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题名: 抗生素对废水处理微生物的影响 及基于酵母菌的调控
作者: 王春艳
学位类别: 博士后
答辩日期: 2016-10
授予单位: 中国科学院研究生院
授予地点: 北京
导师: 杨敏
关键词: 抗生素 ; 硝化作用 ; 氨氧化菌 ; 酵母菌预处理
其他题名: Study on the effect of antibiotics on microorganisms in wastewater treatment and the control technology of a yeast-based process
学位专业: 环境工程
中文摘要: 生物脱氮是废水处理的重要环节,硝化微生物缓慢的生长速率以及对环境的高度敏感性导致硝化作用成为污水氮素去除的限速步骤。前期对不同抗生素生产废水系统进行氨氧化微生物调查表明,废水中高浓度的螺旋霉素能够促进氨氧化古菌相对丰度的提高。因此,长期暴露条件下,不同作用机制的抗生素对硝化微生物影响的系统性研究,具有重要意义。
本论文首先在实验室采用摇瓶实验的方式,通过280天的运行,考察不同作用机制的抗生素(链霉素、土霉素、螺旋霉素、氨苄青霉素)在不同浓度梯度下,对硝化系统的处理效果、微生态以及抗性基因的影响。同时对交替生产氨基糖苷类(核糖/巴龙)、大环内脂类(螺旋霉素)抗生素的某药厂废水处理系统进行长周期氨氧化微生物进行定量追踪及抗性基因的种类和丰度调查。最后针对该厂实际废水特点,构建复合酵母菌群预处理平台,优化运行参数,评价处理效果。本论文取得以下主要成果:
    (1)不同作用机制的抗生素对硝化系统处理效果的影响不同。其中链霉素对亚硝酸盐氧化过程抑制明显,25 mg/L阶段亚硝酸盐氮最高累积量达78 mg/L;高浓度土霉素对氨氧化过程影响显著,氨氮去除率降低至69%。针对这两个影响显著的体系进行微生物群落结构分析,发现Nitrosomonas europaea是两个体系的优势氨氧化细菌,与对照体系(Nitrosomonas unclassified)差异显著。随着链霉素浓度的升高,该体系中的硝化菌出现Nitrospira与Nitrobacter更替,而土霉素与对照体系一致,仅检出丰度极高的Nitrospira。氨氧化微生物的qPCR结果显示,链霉素、氨苄青霉素体系在25 mg/L阶段检出氨氧化古菌amoA基因,50 mg/L迅速增长2个数量级,螺旋霉素在此阶段开始有检出。表明抗生素能够促进氨氧化古菌的出现和生长,但达到一定丰度需要一个长期驯化过程。链霉素、土霉素体系在50 mg/L阶段抗性基因(ARGs)总丰度显著增加,统计分析结果显示两体系ARGs与氨氧化细菌amoA基因的相对丰度均呈显著正相关性(p < 0.01),在长期抗生素暴露过程中,氨氧化细菌可能通过获得耐药性存活下来并保持其在氨氮氧化过程中的关键作用。
    (2)针对氨基糖苷(巴龙霉素/核糖霉素)、大环内酯类(螺旋霉素)抗生素交替生产的废水处理系统,利用qPCR、高通量定量PCR(HT-qPCR)等技术对氨氧化微生物进行长周期定量追踪及抗性基因的种类和丰度调查。发现氨基糖苷类抗性基因相对丰度明显高于大环内脂类,好氧污泥高于厌氧污泥,但并未见某种抗生素压力下该类型抗性基因的明显增长。最初运行阶段,Arch-amoA基因未出,随着抗生素压力的持续作用,Arch-amoA基因相对丰度逐渐升高。调查结果再次证实,螺旋霉素、氨基糖苷等抗生素能够促进氨氧化古菌的产生和增长,对氨氧化微生物群落结构产生比较显著影响。
    (3)针对油污染严重的发酵类抗生素生产废水特征,酵母菌为主体的活性污泥预处理系统被构建。通过小试试验优化操作参数后,该技术在无锡某制药厂进行了日处理量80m3抗生素废水处理工程应用。该预处理系统能够去除大部分油,明显促进菌渣与废水固液分离。采用定量PCR与克隆文库的方法对酵母菌进行追踪,结果表明酵母菌定植成功且体系中细菌污染较少。这项研究表明真菌在抗生素生产废水处理中的应用潜力。
英文摘要:       Nitrification is the key in biological nitrogen removal schemes and is believed to be the rate-limiting step due to the slower growth rate and higher sensitivity to environmental stresses of nitrification microorganisms. Our previous study had revealed the selection of AOA in the presence of high concentrations of spiramycin in activated sludge systems. Therefore, it is important to further study on the effects of antibiotics on nitrification processes in the long-term exposure of antibiotics.
