土壤环境中肠道致病菌的多重PCR检测研究初探

建立同时检测大肠杆菌、沙门氏菌、金黄色葡萄球菌、福氏志贺氏菌、铜绿假单胞菌等5种土壤常见肠道致病菌的多重PCR检测技术,为这些肠道致病菌感染的快速诊断提供实验依据.根据这些肠道病原菌的毒素基因、高度保守基因及特异性基因分别合成5对特异性引物,应用PCR扩增技术对目的菌株进行特异性检测.实验结果表明,5对寡核苷酸引物都具有较高的特异性和专一性,多重PCR检测限达到104cfu·g-1.多重PCR应用于土壤样品分析,极大的缩短了检测时间(仅需3～4h)、降低了检测成本,对控制病原菌的传播具有重要意义,可推广应用于环境监测、水源检测、食品卫生监督、商品检验检疫等领域.     

Uptake of radioiodide by Paenibacillus sp., Pseudomonas sp., Burkholderia sp. and Rhodococcus sp. isolated from a boreal nutrient-poor bog

Radionuclides, like radioiodine(~(129)I), may escape deep geological nuclear waste repositories and migrate to the surface ecosystems. In surface ecosystems, microorganisms can affect their movement. Iodide uptake of six bacterial strains belonging to the genera Paenibacillus,Pseudomonas, Burkholderia and Rhodococcus isolated from an acidic boreal nutrient-poor bog was tested. The tests were run in four different growth media at three temperatures. All bacterial strains removed iodide from the solution with the highest efficiency shown by one of the Paenibacillus strains with 99% of iodide removed from the solution in one of the used growth media. Pseudomonas, Rhodococcus and one of the two Paenibacillus strains showed highest iodide uptake in 1% yeast extract with maximum values for the distribution coefficient(K_d) ranging from 90 to 270 L/kg DW. The Burkholderia strain showed highest uptake in 1% Tryptone(maximum K_d170 L/kg DW). The Paenibacillus strain V0-1-LW showed exceptionally high uptake in 0.5% peptone + 0.25% yeast extract broth(maximum K_d 1,000,000 L/kg DW). Addition of 0.1% glucose to the 0.5% peptone + 0.25% yeast extract broth reduced iodide uptake at 4℃ and 20℃ and enhanced iodide uptake at 37℃ compared to the uptake without glucose. This indicates that the uptake of glucose and iodide may be competing processes in these bacteria. We estimated that in in situ conditions of the bog,the bacterial uptake of iodide accounts for approximately 0.1%–0.3% of the total sorption of iodide in the surface, subsurface peat, gyttja and clay layers.     

摇蚊幼虫与霍普水丝蚓扰动对河流沉积物中氮、磷、氧释放的影响

定量研究河床沉积物中的生物扰动对污染物的影响效应,对于维护河流健康具有重要的理论指导作用.基于室内实验模拟摇蚊和霍普水丝蚓对河床沉积物的扰动过程,研究两种生物扰动作用对沉积物中氮、磷释放以及生物扰动对上覆水中溶解氧的影响.在相同的沉积物环境中分别放入单一物种,实验结果显示,相对于空白组,摇蚊幼虫组的上覆水中氨氮、总氮、总磷平均净增量分别为2.32、0.787、0.105 mg · L^-1,霍普水丝蚓组的氨氮、总氮、总磷平均净增量分别为0.72、0.462、0.063 mg · L^-1,表明摇蚊幼虫和霍普水丝蚓的扰动作用均能促进沉积物中氨氮、总氮、总磷向上覆水中的释放,且摇蚊幼虫对污染物释放的作用效果比水丝蚓更明显;当摇蚊幼虫和水丝蚓混合放入后,摇蚊幼虫对沉积物中氨氮、总氮、总磷释放起主要促进作用;在混合实验组(摇蚊23条,霍普水丝蚓47条)中,氨氮、总氮、总磷的平均净增量均低于单一物种的摇蚊组和水丝蚓组,表明在该组中,摇蚊幼虫和霍普水丝蚓在沉积物中共同扰动作用对氮、磷释放的促进效果减弱;对上覆水中溶解氧浓度而言,摇蚊所占比例越高,溶解氧浓度就越低,且上覆水中的溶解氧浓度变化,会影响生物扰动对沉积物氮、磷释放的效果.     

