906.
[17]曹星星,杨秀清.联苯/多氯联苯对红球菌R04细胞形态及隔膜的影响[J].山西大学学报(自然科学版), 2016,39(2):287-294.Cao X X, Yang X Q. The effect of biphenyl/polychlorinated biphenyl on cell morphology and septum of Rhodococcus sp. R04[J]. Journal of Shanxi University(Natural Science Edition), 2016,39(2):287-294.
[18] Hayteas D L, Duffield D A. The determination by HPLC of PCB and p,p′-DDE residues in marine mammals stranded on the Oregon Coast[J]. Marine Pollution Bulletin, 1997,34(10):844-848.
[19] Yamada T, Shimomura Y, Hiraoka Y, et al. Oxidative stress by biphenyl metabolites induces inhibition of bacterial cell separation[J].Applied Microbiology&Biotechnology, 2006,73(2):452-457.
[20] Mizukami-Murata S, Fujita K, Nakano T. Effect of lower chlorinated hydroxylated-polychlorobiphenyls on development of PC12cells[J].Environmental Science&Pollution Research, 2017,25(17):1-12.
[21] Subramanian S, Schnoor J L, Aken B V. Effects of Polychlorinated Biphenyls(PCBs)and Their Hydroxylated Metabolites(OH-PCBs)on Arabidopsis thaliana[J]. Environmental Science&Technology, 2017,51(12):7263-7270.
[22] Wang L, Li F. Study on the Quantitative Structure-activity Relationship of Acute Toxicities of Substituted Aromatic Compounds to Daphnia magna[J]. Journal of Anhui Agricultural Sciences, 2010,38(23):12542-12544.
[23]陈景文,廖宜勇,赵元慧,等.取代氮杂环类化合物对大型蚤(Daphnia magna Straus)的定量结构-活性关系研究[J].农药, 1996,35(5):21-24.Chen J W, Liao Y Y, Zhao Y H, et al. Study on quantitative structureactivity relationship of substituted nitrogen heterocyclic compounds to Daphnia magna Straus[J]. Pesticide, 1996,35(5):21-24.
[24] Struck S, Schmidt U, Gruening B, et al. Toxicity versus potency:elucidation of toxicity properties discriminating between toxins, drugs,and natural compounds[J]. Genome informatics, 2008,20(20):231-242.
[25] Albert J O. Nitrogenremoval in constructed wetlands treating nitrified meat processing effluent[J]. Water Science and Technology, 1995,32(3):137-147.
[26]徐文玉.细菌细胞壁的教学方案[J].微生物学通报, 1985,12(5):44-48.Xu W Y. Teaching program of bacterial cell wall[J]. Acta Mcrobiologica Sinica, 1985,12(5):44-48.
[27]滕菲,郭桂萍,赵勇,等.革兰氏阳性菌和阴性菌对山梨酸钾的耐受差异性[J].食品与生物技术学报, 2012,31(4):417-422.Teng F, Guo G P, Zhao Y, et al. Study of the Tolerance Difference between Gram Positive and Gram Negative Bacteria to Potassium Sorbate[J]. Food Science and Biotechnology, 2012,31(4):417-422.
[28] Rundegren J, T Sj?din, Petersson L, et al. Delmopinol interactions with cell walls of gram-negative and gram-positive oral bacteria[J].Oral Microbiology&Immunology, 1995,10(2):102-109.