污水处理厂中红霉素抗药性基因的污染特征及选择性因子

采用固相萃取-高效液相色谱串联质谱(SPE-HPLC-MS/MS)检测分析了上海某污水处理厂中12种不同药品和个人护理品(pharmaceutical and personal care products,PPCPs)的含量水平和分布特征,采用实时荧光定量PCR方法定量检测分析了7种红霉素抗药性基因(ERY-ARGs)在污水处理厂中的分布和丰度变化.结果表明,在污水处理厂中共检出5种PPCPs(包括磺胺甲唑、红霉素、四环素、卡马西平和三氯生),浓度范围分别为24.5~38.7、47.5~49.2、43.1~85.4、2.5~3.9和423.2~8 973.3ng·L-1.两段A/O生物处理工艺对三氯生具有较好的去除效果,但对其余4种PPCPs的去除效果较差.与此同时,目标ERY-ARGs在污水处理厂中均被检出,在原水中的丰度达9.28×103(erm A)~1.83×108(ere A)copies·L-1,虽然两段A/O工艺能够显著降低污水中的ERY-ARGs(1.19 log~3.97 log),但其在最终处理出水中的浓度仍然较高.通过相关性分析可知,ERY-ARGs与红霉素和三氯生之间均具有显著相关性(P<0.05),表明污水处理厂中红霉素对ERY-ARGs的产生和传播具有重要影响,而三氯生对ERY-ARGs可能存在交叉选择性作用.     

2,3,7,8-Tetrachlorodibenzo-p-dioxin promotes migration ability of primary cultured rat astrocytes via aryl hydrocarbon receptor

Emerging evidence showed that 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) could induce expression of certain reactivation-associated genes in astrocytes, however, the consequent cellular effects and molecular mechanisms are still unclear. During the process of astrocyte reactivation, migration is a critical cellular event. In the present study, we employed wound-healing assay and Transwell® motility assay to explore the effects of TCDD on cell migration in primary cultured rat cortical astrocytes. We found that upon TCDD treatments at relative low concentrations (10^? 10 and/or 10^? 9?mol/L), the ability of primary astrocytes to migrate horizontally and vertically was promoted. In line with this cellular effect, the mRNA expression of two pro-migratory genes, including cell division cycle 42 (CDC42) and matrix metalloproteinase 2 (MMP2) was induced by TCDD treatment. Dioxin exerts its toxic effects mainly through aryl hydrocarbon receptor (AhR) pathway. So the role of AhR pathway in the pro-migratory effects of TCDD was examined using an AhR antagonist, CH223191. We found that application of CH223191 significantly reversed the pro-migratory effects of TCDD. Interestingly, the basal ability of horizontal migration as well as basal levels of CDC42 and MMP2 expression were dramatically reduced suggesting a possible physiological role of AhR in maintaining the endogenous migration ability of the primary astrocytes. These findings support the notion that dioxin promotes astrocyte reactivation at molecular and cellular levels.     

四环素对大肠杆菌抗生素抗性基因进化的影响

为了解环境中低浓度抗生素对抗生素抗性发展的影响,以大肠杆菌(Escherichia coli,E.coli)为基础材料,通过亚抑菌浓度四环素处理,得到由敏感到抗性的菌株,研究了四环素对大肠杆菌产生抗生素抗性基因(antibiotic resistance genes,ARGs)的影响,采用同位素标记相对和绝对定量技术(isobaric tags for relative and absolute quantification,iTRAQ),分析四环素处理前后,大肠杆菌总蛋白差异表达情况.结果表明,大肠杆菌经1/2MIC浓度的四环素诱导,20 d后可由敏感转至耐药,表明环境中的低浓度四环素的刺激对大肠杆菌耐药性的形成影响较大.并且,检测到四环素抗性基因tetA、tetE和tetZ,β-内酰胺类抗性基因blaTEM和blaFOX,多重耐药性基因acrA、marA和marR.蛋白结果表明大肠杆菌在四环素胁迫下以特定的节能存活模式,通过诱导细胞防御和修复系统来应对环境压力,并表现出对离子传输的严格调节,来抵抗四环素毒性的影响.研究结果为进一步解析环境中抗生素胁迫的抗性机制提供了参考.     