农业废物好氧堆肥中环境因子对nirK、nirS和nosZ数量的影响

应用定量聚合酶链式反应(real-time PCR)技术对农业废物好氧堆肥过程中参与反硝化过程的功能基因(nirK、nirS和nosZ)丰度在堆体不同位置处随时间的变化情况进行了研究.结果表明,随着堆肥进程,3种基因数量整体呈现出先升后降的变化规律,且不同位置处的反硝化基因数量之间存在着显著的差异性.使用Canoco 4.5软件对获得的反硝化功能基因丰度数据与不同时期不同层次堆体温度、pH、含水率、NH4+-N、NO3--N和水溶性有机碳(WSC)等环境因子的相关性进行冗余分析(redundancy analysis,RDA).基于手动选择的RDA分析结果表明,WSC、NH4+-N和堆体温度对反硝化基因丰度有着显著的影响(P<0.05),且前2个因子达到了极显著水平(P<0.01).应用t-value回归分析方法单独分析每种环境因子与3种基因的相关性,其中nirK与温度和pH显著正相关(P<0.05),nirS与温度显著正相关(P<0.05),nosZ与NH4+-N显著正相关(P<0.05)、与WSC显著负相关(P<0.05).     

给水管网中耐氯性细菌的灭活特性研究

使用4种常见消毒剂对从实际管网中分离出来的7株耐氯性细菌进行消毒实验.结果表明,这7株细菌均具有较高的耐氯性,其中1株耐氯性最高的类龟分支杆菌自由氯99.9%灭活的CT值为120 mg.(L.min)-1,另外2株血红鞘氨醇单胞菌和甲基杆菌99.9%灭活的CT值分别为7 mg.(L.min)-1和4 mg.(L.min)-1.比较4种消毒剂的消毒效果发现,二氧化氯和单过硫酸氢钾的消毒效果较好,能够在30 min内使分支杆菌的灭活率达到5个数量级.自由氯由于衰减较快,消毒效果不佳.一氯胺能够维持一定的消毒剂浓度,但由于其氧化性较弱,因此需要提高浓度,才能满足消毒要求.能在1 h内灭活3个数量级以上分支杆菌的消毒剂投加量为:3.0 mg/L一氯胺、1.0 mg/L二氧化氯(以Cl2计)和1.0 mg/L单过硫酸氢钾(以Cl2计).考虑到我国水厂消毒的实际情况,建议采用间歇性提高一氯胺浓度或改换二氧化氯消毒的方法,提高对耐氯性细菌的灭活效果.     

白洋淀荷茎叶提取液对铜绿微囊藻及四尾栅藻化感效应

本研究探讨了活体荷不同部位(茎和叶)提取出液对铜绿微囊藻、四尾栅藻生长化感效应,为推广荷遏制水华暴发及制作抑藻制剂提供理论基础.实验设计了5个浓度梯度,结果表明,荷叶浸出液对藻类的抑制效果优于荷茎浸出液,当荷叶浸出质量浓度为25 g.L-1时,对铜绿微囊藻及四尾栅藻的抑制率分别为71.33%、78.14%.浸出液成分的GC-MS分析表明,荷叶和茎浸出液都含有丙酰胺,质量浓度分别为1.1 mg.L-1、0.2 mg.L-1.并根据概率计算法,分别计算了两种藻类的半浓度效应.     

