固定化工程菌对偶氮染料脱色及强化作用

利用聚亚胺酯大孔泡沫吸附固定基因工程菌Escherichia coli JM109 (pGEX-AZR),研究其对偶氮染料的脱色动力学及生物强化作用.实验表明,固定的E.coli JM109(pGEX-AZR) 对酸性大红GR的脱色动力学符合Andrews方程,动力学常数为μ_max,c、K_c、K_ic分别为49 .2 mg·(g·h)^-1、710 .43 mg·L^-1和681 .62　mg·L^-1,R²为0 .995.将固定的E.coli JM109(pGEX-AZR)按10%的比例投加到厌氧序批式活性污泥反应器中连续运行32 d,含有固定化工程菌的强化体系耐浓度冲击的能力和脱色率均高于对照体系,脱色率可以稳定在90%以上.利用RISA对其微生物群落结构进行分析,E.coli JM109(pGEX-AZR)及降解酸性大红GR的优势菌群可以在污泥体系中稳定存在.     

有机膨润土在Pb2+和p-硝基苯酚复合污染中的吸附及机理

以Pb²⁺、p-硝基苯酚为代表污染物,用实验室模拟法研究了有机膨润土在重金属-有机物混合污染体系中的吸附行为.结果表明,在有机膨润土/Pb²⁺/p-硝基苯酚体系中,溶液中共存Pb²⁺会延缓p-硝基苯酚在有机膨润土上的吸附速率;Pb²⁺与p-硝基苯酚产生竞争吸附,竞争作用的大小与溶液中p-硝基苯酚和Pb²⁺的浓度比c(酚/Pb)以及有机膨润土对污染物的吸附机制有关,相对于吸附机制以分配作用为主的100CTMAB-膨润土,Pb²⁺与p-硝基苯酚在以表面吸附为主的100TMAB-膨润土上的竞争吸附更强.     

Sequential use of ultraviolet light and chlorine for reclaimed water disinfection

Several disinfection processes of ultraviolet (UV), chlorine or UV followed by chlorine were investigated in municipal wastewater according to the inactivation of Escherichia coli, Shigella dysenteriae and toxicity formation. The UV inactivation of the tested pathogenic bacteria was not affected by the quality of water. It was found that the inactivated bacteria were obviously reactivated after one day in dark. Fluorescent light irradiation increased the bacteria repair. The increase of UV dosage could cause more damage to bacteria to inhibit bacteria self-repair. No photoreactivation was detected when the UV dose was up to 80 mJ/cm² for E. coli DH5αup, and 23 mJ/cm² for S. dysenteriae. Nevertheless, sequential use of 8 mJ/cm² of UV and low concentration of chlorine (1.5 mg/L) could effectively inhibit the photoreactivation and inactivate E. coli below the detection limits within seven days. Compared to chlorination alone, the sequential disinfection decreased the genotoxicity of treated wastewater, especially for the sample with high NH₃-N concentration.     

纳米Fe3O4负载的浮游球衣菌去除重金属离子的工艺研究

以纳米Fe₃O₄负载浮游球衣菌（Sphaerotilus natans）制备复合生物吸附剂,对此吸附剂进行表征并考察了其吸附水中重金属离子的性能.红外光谱分析表明,此复合生物吸附剂表面的主要活性基团为酰胺基（—CONH—）和羟基（—OH）.吸附性能研究表明,菌含量和流量是影响复合生物吸附剂吸附重金属离子的主要因素,在Cu²⁺初始浓度c₀<20 mg/L,菌含量1.5 g/L（菌/ Fe₃O₄＝3∶2）,流量0.96 L/h 时吸附剂对Cu²⁺的吸附效果最好;用稀盐酸对复合生物吸附剂进行再生,吸附剂可重复使用10次以上,再生液可重复使用3次; 吸附选择性为：Pb²⁺ > Cu²⁺ > Zn²⁺>Cd²⁺.     