甲基汞神经毒性检测和评价效应指标--即刻早期基因研究

论述了甲基汞神经毒性研究现状，通过分析即刻早期基因（IEG）在甲基汞神经毒性机制中的作用，得出即刻早期基因能作为甲基汞神经毒性检测和评价的效应指标，同时也简要介绍即刻早期基因的研究方法，并指出利用基因芯片技术能够被步筛选出甲基汞对大鼠脑基因表达影响的相关基因。     

死菌DNA对厌氧消化污泥中抗生素抗性基因及微生物群落分析的干扰

为评估死菌DNA对厌氧消化污泥抗生素抗性基因(ARGs)和微生物群落分析的潜在干扰,本研究对3种不同类型厌氧消化污泥进行叠氮溴化丙锭(PMA)处理,比较在PMA屏蔽死菌DNA PCR扩增情况下污泥ARGs和微生物群落分析结果与未经PMA处理情况下的差异.结果表明,经PMA处理后,剩余污泥自厌氧消化样品和高含固厌氧消化污泥样品中的ARGs丰度分别下降了41%～86%和74%～98%;污泥水解液厌氧消化15 d后的污泥样品中ARGs下降幅度相对较小,但降幅最高也达到34%.PMA处理对3个来源不同的厌氧消化污泥微生物群落组成分析结果呈现不同程度的影响,对高含固厌氧消化污泥的微生物群落结构分析影响最为显著.在经PMA处理与未经PMA处理两种情况下,厌氧消化污泥ARGs与微生物群落组成相关性分析的结果也截然不同.研究证明了死菌DNA对厌氧消化污泥ARGs和微生物群落分析的潜在干扰,采用PMA预处理能够更准确地反映厌氧消化污泥中的微生物群落及菌体携带ARGs的特征.     

四环素胁迫对Shigella flexneri细菌四环素抗性基因抗性表达的影响过程

四环素(TC)抗生素在不同环境介质中已被广泛检出,为研究其对四环素抗性基因(TC-ARGs)丰度变化及表达水平的影响过程,以从活性污泥中筛选和纯化分离获得的弗氏志贺氏菌(Shigella flexneri)为研究对象,考察了不同浓度TC对其生长过程的作用影响,采用荧光定量PCR和逆转录PCR方法定量检测了不同抗性机制TC-ARGs,包括tetC、tetO和tetX基因的丰度变化及表达水平,并探讨了TC浓度与TC-ARGs丰度及其表达水平之间的相关关系.结果表明,在培养周期内(24 h),TC胁迫对Shigella flexneri细菌的生长具有抑制作用,细菌细胞浓度增长速率随TC暴露浓度的升高而降低,但对TC-ARGs丰度变化影响较小.TC胁迫能够促进Shigella flexneri细菌TC-ARGs的转录表达,tetC、tetO和tetX基因表达水平在整个培养周期内均先升高后降低.由相关性分析可知,TC浓度与TC-ARGs丰度及其表达水平之间相关关系不显著,但tetC和tetO基因丰度与其转录表达水平之间存在显著的正相关关系,表明其基因丰度一定程度上可用来衡量和评价其抗性表达水平.     

A screening assay for thyroid hormone signaling disruption based on thyroid hormone-response gene expression analysis in the frog Pelophylax nigromaculatus

Amphibian metamorphosis provides a wonderful model to study the thyroid hormone (TH) signaling disrupting activity of environmental chemicals, with Xenopus laevis as the most commonly used species. This study aimed to establish a rapid and sensitive screening assay based on TH-response gene expression analysis using Pelophylax nigromaculatus, a native frog species distributed widely in East Asia, especially in China. To achieve this, five candidate TH-response genes that were sensitive to T3 induction were chosen as molecular markers, and T3 induction was determined as 0.2 nmol/L T3 exposure for 48 hr. The developed assay can detect the agonistic activity of T3 with a lowest observed effective concentration of 0.001 nmol/L and EC50 at around 0.118–1.229 nmol/L, exhibiting comparable or higher sensitivity than previously reported assays. We further validated the efficiency of the developed assay by detecting the TH signaling disrupting activity of tetrabromobisphenol A (TBBPA), a known TH signaling disruptor. In accordance with previous reports, we found a weak TH agonistic activity for TBBPA in the absence of T3, whereas a TH antagonistic activity was found for TBBPA at higher concentrations in the presence of T3, showing that the P. nigromaculatus assay is effective for detecting TH signaling disrupting activity. Importantly, we observed non-monotonic dose-dependent disrupting activity of TBBPA in the presence of T3, which is difficult to detect with in vitro reporter gene assays. Overall, the developed P. nigromaculatus assay can be used to screen TH signaling disrupting activity of environmental chemicals with high sensitivity.     