典型油田多环芳烃污染对土壤反硝化微生物群落结构的影响

油田区土壤具有潜在的PAHs(polycyclic aromatic hydrocarbons)污染风险,而以硝酸根为电子受体的反硝化作用可能在PAHs的厌氧代谢中起到重要作用.以具有50多年历史的江汉油田区域为对象,从该油田的油井口附近采集了9个土壤样品,编号为JH-1~JH-9,以反硝化相关的nir K(Cu-亚硝酸还原酶基因)和nirS(细胞色素cd1-亚硝酸还原酶基因)为分子标识,通过定量PCR及克隆文库结合T-RFLP(terminal-restriction fragment length polymorphism)的方法,研究典型油田区土壤反硝化微生物的群落结构,并探讨其与土壤环境因子之间的关系.结果表明,该油田区土壤中nirK基因的丰度高于nirS基因,PAHs含量最高的土壤样品(JH-4)中反硝化功能基因nir K和nir S的丰度均最低,相关性分析表明,土壤nir K及nirS基因的丰度均与土壤PAHs含量呈显著负相关(nirK:R2=0.54,P0.05;nirS:R~2=0.58,P0.05).克隆文库及T-RFLP的结果则表明,该油田土壤中nirK基因的群落组成在不同样品间的变异较大,且PAHs含量最高的JH-4中该基因的群落组成与其它各样品有明显的不同,RDA(redundancy analysis)的分析结果进一步表明除有效氮、有效磷外,土壤PAHs含量也是影响nirK型反硝化微生物群落组成的重要因子.相较于nirK,该油田区土壤中nirS基因的群落组成在不同样品间的差异较小,但发现nirS型假单胞菌的丰度与土壤PAHs含量呈正相关,表明具备较强有机污染物降解能力的假单胞菌属可能在该区域土壤PAHs的反硝化代谢中起到重要作用.     

黄连根茎浸提物对绿藻的毒理作用

黄连根茎和制剂具有抗菌等作用,广泛用于水产养殖,所造成水生态风险需要评估.试验设置总生物碱为0(CK),0.088(T1)、0.44(T2)和1.76 mg·L-1(T3)的黄连根茎浸提液(CRE)4种处理,研究了对斜生栅藻和蛋白核小球藻的毒理作用.结果表明,T1抑制绿藻生长,T2和T3使绿藻生长和繁殖停止;它们均显著降低绿藻叶绿素和蛋白质含量,说明CRE抑制光合作用和蛋白质合成是绿藻生长繁殖速率降低和死亡的直接原因.CRE使氢离子和胞内物质外流,导致藻液p H值显著降低和电导率提高.在T1和T2处理中,绿藻细胞SOD活性先升后降;在T3处理中,SOD活性显著降低.说明在CRE暴露初期,低中浓度的CRE诱导绿藻细胞产生抗性,随暴露时间增长或直接暴露在高浓度的CRE下,抗氧化酶系统被破坏.同样,随着CRE浓度增大,丙二醛含量增加,意味着绿藻细胞膜结构破坏,透性增加.CRE总体上对蛋白核小球藻的危害作用大于斜生栅藻.在水产养殖中,滥用黄连根茎或制剂,以及大规模集约化种植黄连对水体初级生产力具有潜在的生态风险.     

Differences in nitrite-oxidizing communities and kinetics in a brackish environment after enrichment at low and high nitrite concentrations

Nitrite accumulation in shrimp ponds can pose serious adverse effects to shrimp production and the environment.This study aims to develop an effective process for the enrichment of ready-to-use nitrite-oxidizing bacteria(NOB)inocula that would be appropriate for nitrite removal in brackish shrimp ponds.To achieve this objective,the effects of nitrite concentrations on NOB communities and nitrite oxidation kinetics in a brackish environment were investigated.Moving-bed biofilm sequencing batch reactors and continuous moving-bed biofilm reactors were used for the enrichment of NOB at various nitrite concentrations,using sediment from brackish shrimp ponds as seed inoculum.The results from NOB population analysis with quantitative polymerase chain reaction(q PCR)show that only Nitrospira were detected in the sediment from the shrimp ponds.After the enrichment,both Nitrospira and Nitrobacter coexisted in the reactors controlling effluent nitrite at 0.1 and 0.5 mg-NO_2~--N/L.On the other hand,in the reactors controlling effluent nitrite at 3,20,and 100 mg-NO_2~--N/L,Nitrobacter outcompeted Nitrospira in many orders of magnitude.The half saturation coefficients(Ks)for nitrite oxidation of the enrichments at low nitrite concentrations(0.1 and 0.5 mg-NO_2~--N/L)were in the range of 0.71–0.98 mg-NO_2~--N/L.In contrast,the Ksvalues of NOB enriched at high nitrite concentrations(3,20,and 100 mg-NO_2~--N/L)were much higher(8.36–12.20 mg-NO_2~--N/L).The results suggest that the selection of nitrite concentrations for the enrichment of NOB inocula can significantly influence NOB populations and kinetics,which could affect the effectiveness of their applications in brackish shrimp ponds.     