生物炭的13C标记方法及对镉、铜的吸附性能探究

生物炭施入土壤后很难分离和检测，因而难以直观研究生物炭在土壤中的环境行为.本文采用稳定同位素标记法制备¹³C标记的生物炭，研究了¹³C标记生物炭的制备方法，以及标记前后生物炭的基本理化性质及其对Cd²⁺和Cu²⁺的吸附效果和吸附机理.结果表明，脉冲标记法可有效提高玉米植株的δ¹³C值，经过550℃限氧热解法制备的¹³C玉米秸秆生物炭（¹³C BC）的δ¹³C值为249.3‰，显著高于土壤（-23.5‰）及未标记的玉米秸秆生物炭（-25.7‰），具有显著的生物炭示踪效果.¹³C标记的生物炭的pH、CEC、比表面积、表面官能团种类及数量等理化性质与未标记生物炭相似，说明脉冲标记法制备的¹³C标记生物炭不改变生物炭的基本理化性质.吸附动力学和吸附热力学实验结果表明，¹³C标记生物炭与未标记生物炭对Cu²⁺、Cd²⁺的吸附均符合准二级动力学方程和Langmuir等温吸附方程，且吸附平衡时间、最大吸附量基本相同，表明¹³C标记生物炭不改变生物炭对Cd²⁺、Cu²⁺的吸附效果和吸附特征.XRD、FT-IR结果表明，¹³C标记生物炭和未标记生物炭具有相同的吸附机理，均为离子交换作用、静电吸附作用和阳离子-π键作用.因此，脉冲标记法制备的¹³C标记生物炭具有与生物炭相同的基本理化性质和对镉、铜的吸附行为，同时具有显著高于土壤背景的δ¹³C值，可为土壤中生物炭迁移、生物炭与重金属相互作用行为的研究提供一条途径.     

Effect of excessive cadmium chloride on the plasmids of E. coli HB101 in vivo

After Escherichia coli HB101 with plasmid pWH58, pWH98, or pTBa₅ were cultered respectively in amp LB broth which contained 50 mg/L CdCl₂ constantly for 24h, these plasmids were isolated from E. coli, and the effect of excessive CdCl₂ on the E. coli HB101 and plasmid DNA was studied by surveying the growth of E. coli HB101 and plasmid, argarose gel electrophoresis and analysis of restriction fragment length polymorphism (RFLP) of plasmids, and plasmid transformation. The results showed that 50 mg/L CdCl₂ treatment lagged the growth of E. coli HB101 for at least 4h, but after grown for 24h there were not significant differences in the growths of E. coli HB101s and the productions of plasmids between the treatment and control. These results implified that E. coli HB101 have induced adaptability to cadmium stress and excessive CdCl₂ did not inhibit the replication and amp + genes expression of plasmid DNA in vivo of E. coli significantly. 50 mg/L CdCl₂ treatment for 24 hours might cause the sequences change of plasmid DNA, but could not lead to the random breakage of plasmid DNA strands. Moreover, after 50 mg/L of CdCl₂ treatment in vivo the transformation activities of plasmid did not altered, implied excessive CdCl₂ could not affect the superhelical structure of plasmid and also not break the loop of plasmid DNA evidently.     

软锰矿微生物催化氧化烟气脱硫

利用软锰矿与微生物联合催化氧化烟气SO₂.分别选取氧化亚铁硫杆菌（T. ferrooxidans菌）和锰氧化细菌,研究微生物对SO^2-₃的氧化性能和恢复体系中Mn（Ⅱ）和Fe（Ⅲ）的活性性能.采用紫外线诱变法,选育出了对SO^2-₃和Fe²⁺转化效率高T. ferrooxidans菌的优势菌株,对Fe²⁺的完全氧化时间缩短为约24 h.通过分析细菌与软锰矿联合催化氧化脱除SO₂的脱硫效率变化、液相中SO^2-₃离子的变化,以及二者的关系,探讨了细菌在软锰矿脱硫体系中所起的作用,并用透射电子显微镜观察了反应前后锰氧化细菌的形态变化.结果表明Fe²⁺与T. ferrooxidans菌共同存在时,T. ferrooxidans菌对SO₃^2-的转化速率可达到0.015?3　g/(L·min),优于化学氧化.T. ferrooxidans菌和锰氧化菌联合软锰矿脱硫实验结果表明,细菌对软锰矿脱硫存在强化作用;锰氧化细菌促进脱硫存在适应期,T. ferrooxidans菌和锰氧化菌存在协同效应,微生物可以完成铁锰催化剂的再生循环.     

