水厂饮用水肠道病毒工艺控制与污染防范

介绍了饮用水中肠道病毒的控制标准和指示物、工艺控制和防控措施。饮用水中的病毒以大肠杆菌噬菌体作为指示物,以EPA的病毒去除率为标准。从工艺方面可以采取控制滤后水浊度、提高消毒CT值、强化工艺组合等措施,同时水厂可从水源地、工艺废水和污泥、生产环境、接触人员等方面开展病毒污染防控。     

抗狂犬病病毒单链抗体的筛选及鉴定

目的:利用噬菌体展示技术构建的噬菌体抗体库,筛选、表达及鉴定具有体外中和活性的抗狂犬病病毒单链抗体。方法:以21份接受狂犬病病毒疫苗免疫的健康人外周血构建的抗狂犬病病毒scFv噬菌体免疫库为基础,免疫管包被纯化狂犬病病毒颗粒,通过三轮筛选,ELISA检测单克隆噬菌体抗体颗粒的阳性率,DNAStar和Vbase2对阳性克隆进行序列分析。序列正确的单链抗体构建原核表达质粒,转化大肠杆菌BL21(DE3)菌株,IPTG诱导表达单链抗体。经压力破碎、镍亲和层析对表达的蛋白进行纯化。RFFIT检测纯化蛋白对狂犬病病毒的体外中和活性。同时使用间接ELSIA检测获得的scFv对3aG,CVS,CTN及PV株的交叉反应性。结果:共获得40个阳性、基因测序结果正确的单链抗体。其中7个获得原核表达菌株,但只有3个获得较高的可溶性表达。RFFIT结果表明有4个scFv比活大于500 IU/mg。7个scFv对4种病毒株均有一定的交叉反应。结论:成功从该scFv噬菌体免疫库中筛选出特异性scFv;RFFIT结果表明获得的scFv具有体外中和狂犬病病毒的活性。     

噬菌体展示技术及其在病原微生物研究中的应用进展

噬菌体是能特异性地感染细菌的一类病毒,其基因数目少,结构相对简单,容易在分子水平上操控其基因。噬菌体展示技术是将外源肽或蛋白以融合蛋白形式表达并呈现在噬菌体衣壳表面的分子生物学技术。与传统的研究方法相比,噬菌体展示技术具有通量高、成本低、操作简单、耗时短等优点。本综述主要聚焦噬菌体展示技术在病原微生物抗原筛选、抗体制备和疫苗研制等方面的应用进展。     

浮游病毒的检测方法

浮游病毒包括噬藻体（Cyanophage,即蓝藻病毒）、藻病毒（Phycovirus,真核藻病毒）和噬菌体（Bacteriophage）等,它们在水体中大量存在,而且在控制水体的初级生产力和有机碳循环方面发挥着重要作用.     

高铁酸盐铁（Ⅵ）和铁（Ⅴ）：环保的氧化剂和消毒剂

病毒 研究高铁酸盐对病毒的破坏和去除效果已有报导。试验的病毒是肠细菌噬菌体f2和Qβ。结果表明在pH6～8的条件下，低浓度的铁（Ⅵ）能迅速灭活水与二次污水中的病毒。消毒过程没有遵循一级动力学。在pH值为6和7的磷酸盐缓冲溶液里，铁（Ⅵ）也可有效的灭活Qβ病毒。pH值下降，灭活率升高。灭活率可由ChickWatson定律表示（方程12）     

污水中包膜病毒离心超滤浓缩方法研究

针对污水中包膜病毒高效富集浓缩方法缺乏的问题，以Phi6噬菌体为模式包膜病毒，系统考察了超滤浓缩方法对污水样本中Phi6噬菌体的富集浓缩效果。结果表明，超滤筒的截留分子质量越小，对污水中Phi6噬菌体的回收率就越高，但所需浓缩时间也会增加。综合考虑回收率和耗时，推荐采用截留分子质量为10 ku的超滤筒，在4℃条件下3 000g正向离心28 min、1 000g反向收集5 min，浓缩倍数为110倍，回收率可达到（32±3.7）%。     