三种大型海藻对对氯苯胺的去除行为及光合生理响应

以龙须菜、坛紫菜和石莼3种大型海藻为实验材料,通过模拟实验,研究并比较了它们对不同浓度对氯苯胺(PCA)的去除行为,以及石莼对PCA的光合生理响应.实验结果显示,7d实验结束时,3种藻类对海水中PCA的最终去除率排序分别为:6mg·L-1PCA处理组,龙须菜(58.7%)<坛紫菜(75.7%)<石莼(94.9%);60mg·L-1PCA处理组,龙须菜(50.2%)<石莼(61.3%)<坛紫菜(74.9%);海藻的培植密度越大,PCA的去除率越高,在受PCA污染的海域中大面积种植大型海藻来有效地处理修复此类污染在理论上是可行的.在不同浓度PCA处理过程中,石莼的生长受到抑制,有效光化学效率、光合作用和呼吸作用呈现出低浓度(<5mg·L-1)促进、高浓度(>10mg·L-1)抑制的现象.叶绿素、类胡萝卜素和可溶性蛋白含量在各处理组之间没有表现出确定趋势,表明PCA对石莼生长的影响可能是由于呼吸速率的增加而引起的.     

Enhanced methane production in an anaerobic digestion and microbial electrolysis cell coupled system with co-cultivation of Geobacter and Methanosarcina

The anaerobic digestion(AD)and microbial electrolysis cell(MEC)coupled system has been proved to be a promising process for biomethane production.In this paper,it was found that by co-cultivating Geobacter with Methanosarcina in an AD–MEC coupled system,methane yield was further increased by 24.1%,achieving to 360.2 m L/g-COD,which was comparable to the theoretical methane yield of an anaerobic digester.With the presence of Geobacter,the maximum chemical oxygen demand(COD)removal rate(216.8 mg COD/(L·hr))and current density(304.3 A/m_3)were both increased by 1.3 and 1.8 fold compared to the previous study without Geobacter,resulting in overall energy efficiency reaching up to 74.6%.Community analysis demonstrated that Geobacter and Methanosarcina could coexist together in the biofilm,and the electrochemical activities of both were confirmed by cyclic voltammetry.Our study observed that the carbon dioxide content in total gas generated from the AD reactor with Geobacter was only half of that generated from the same reactor without Geobacter,suggesting that Methanosarcina may obtain the electron transferred from Geobacter for the reduction of carbon dioxide to methane.Taken together,Geobacter not only can improve the performance of the MEC system,but also can enhance methane production.     

模拟自然水体中藻-溞-草间的相互作用研究

为了解氮磷浓度对生物操纵效果及同时恢复水生植被的影响,以小球藻、大型溞和金鱼藻分别作为浮游植物、浮游动物和大型沉水植物的代表,模拟自然条件研究了不同氮磷浓度对三者生长和相互作用的影响.结果表明:金鱼藻和小球藻共培养时,在氮浓度介于2.92～12.60mg·L-1、磷浓度介于0.06 ～0.85 mg·L-1,金鱼藻增长不明显,甚至出现负增长;而小球藻的增长率则远高于金鱼藻.小球藻和大型溞共培养时,氮、磷浓度分别为1.26～ 10.53 mg·L-1和0.04～1.16 mg·L-1时均占据优势,对小球藻的抑制效果显著,抑藻及氮磷去除效果明显好于金鱼藻和小球藻共培养,而且磷的去除效果优于氮.三者共培养时,在氮浓度介于3.15 ～23.92 mg·L-1、磷浓度介于0.07 ～0.64 mg·L-1时,大型溞与金鱼藻的增长率都较高,而小球藻则维持在较低的增长水平,水质改善效果较好;当氮、磷浓度分别升至25.95 mg·L-1和1.18 mg·L-1时,大型潘和金鱼藻的增长率均下降,水质变差.大型溞和金鱼藻的联合控藻效果好于其单一的控制效果,该效果明显受到氮磷浓度的影响.     