污水河道污泥中大肠杆菌抗生素抗性与金属耐受性的耦合分析

城市污水河道受人为活动的影响,可能作为抗性细菌与抗性基因的储库,给人类生产生活带来巨大的威胁.本研究以中国寒区城市污水河道污泥为取样点,研究大肠杆菌（Escherichia coli）的抗生素与金属的耐药现状,以评估中国寒区城市水环境受抗性细菌污染的情况与传播抗性的风险.利用选择性培养基从河道淤泥中分离出455株肠杆菌,并从中筛选出110株E. coli.抗生素表型结果显示：69.1%的E. coli分离株对至少1种抗生素具有抗性,其中,抗性最高的是氧四环素（42.7%）;所有的E. coli对亚胺培南和呋喃妥因没有抗性;分离株E. coli DL27对检测的17种抗生素中的13种具有抗性,分离株E. coli DL18、分离株E. coli DL46对12种抗生素具有抗性.金属抗性表型检测结果显示：40.9%的E. coli对银（Ag）具有耐受性,3.6%的分离株对汞（Hg）、锌（Zn）具有耐受性,所有菌株对铜（Cu）无耐受性;且发现分离株E. coli DL35对9种抗生素和2种金属具有抗性.对E. coli群体15种抗生素抗性基因（Antibiotic Resistance Genes,ARGs）、4种金属耐受性基因（Metal Resistance Genes,MRGs）、1种整合酶基因的检测结果显示,ARGs中tetA（56.4%）、blaTEM（26.4%）、sul2（24.5%）、sul1（22.7%）基因的检出率最高,MRGs的检出率分别为20.0%（silB）、13.6%（silE）、8.2%（merC）、16.4%（pcoE）,整合酶基因intI1的检出率为2.7%.研究表明,在河道污泥中不仅存在多重抗生素抗性E. coli,而且存在同时具有抗生素抗性和金属耐受性的E. coli.河道污泥是抗性细菌和抗性基因的储库,对人类生产生活构成了潜在的巨大威胁.     

Ni2+生物吸附动力学及吸附平衡研究

研究了金属离子Ni²⁺在酿酒酵母上的生物吸附特性,内容包括生物吸附动力学和吸附等温线.生物吸附动力学结果表明,当Ni²⁺初始浓度为65.6 mg/L时,Ni²⁺在酿酒酵母上的生物吸附可以分为2个阶段,第1阶段为物理吸附,在10　min内快速达到平衡.Ni²⁺在酵母上的吸附过程可以很好地用准二级动力学方程来描述 （R²=0.999）,动力学参数k₂为0.018 4 g/(mg·min),q_e为5.96 　mg/g.吸附等温线结果表明,Ni²⁺在酿酒酵母上的生物吸附可以用Langmuir和Freundlich方程来描述,最大吸附量q_max为6.32 　mg/g.酿酒酵母可用于处理低浓度的含Ni²⁺废水.     

镍钴转运酶NiCoT基因的克隆表达及基因工程菌对镍离子的富集

利用PCR技术从Staphylococcus aureus/ ATCC6538基因组中扩增出大小为1?053 bp的镍钴转运酶基因NiCoT gene,将其连接到pET-3c载体上构建重组质粒,并转化至E.coli BL21.筛选阳性菌并经酶切分析和PCR扩增双重鉴定.核苷酸序列测定及分析结果与GenBank中报道的同类基因相似性高达97%以上,表明其具有正确的NiCoT基因核苷酸序列.重组菌的SDS-PAGE结果图谱中,在相对分子量为39?000附近有特异性蛋白条带,大小符合预测值,表明NiCoT基因在E.coli BL21中成功表达.基因工程菌在IPTG用量为1.00 mmol·L^-1,诱导时间为4 h的条件下培养对镍离子的富集能力最高.在不同镍离子浓度时,基因工程菌对溶液中Ni²⁺的平衡富集量为11.33 mg·g^-1,与原始宿主菌相比提高了3倍.对基因工程菌吸附镍和钴的实验表明,Staphylococcus aureus ATCC6538的NiCoT对镍具有较高的特异性和富集容量,属于第Ⅲ类镍钴转运酶.     

基于CellSense生物传感器的垃圾渗滤液毒性分析

采用基于大肠杆菌(E.coli)的CellSense生物传感器,对上海某垃圾填埋场渗滤液处理系统垃圾渗滤液原液(1号)、兼性塘(2号)、曝气氧化塘(3号)、矿化垃圾生物床(4号)和FeC内电解(5号)各处理单元的水质毒性进行跟踪分析.结果表明,在该垃圾渗滤液的处理过程中,由于渗滤液的基质作用和难降解有机物生成的毒性中间产物等的影响,处理工艺中部分单元的水质毒性出现升高现象.CellSense生物传感器分析得上述1~5号单元的水质综合毒性分别为90%(EC10)、80%(EC30)、60%(EC50)、80%(EC50)和2%(EC50),毒性分析结果与对应的COD和氨氮等指标无明显相关性.     