多孔建筑涂层抗病毒测试方法研究

多孔建筑涂层由于多孔性的特点对病毒具有吸附作用，从而赋予涂层抗病毒活性，而多孔涂层的抗病毒活性可能受测试中多种因素影响。选用多孔硅藻土制备多孔建筑涂层，以噬菌体MS2和φA039作为非包膜病毒模型测试多孔涂层的抗病毒活性，研究抗病毒测试中病毒储备液种类、病毒悬液滴加量及其浓度对抗病毒测试结果的影响。结果表明，应尽可能选择与稀释液有机组成相同的储备液；涂层表面悬液滴加量应视样品大小而定；涂层表面滴加病毒悬液浓度以1×107PFU/mL为宜。分析了多孔建筑涂层抗病毒测试中最佳测试条件，提出测试中需要注意的问题，为多孔涂层抗病毒测试和标准制定提供参考。     

水中f2噬菌体的浓集效果观察

水中ｆ２噬菌体的浓集效果观察水中肠道病毒对人类健康危害之大、影响之深，已是各国政府和专业工作者倍受关注的问题。但对水病毒的监控、检测，到目前为止国内外尚未有标准方法。有关浓集病毒的方法国内外报道的很多，士。滤膜法、盐沉淀法、透析法等，但均存在操作复杂...     

废水处理中的微型动物及其作用

一、引言 原生动物和后生动物是废水生物处理中常见的微生物,统称为微型动物,它在废水生物处理中起着很重要的作用,但一般文献资料只提及在吞食细菌净化水质和作为指示生物这两方面的作用,而忽略了其它作用。本文对废水生物处理中的微型动物及其各种作用,进行全面的探讨。 二、废水处理中的主要微型动物 1.活性污泥法中的微型动物 (1)原生动物 活性污泥在运行正常、处理水质较好的情况下,混合液中一般存活着5000     

一种水中诺如病毒富集方法的建立及应用

目的建立混合纤维素膜—聚乙二醇(PEG)沉淀法富集水中诺如病毒的方法并评价效果,以便应用于不同水源中诺如病毒的检测。方法采集乌鲁木齐市地区自来水、河水、污水样品15份,在样品中加入MS2噬菌体,通过混合纤维素膜—聚乙二醇(PEG)沉淀法富集;计算外加MS2回收率,并采用实时荧光RT-PCR方法检测样品中诺如病毒。结果自来水、河水和污水中MS2的平均回收率分别为4.8%,5.4%和9.7%;15份水样中未检出诺如病毒。结论本方法富集效果较好,可用于本地自来水、河水和污水中诺如病毒的检测,但仍需进一步优化以增强病毒的富集效果。     

Occurrence and persistence of enteroviruses, noroviruses and F-specific RNA phages in natural wastewater biofilms

Enteroviruses and noroviruses are pathogenic viruses excreted by infected individuals. Discharged in wastewaters, some of these viruses can be captured by biofilms. In the present study, we assessed the occurrence and persistence of these viruses in wastewaters and in corresponding biofilms. Natural wastewaters and biofilms were analyzed monthly from January to July using real-time RT-PCR. Enterovirus RNA was detected in wastewater in June while norovirus RNA was detected from January to March. In contrast, biofilm analysis revealed the presence of both enterovirus and norovirus genomes throughout the study period. For instance, enterovirus and norovirus genogroups (GG) I and II were detected in 50, 46 and 37% of the biofilm samples, respectively (n = 24). In a laboratory experiment, persistence of norovirus GGI RNA (quantified using molecular techniques) and F-specific bacteriophages (quantified using both culture and molecular techniques) was assessed in wastewater and corresponding naturally-contaminated biofilms at both 4 and 20 °C. The concentrations of viral genomes (norovirus GGI and F-specific RNA phage) were very stable in biofilms. Indeed, no significant decrease was observed during the persistence experiment that lasted 49 days. Furthermore, regardless of our experimental conditions, viral genome and infectious F-specific bacteriophages persisted longer in biofilm than in wastewater. According to our results, wastewater biofilms may contribute to the persistence and dispersal of pathogenic viruses outside of epidemic periods.     