Rhizobium sp.W33对不同植物吸收铜和根际分泌物的影响

通过盆栽试验,采用1株植物促生细菌Rhizobium sp.W33接种黑麦草、狼尾草、高羊茅、紫花苜蓿、猪屎豆、油菜和印度芥菜等7种不同植物,考察了Cu胁迫下菌株对植物生物量、Cu含量和根际土壤中分泌物的影响,并筛选根瘤菌-植物联合修复重金属污染土壤的组合体系,以期初步探明细菌-植物联合修复重金属污染土壤的机制.结果表明,接种Rhizobium sp.W33后,有5种植物的干重增加了11%~56%,增加幅度为印度芥菜黑麦草高羊茅狼尾草油菜,紫花苜蓿和猪屎豆的干重下降.黑麦草、狼尾草、高羊茅和猪屎豆根内Cu含量接近或超过1000mg·kg-1,Cu大量积累在植物根部.Rhizobium sp.W33能够显著促进黑麦草吸收Cu(p0.05),并提高其对Cu的富集系数和转移系数,增加根部和地上部的Cu总量.苹果酸是供试植物-细菌联合修复Cu污染土壤的主要有机酸种类,接种根瘤菌W33后黑麦草、狼尾草根际的苹果酸和水溶性糖含量降低,过氧化氢酶活性增加,其中,苹果酸、水溶性糖含量与黑麦草中Cu含量呈显著相关性(p0.05).因此,根瘤菌W33-黑麦草联合体系修复重金属Cu污染土壤的效果最好,具有应用于重金属Cu污染土壤稳定修复的潜力.     

胶质芽孢杆菌对印度芥菜富集土壤Cd及土壤pH的影响

采用根袋法盆栽试验,研究接种活菌量浓度分别为1×1010 cfu·kg-1 (T1)和2×1010 cfu·kg-1(T2)的胶质芽孢杆菌,对印度芥菜富集土壤Cd、土壤有效态Cd含量及土壤pH的影响.结果表明,以不接种菌液作为对照,处理T1、T2地上部分生物富集系数分别为13.58、16.83,与对照相比分别提高42.95％、77.16％;地下部分生物富集系数分别为4.35、4.83,分别比对照提高24.64％、38.40％;土壤的净化率分别是对照的1.73、2.20倍.对照组土壤有效态Cd含量随着时间的延长而降低,根际和非根际土壤分别降低32.93％和45.54％;但T1和T2处理中,根际、非根际土壤有效态Cd含量分别提高15.15％、31.73％和36.54％、45.28％.收获时,处理T1、T2根际土壤pH降低率分别是对照的1.08、1.10倍.不同浓度胶质芽孢杆菌处理下土壤pH与有效态Cd含量均呈显著负线性关系.综上,胶质芽孢杆菌主要是通过改变土壤pH来影响土壤Cd生物有效性,从而促进超富集植物的修复效果.本研究可为微生物辅助植物修复重金属污染土壤提供一定理论依据和实践指导.     

凤眼莲化感物质对铜绿微囊藻、斜生栅藻生长及细胞数相对比例的影响

为探索化感物质在蓝藻水华治理方面的应用潜力,选取在凤眼莲根系分泌物中发现的5种化感物质（亚油酸、亚油酸甘油酯、丙酰胺、N-苯基—2-萘胺和壬酸）,研究了3种剂量下（0.1、1.0、10.0mg.L-1）其对单一、混合培养铜绿微囊藻和斜生栅藻生长的影响.结果发现,化感物质成分、剂量和藻类培养方式对铜绿微囊藻和斜生栅藻生长...     