基于枯草芽孢杆菌微生物传感器的毒性分析

采用枯草芽孢杆菌(Bacillus subtilis)为指示生物的微生物传感器毒性分析系统,对重金属(Hg2+、Cu2+、Zn2+、Cr6+、Cd2+、Pb2+和Co2+)、有机污染物[邻氯苯酚(2-CP)、2,4-二氯酚(2,4-DCP)、邻硝基酚(2-NP)、对硝基酚(4-NP)、四环素和十二烷基苯磺酸钠]及石油废水等的生物急性毒性进行分析.结果表明,对数生长后期和稳定期的Bacillussubtilis微生物传感器具有良好的毒性分析性能,Cd2+、Zn2+、Cr6+、Cu2+、Hg2+、Pb2+对Bacillus subtilis的EC50分别为47.3,10.9,14.0,2.6,0.8,100.1mg/L,Co2+的EC30为56.6mg/L,2-CP、2,4-DCP、2-NP、4-NP、四环素和十二烷基苯磺酸钠的EC50分别为559.6,450.8,588.5,487.0,121.3,558.9mg/L,该微生物传感器能真实反映石油废水的毒性情况.     

The study of biological activity of transformation products of diclofenac and its interaction with chlorogenic acid

In the present work we compared the biological activity of DCF, 4′-OHDCF and 5-OHDCF as molecules of most biodegradation pathways of DCF and selected transformation products (2-hydroxyphenylacetic acid; 2,5-dihydroxyphenylacetic acid and 2,6-dichloroaniline) which are produced during AOPs, such as ozonation and UV/H₂O₂. We also examined the interaction of DCF with chlorogenic acid (CGA). CGA is commonly used in human diet and entering the environment along with waste mainly from the processing and brewing of coffee and it can be toxic for microorganisms included in activated sludge. In the present experiment the evaluation of following parameters was performed: E. coli K-12 cells viability, growth inhibition of E. coli K-12 culture, LC₅₀ and mortality of Chironomus aprilinus, genotoxicity, sodA promoter induction and ROS generation. In addition the reactivity of E. coli SM recA:luxCDABE biosensor strain in wastewater matrices was measured. The results showed the influence of DCF, 4′-OHDCF and 5-OHDCF on E. coli K-12 cells viability and bacteria growth, comparable to AOPs by-products. The highest toxicity was observed for selected, tested AOPs by-products, in comparison to the DCF, 4′-OHDCF and 5-OHDCF. Genotoxicity assay indicated that 2,6-dichloroaniline (AOPs by-product) had the highest toxic effect. The oxidative stress assays revealed that the highest level of ROS generation and sodA promoter induction were obtained for DCF, 4′-OHDCF and 5-OHDCF, compared to other tested compounds. We have also found that there is an interaction between chlorogenic acid and DCF, which resulted in increased toxicity of the mixture of the both compounds to E. coli K-12, comparable to parent chemicals. The strongest response of E. coli SM biosensor strain with recA:luxCDABE genetic construct in filtered treated wastewaters, comparable to control sample was noticed. It indicates, that E. coli SM recA:luxCDABE biosensor strains is a good tool for bacteria monitoring in wastewater environment. Due to toxicity and biological activity of tested DCF transformation products, there is a need to use additional wastewater treatment systems for wastewater contaminated with pharmaceutical residues.     

Effects of lead , cadmium , mercury chlorides and aquaticenvironmental samples on the growth of Escherichia coli

In this paper it was reported that PbCl₂ , CdCl₂ , HgCl₂, and aquatic environmental samples containing Al, Cd, Cu, Pb, Sr, Zn and other heavy metal ions as well as organic chemicals suppressed thegrowth of E. coli only at higher exposed concentrations. The stimulative effects of PbCl₂ and HgCl₂ on thegrowth of E. coli were clearly showed at lower concentrations and longer time. It suggested that the toxic effects of heavy metal ions and other pollutants on the growth of E. coli are variable according to the differentexposed levels.     

Effects of lead，cadmium，mercury chlorides and aquatic environmental samples on the growth of Escherichia coli

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A GFP-based bacterial biosensor with chromosomally integrated sensing cassette for quantitative detection of Hg(II) in environment

A mercury biosensor was constructed by integrating biosensor genetic elements into E. coli JM109 chromosome in a single copy number, using the attP/attB recombination mechanism of λphage. The genetic elements used include a regulatory protein gene (merR) along with operator/promoter (O/P) derived from the mercury resistance operon from pDU1358 plasmid of Serratia marcescens. The expression of reporter gene gfp is also controlled by merR/O/P. Integration of the construct into the chromosome was done to increase the stability and precision of the biosensor. This biosensor could detect Hg(II) ions in the concentration range of 100-1700 nmol/L, and manifest the result as the expression of GFP. The GFP expression was significantly different (P ≤ 0.05) for each concentration of inducing Hg(II) ions in the detection range, which reduces the chances of misinterpretation of results. A model using regression method was also derived for the quantification of the concentration of Hg(II) in water samples.     