Fluorescent dye labeled bacteriophages--a new tracer for the investigation of viral transport in porous media: 1. Introduction and characterization

A new method for the study of pathogen transport in porous media is presented. The method is based on conjugation of fluorescent dyes to target bacteriophages and application of the modified bacteriophages for tracer studies. We demonstrate that the relevant transport determining properties of Rhodamine and several fluorescein-labeled phages are practically identical to those of the native bacteriophages. The advantages of the proposed method relative to direct enumeration of bacteriophages by plaque forming unit method, turbidity, fluorescent microspheres, and other alternative tracers are discussed. Notable advantages include simple quantitation by optical methods, unbiased signals even when virus aggregates are formed, and the ability to decouple inactivation kinetics from transport phenomena. Additionally, the signal reflects the removal and transport of the studied microorganism and not a surrogate.     

Occurrence and levels of indicator bacteriophages in bathing waters throughout Europe

Somatic coliphages, F-specific RNA bacteriophages, bacteriophages infecting Bacteroides fragilis, Escherichia coli and enterococci were counted in bathing waters in the late spring and summer. We tested fresh and marine bathing waters from North, South, East and West Europe expected to contain between 100 and 500 E. coli per 100 ml, although wider ranges were sometimes found. Bacteriophages were counted after concentration, since a preliminary study proved that this step was necessary to obtain positive counts. During monitoring, a first-line quality control with reference materials for bacteria and bacteriophages was performed by all the laboratories participating in the study. The same microbes were also counted in raw sewage samples from various areas in Europe, where the bacterial indicators and the three groups of bacteriophages were detected in roughly the same numbers. All groups of bacteriophages were detected in both fresh and marine bathing waters throughout Europe. Reliable and complete results from 147 samples showed that for log-transformed values, E. coli and bacteriophages were slightly correlated. However, the slope of the regression line changed according to E. coli concentration and the correlation diminished when this concentration was close to zero per 100 ml. The ratios between E. coli and phages in bathing waters differed significantly from those in sewage. The relative amounts of bacteriophages, mainly somatic coliphages and phages infecting Bact. fragilis RYC2056, increased in bathing waters with low E. coli concentration, especially in seawater samples containing <100 E. coli per 100 ml. The relationship of bacteriophages with respect to enterococci paralleled that of bacteriophages with respect to E. coli. Somatic coliphages and bacteriophages infecting Bact. fragilis are useful to predict the presence of some pathogens with the same origin as present bacterial indicators but with higher survival rates.     

Inactivation of bacteriophages in water by means of non-ionizing (UV-253.7 nm) and ionizing (gamma) radiation: a comparative approach

Thc inactivation behaviour of the bacteriophages PHI X 174 (ssDNA virus). MS2 (ssRNA virus) and B40-8 (dsDNA) toward non-ionizing (UV-253.7 nm) as well as to ionizing radiation (gamma radiation) was studied in order to evaluate their potential as viral indicators for water disinfection by irradiation. Previous findings of the high UV-253.7 nm resistance of MS2 were confirmed whereas an unexpected high sensitivity to gamma radiation compared to the two other phages was found. On the other hand, PHI X 174 revealed an enhanced UV sensitivity but a high resistance to ionizing radiation. B40-8 had an intermediate position between the other two bacteriophages relative to both types of radiation. As expected, the data of E. coli reconfirmed the unreliability of fecal indicator bacteria for the purpose of predicting responses of viruses to water treatment. In UV disinfection the influence of water matrix may be adequately controlled by considering the UV (253.7 nm) absorption of the water whereas so far no such parameter has existed for the influence of the water quality on ionizing irradiation with respect to the scavenger concentration.     