大气NOx测量准确性问题分析

采用美国热电公司的NO-N02-NOx分析仪(Model 42i TL)和NOy分析仪(Model 42i NOy),在2011年1月-10月期间,对北京城市大气中的NO、NOx和NOy进行了连续在线测量.两台仪器对NO的测量结果具有较高的一致性(r＞0.998,p＜0.01),说明两台仪器测量精度基本一致.将NOy分析仪(Model 42i NOy)采样口处安装和不安装颗粒物过滤膜两种条件下测量的NOy结果与NO-NO2-NOx分析仪所测的NOx结果分别进行了相关性分析([NOy] =0.989x[NOx],R2 =0.993;[NOy] =1.134 ×[NOx],R2=0.959),得出以下两个结论:①以颗粒态硝酸盐为代表的颗粒物是造成NOx与NOy偏差的主要原因,其所占比例在10％左右;②目前Model 42i-TL所测NOx的浓度水平较真实值偏高,其测量值更接近气态NOy的浓度水平.为估算NO2测值被高估的程度(以2011年8月6日-15日观测数据为例),用扣除部分气态NOz物种(HONO、HNO3、PAN、PPN)的修正方法,推论出在夏季N02被高估约7％(R2=0.968).     

铜绿假单胞菌对白腐菌产木质纤维素降解酶的影响

采用固态发酵的方法,考察了两株不同型号的铜绿假单胞菌（AB93066和ATCC9027）对白腐菌（黄孢原毛平革菌）产木质纤维素降解酶（木素过氧化物酶LiP、锰过氧化物酶MnP和羧甲基纤维素酶CMCase）的影响,并对酶提取液的pH值、表面张力及稻草降解残渣中挥发性有机质含量的变化进行了分析.结果表明,添加5.72%的A...     

Effects of silver adsorbed on fumed silica, silver phosphate glass, bentonite organomodified with silver and titanium dioxide in aquatic indicator organisms

In order to reduce the level of transmission of diseases caused by bacteria and fungi, the development of antimicrobial additives for use in personal care, hygiene products, clothing and others has increased. Many of these additives are based on metals such as silver and titanium. The disposal of these products in the environment has raised concerns pertaining to their potential harmfulness for beneficial organisms. The objective of this study was to evaluate the influence of the shape, surface chemistry, size and carrier of three additives containing silver and one with titanium dioxide (TiO₂) on microcrustacean survival. Daphnia magna was used as a bioindicator for acute exposure test in suspensions from 0.0001 to 10,000 ppm. Ceriodaphnia dubia was used for chronic test in TiO₂ suspensions from 0.001 to 100 ppm. D. magna populations presented high susceptibility to all silver based additives, with 100% mortality after 24 hr of exposure. A different result was found in the acute experiments containing TiO₂ suspensions, with mortality rates only after 48 hr of incubation. Even on acute and chronic tests, TiO₂ did not reach a linear concentration-response versus mortality, with 1 ppm being more toxic than 10,000 ppm on acute test and 0.001 more toxic than 0.01 ppm on chronic assay. Silver based material toxicity was attributed to silver itself, and had no relation to either form (nano or ion) or carrier (silica, phosphate glass or bentonite). TiO₂ demonstrated to have a low acute toxicity against D. magna.     

毛竹入侵对常绿阔叶林主要树种的化感作用研究

为探讨毛竹入侵对周边森林环境的影响,以毛竹浸提液处理苦槠、青冈种子,测定种子萌发效应.结果表明,两树种种子萌发与浸提液浓度和来源有关.浓度效应表现为高抑低促的作用规律,高浓度处理下苦槠的发芽率为对照的82.3%~102.2%,青冈为对照的80%~90.9%,而低浓度处理下分别为对照的101.7%~107.6%和94.9%~109.1%.除低浓度处理对苦槠发芽速度无影响外,其他处理均对两树种发芽速度有显著的延缓作用(P<0.05).高浓度浸提液对苦槠根系活力有显著的抑制作用(P<0.05),抑制率为41.1%~62.4%.不同来源浸提液处理间差异明显,根系浸提液处理的种子发芽率最低.苦槠芽苗的芽长与不同来源浸提液间差异不显著(P>0.05),而青冈芽苗芽长与不同来源浸提液间差异显著(P<0.05),茎叶、根系与枯落物浸提液高于土壤浸提液.这些结果说明毛竹具有潜在的化感作用,可能有利于其入侵周边森林,从而危害周边森林环境.