石油污染土壤的生物修复技术及微生物生态效应

利用投菌法和生物刺激法对陕北子长石油污染土壤进行微生物修复研究.通过利用红外分光光度法测定不同处理方法对石油烃的去除效果确定了修复陕北石油污染土壤的最佳方案.修复过程中利用最大可能计数法(MPN)、PCR-琼脂糖电泳法、PCR-DGGE法分别测定了石油烃降解菌数目、催化基因、土壤微生物多样性对土壤微生物生态效应进行研究.结果发现石油污染土壤不同生物处理修复效果为:生物刺激(加入N、P营养物质)生物强化(投加降解菌)其他.土壤中石油烃降解率与可降解石油烃的催化基因含量之间存在正相关关系,修复过程中土壤中的石油烃和烷烃降解菌数量显著多于多环芳烃降解菌数量,投加外源降解菌SZ-1可以显著提高土壤细菌群落的多样性.研究结果有助于深入理解生物修复石油土壤过程中的微生物生态效应变化.     

Piriformospora indica confers cadmium tolerance in Nicotiana tabacum

Piriformospora indica, a root-colonizing endophytic fungus of Sebacinales, promotes plant growth and confers resistance against biotic and abiotic stresses. In order to confirm the influence of P. indica on growth, proline, malondialdehyde (MDA), chlorophyll, and cadmium (Cd) amounts in Nicotiana tabacum under Cd stress, hydroponics, pot and field trials were conducted. The results showed that P. indica can store Cd in plant roots and reduce leaf Cd content, reduce the concentration of MDA, and increase the proline and chlorophyll content and the activities of catalase, peroxidase, and superoxide dismutase under hydroponic Cd stress. RT-PCR analysis showed that the relative expression level of genes Gsh2, TaPCS1, oas1, GPX, and Hsp70 in colonized plants was 4.3, 1.4, 2.9, 1.7, and 6.9 fold higher than in un-colonized plants respectively. Cd exposure significantly reduced un-colonized plants'' agronomic traits compared to P. indica-colonized ones. Our results suggested that P. indica can sequester Cd in roots, so that much less cadmium was transported to leaves, and the increased concentrations of antioxidant enzymes, pigments and proline contents, as well as the higher expression of stress-related phytochelatin biosynthesis genes in P. indica-inoculated plants, may also serve to protect N. tabacum plants against oxidative damage, enhancing Cd tolerance.     

历史抗生素胁迫改变磺胺甲唑和甲氧苄啶对活性污泥的影响:ARGs及其潜在宿主

抗生素共暴露对污水处理厂抗生素抗性基因（ARGs）及微生物群落聚集过程有着重要影响.然而,历史抗生素接触胁迫差异（遗留效应）是否能决定微生物和ARGs对抗生素复合污染的响应尚不清楚.通过选择高浓度（30 mg ·L^-1）磺胺甲唑（SMX）和甲氧苄啶（TMP）作为历史接触胁迫条件,探究SMX和TMP复合污染对ARGs、细菌群落及其交互作用的短期影响.基于高通量荧光定量PCR,共检测出13种ARGs,它们的绝对丰度介于2.21~5.42 copies ·μL^-1（对数值,以DNA计,下同）,其中sul2、ermB、mefA和tetM-01是样品中最主要的亚型,绝对丰度在2.95~5.40 copies ·μL^-1之间;SMX和TMP复合污染会引起ARGs和可移动基因元件（MGEs）的富集,但其对各亚型的影响不同,且SMX的历史遗留效应高于TMP;不同接触史的复合污染下,ARGs间的共现与互斥模式均存在;MGEs （尤其是intI-1）与磺胺类（sul1、sul2）、四环素类［tet （32）］和大环内酯-林可酰胺-链霉菌素类抗性基因（ermB）均呈显著正相关性;基于微生物全尺度分类发现各类群微生物群落结构对复合污染的响应不同,且条件丰富菌属（CAT）得到了显著富集;陶厄氏菌属（Thauera）、假黄单胞菌属（Pseudoxanthomonas）和副球菌属（Paracoccus）是优势的抗性菌属;网络分析共发现31个ARGs的潜在宿主,以条件稀有菌属（CRT）为主,尤其是Candidatus_Alysiosphaera和纺锤状菌属（Fusibacter）与多种ARGs呈现正相关,是多种ARGs的潜在公共细菌宿主;部分稀有菌属［RT,变异杆菌属（Variibacter）、气单胞菌属（Aeromonas）和管道杆菌属（Cloacibacterium）等］是转座子IS 613的潜在宿主,在ARGs的增殖传播中发挥重要作用.综上,研究揭示了污水处理系统中抗生素历史胁迫对ARGs赋存特征和及其宿主细胞的遗留效应,可为削减抗生素复合污染引起的ARGs污染提供新思路与理论依据.