Bacteriophages isolated from activated sludge processes and their polyvalency

In this study, bacteriophages were isolated from activated sludge and their host range was studied. Bacterial isolates were obtained from an activated sludge process treating urban sewage, and bacteriophages were obtained by plaque assay using the bacterial isolates obtained in this study as the host. Out of 15 bacteria isolated, 9 supported plaque formation. The host range test was conducted with a combination of 8 bacteriophage isolates and 9 bacterial isolates. All of the 8 bacteriophages tested were found to form plaques on more than 1 host, and 4 of them formed plaques on both gram-positive and gram-negative bacterial isolates. Three of the 8 bacteriophages failed to form plaques on their original bacterial host. The experimental result indicates that bacteriophages are an active part of the activated sludge microbial ecosystem, having a very close ecological relationship with their host bacteria.     

Differential persistence of F-specific RNA phage subgroups hinders their use as single tracers for faecal source tracking in surface water

The four subgroups of F-specific RNA bacteriophages (I-IV) have been proposed as potential tracers for faecal source tracking. Groups II and III predominate in human sources while groups I and IV are most abundant in animal sources. The four subgroups of naturally occurring F-specific RNA bacteriophages were identified in different samples by plaque hybridization with genotype-specific probes and the persistence of each subgroup was evaluated. The proportions of the F-specific RNA bacteriophage subgroups were measured in wastewaters, after inactivation in surface waters or after wastewater treatment and in mixtures of wastewater of human and animal origin. Our results indicate that phage groups differ in their persistence in the environment and to different disinfecting treatments. The greater survival of subgroups I and II in treated samples hinders the interpretation of results obtained with F-specific RNA bacteriophages. The phages of subgroups III and IV were the least resistant to all treatments. These results should be considered when using genotypes of F-specific RNA as sole tracers for faecal source tracking.     

Photoinactivation of virus on iron-oxide coated sand: enhancing inactivation in sunlit waters

Adsorption onto iron oxides can enhance the removal of waterborne viruses in constructed wetlands and soils. If reversible adsorption is not coupled with inactivation, however, infective viruses may be released when changes in solution conditions cause desorption. The goals of this study were to investigate the release of infective bacteriophages MS2 and ΦX174 (two human viral indicators) after adsorption onto an iron oxide coated sand (IOCS), and to promote viral inactivation by exploiting the photoreactive properties of the IOCS. The iron oxide coating greatly enhanced viral adsorption (adsorption densities up to ∼10⁹ infective viruses/g IOCS) onto the sand, but had no affect on infectivity. Viruses that were adsorbed onto IOCS under control conditions (pH 7.5, 10 mM Tris, 1250 μS/cm) were released into solution in an infective state with increases in pH and humic acid concentrations. The exposure of IOCS-adsorbed MS2 to sunlight irradiation caused significant inactivation via a photocatalytic mechanism in both buffered solutions and in wastewater samples (4.9 log₁₀ and 3.3 log₁₀ inactivation after 24-h exposure, respectively). Unlike MS2, ΦX174 inactivation was not enhanced by photocatalysis. In summary, IOCS enhanced the separation of viruses from the water column, and additionally provided a photocatalytic mechanism to promote inactivation of one of the surrogates studied. These qualities make it an attractive option for improving viral control strategies in constructed wetlands.     

Fates of bacteriophages and bacterial indicators in the Moselle river (France)

It has been suggested that bacteriophages can provide useful information about the pathogenic microorganisms, particularly enteric viruses, present in water. This information is complementary to that obtained from bacterial indicators of faecal contamination, which would be of great value for evaluating the risks associated with the use of certain types of water. Before bacteriophages can be used as indicators of faecal contamination, we need to confirm that bacteriophages give a different response to that given by the well-known bacteria indicators and to determine what happens to bacteriophages in river water. Indeed, drinking water is often produced from river water, either by natural filtration through the soil or after undergoing various treatments. We collected 96 river water samples from six different sites between February and November 2000. The samples were analysed for three faecal indicator bacteria (thermotolerant coliforms, enterococci and spores of sulphite-reducing anaerobes) and three types of bacteriophages (somatic coliphages, F-specific phages and Bacteroides fragilis phages). The densities of thermotolerant coliforms and enterococci depended mainly on physical factors such as flow rate and water temperature. High temperature and low flow rate led to a decrease in the density of these microorganisms, especially in the absence of a major input of faecal pollution. Conversely, the densities of somatic coliphages, F-specific phages and spores of sulphite-reducing anaerobes remained constant regardless of the flow rate and temperature. The density of Bacteroides fragilis phages was too low for unambiguous determination of their fate in river water.     