      Firstly in this study, shake flask reactors for 280 days of operation were established to investigate the effects of different antibiotics including oxytetracycline, spiramycin, streptomycin and ampicillin on the performance of nitrification processes, Microbial community structure and antibiotic resistant genes (ARGs). Secondly, a long period quantitative tracking survey were carried out about nitrifying populations and ARGs from wastewater treatment system for the alternate production of aminoglycoside (Paromomycin/Ribostamycin) and macrolides (spiramycin). Finally, a yeast-based pretreatment system was established according to the characteristics of the field wastewater. The main contents are as follows:
      (1) The effects of performance of nitrification processes differ from different antibiotics. Nitrite oxidation process was seriously affected with accumulation of nitrite in the streptomycin system and ammonia nitrogen removal was significantly inhibited at 50 mg/L oxytetracycline. We laid emphasis of further study on microbial community structure in the two systems. For ammonia-oxidizing bacteria (AOB),Nitrosomonas europaea was as the dominant ammonia-oxidizing bacteria differented from the control system (Nitrosomonas unclassified). For nitrite-oxidizing bacteria (NOB), Nitrospira disappeared and Nitrobacter was detected in the streptomycin system and only Nitrospira with high relative abundance in the oxytetracycline system. The results of qPCR showed that archaeal amoA (Arch-amoA) gene was found in streptomycin and ampicillin system at 25 mg/L stage, respectively, subsequently had the rapid growth with 2 orders of magnitude. At 50 mg/L stage Arch-amoA gene was detected in spiramycin system. No Arch-amoA genes were detected in the control and oxytetracycline systems. Statistical results showed that the relative abundance of ARGs and AOB amoA genes had a significant positive correlation (P < 0.01), which indicated AOB could survive by acquiring antibiotic resistance through long-term exposure and would keep the position as the key player in ammonia removal.
      (2) The long period quantitative tracking of ammonia-oxidizing microorganisms and abundance of ARGs were determined by qPCR and high-throughput quantitative PCR (HT-qPCR) from wastewater treatment system for the alternate production of aminoglycoside (Paromomycin/Ribostamycin) and macrolides (spiramycin). There was no significant increase in relative abundance of some antibiotic ARGs under this antibiotic exposure. The relative abundance of Arch-amoA gene increased gradually with operation of antibiotic production wastewater treatment system. The results of the investigation confirmed that the antibiotics such as spiramycin and some aminoglycosides could promote the presence and growth of ammonia-oxidizing archaea (AOA).
      (3) High residual oil content in antibiotic production waste mother liquor makes the solid-liquid separation difficulty. A yeast-based pretreatment process was established for the removal of oil and promotion of solid-liquid separation in antibiotic production wastewater treatment systems. High oil removal rates (pilot-scale: 85.0-92.0%; full-scale: 61.4-74.2%) and excellent sludge settling velocity (pilot-scale: 16.6-21.3%; full-scale: 22.6-32.0%) were found from the pilot and full-scale operation. 18S rRNA gene clone libraries and quantitative PCR were established for tracking the fates of inoculated yeasts, which indicated that yeast successfully colonized. Both of pilot and full-scale studies have proved that yeast can be used to promote the solid-liquid separation and yeast systems are a stable and effective method for oil-containing fermentation antibiotic production wastewater pretreatment.
内容类型: 学位论文
URI标识: http://ir.rcees.ac.cn/handle/311016/36968
Appears in Collections:环境水质学国家重点实验室_学位论文

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Recommended Citation:
王春艳. 抗生素对废水处理微生物的影响 及基于酵母菌的调控[D]. 北京. 中国科学院研究生院. 2016.
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