Chemical composition and Zn bioavailability of the soil solution extracted from Zn amended variable charge soils

A study on variable charge soils (volcanic Italian and podzolic Scottish soils) was performed to investigate the influence of soil properties on the chemical composition of soil solution. Zinc speciation, bioavailability and toxicity in the soil solution were examined. The soils were spiked with increasing amounts of Zn (0, 100, 200, 400 and 1000 mg/kg) and the soil solutions were extracted using rhizon soil moisture samplers. The pH, total organic carbon (TOC), base cations, anions, total Zn and free Zn2+ in soil solution were analysed. A rapid bioassay with the luminescent bacterium Escherichia coli HB101 pUCD607 was performed to assess Zn toxicity. The influence of soil type and Zn treatments on the chemical composition of soil solution and on Zn toxicity was considered and discussed. Di erent trends of total and free Zn concentrations, base cations desorption and luminescence of E. coli HB101 pUCD607 were observed. The soil solution extracted from the volcanic soils had very low total and free Zn concentrations and showed specific Zn2+/Ca2+ exchange. The soil solution from the podzolic soil had much higher total and free Zn concentrations and showed no evidence of specific Zn2+/Ca2+ exchange. In comparison with the subalkaline volcanic soils, the acidic podzol showed enhanced levels of toxic free Zn2+ and consequently stronger effects on E. coli viability.     

铬污染土壤稳定化处理对蚯蚓的毒性效应

浸出毒性测试是评价重金属污染土壤稳定化处理效果的主要方法,但该方法不能准确反映稳定化处理后土壤的生物毒性变化.本研究以多硫化钙(CPS)稳定化处理的电镀厂铬(Cr)污染土壤为对象,以赤子爱胜蚓为指示生物,研究评估土壤稳定化处理的生物毒性效应.结果表明,在Cr污染土壤中,随着CPS投加量的增加,蚯蚓的毒性效应逐渐减弱随后增强.高浓度Cr污染土壤经过CPS稳定化处理后仍然具有明显的生物毒性,CPS与土壤中Cr(VI)投加比(物质的量比,下同)为3时土壤浸出毒性虽满足浸出标准,但对蚯蚓的7 d致死效应高达100%,且暴毒期间出现肉眼可见的外部形态变化,回避率为83.3%,在暴毒第5 d时蚯蚓体内总Cr含量为20.9 mg·kg~(-1).CPS对蚯蚓具有较强的生物毒性作用,在不含Cr污染的对照组土壤中,CPS投加量与稳定化处理Cr污染土壤中稳定化药剂投加比为5相同时,蚯蚓的7 d急性毒性致死率为100%.蚯蚓的致死率和外部形态变化可快速直观地反映重金属污染土壤的生物毒性,可为土壤稳定化处理的安全性与生物毒性评估提供参考依据.     

江苏如东滩涂贝类养殖区表层沉积物中重金属来源分析及其潜在生物毒性

通过2010年对江苏如东滩涂贝类养殖区表层沉积物中重金属Cu、Zn、Pb、Cd、Hg、As含量的调查分析,应用地质累积指数(index of geo-accumulation,Igeo)对各重金元素的污染状况进行了评价,结果表明重金属污染可分为3类:Cu、Cd、Hg为无污染;Pb、As为中～无污染;Zn为中度污染,重金属污染程度顺序为Zn>As>Pb>Cd>Cu>Hg.根据沉积物质量基准(sediment quality guideline,SQG),表层沉积物中Cd、As的含量在各采样站位分别表现为无潜在生物毒性效应,偶尔有潜在生物毒性效应,Cu、Pb、Zn、Hg的含量则在部分采样站位偶尔有潜在生物毒性效应,其中仅有Zn在部分采样站位有频繁发生的潜在生物毒性效应.毒性单位评价(toxic unit,TU)结果表明,仅有1个采样站位具有明显的急性毒性,其余站位无急性生物毒性效应.主成分分析(principal component analysis,PCA)的结果表明,前2个主成分的贡献率分别为37.56%、33.71%,揭示了重金属污染主要有2个来源:工业点源排污和交通航运污染源.     

一种新的生物毒性试验方法

提出了一种新的生物毒性试验方法。应用氧对萤光淬灭的原理研制了一种呼吸计量仪。根据有毒物质对微生物细胞的呼吸抑制程度来评估其毒性大小。应用该呼吸计量仪，试验了多种重金属对活性污泥、土壤细菌及大肠杆菌的毒性影响。结果表明，应用该呼吸计量仪进行毒性试验是可行的。它具有灵敏、快速、高效等特点，可望成为一种大规模的生物毒性试验和环境检测装置。