Microbial impact of small tributaries on water and shellfish quality in shallow coastal areas

This study aimed to investigate the impact of small tributaries on seawater and shellfish quality in coastal area subjected to brief episodes leading to fecal contamination. Escherichia coli and F-RNA-specific bacteriophages were selected as fecal indicators and astroviruses were chosen as being representative of pathogens in the human population during winter viral epidemics. A two-dimensional hydrodynamic model was built to simulate the current and dispersion in the model domain, which includes areas uncovered at low tide. The model also includes decay rates to simulate microorganism behavior and assess the influence of fecal input on shellfish quality. The originality lies in the fact that specific features of the study area were considered. Modeling results indicate limited particle movements and long flushing times at the back of the bay, where shellfish are farmed. Computational results showed that under normal conditions, i.e. 94% of the time, when rainfall was less than 10 mm per day, the sector shows acceptable water quality. These results are in agreement with shellfish concentration measured in the field. Under high flow conditions, high concentrations of fecal indicators and astrovirus were measured in the river and tributaries. The corresponding fluxes were over 50 times higher than under normal weather conditions. The location of the shellfish beds near the coast makes them vulnerable and fecal indicators and viruses were detected in shellfish after short rainfall events. Our modeling approach makes a contribution to shellfish management and consumer protection, by indicating the "risk period" as defined by EU regulations. Molecular development such as viral quantification in conjunction with model developments will help to prevent shellfish contamination and thus provide safer products to consumers and an effective tool for shellfish producers.     

Removal of MS2, Qβ and GA bacteriophages during drinking water treatment at pilot scale

The removal of MS2, Qβ and GA, F-specific RNA bacteriophages, potential surrogates for pathogenic waterborne viruses, was investigated during a conventional drinking water treatment at pilot scale by using river water, artificially and independently spiked with these bacteriophages. The objective of this work is to develop a standard system for assessing the effectiveness of drinking water plants with respect to the removal of MS2, Qβ and GA bacteriophages by a conventional pre-treatment process (coagulation-flocculation-settling-sand filtration) followed or not by an ultrafiltration (UF) membrane (complete treatment process). The specific performances of three UF membranes alone were assessed by using (i) pre-treated water and (ii) 0.1 mM sterile phosphate buffer solution (PBS), spiked with bacteriophages. These UF membranes tested in this work were designed for drinking water treatment market and were also selected for research purpose.The hypothesis serving as base for this study was that the interfacial properties for these three bacteriophages, in terms of electrostatic charge and the degree of hydrophobicity, could induce variations in the removal performances achieved by drinking water treatments.The comparison of the results showed a similar behaviour for both MS2 and Qβ surrogates whereas it was particularly atypical for the GA surrogate. The infectious character of MS2 and Qβ bacteriophages was mostly removed after clarification followed by sand filtration processes (more than a 4.8-log reduction) while genomic copies were removed at more than a 4.0-log after the complete treatment process. On the contrary, GA bacteriophage was only slightly removed by clarification followed by sand filtration, with less than1.7-log and 1.2-log reduction, respectively. After the complete treatment process achieved, GA bacteriophage was removed with less than 2.2-log and 1.6-log reduction, respectively.The effectiveness of the three UF membranes tested in terms of bacteriophages removal showed significant differences, especially for GA bacteriophage. These results could provide recommendations for drinking water suppliers in terms of selection criteria for membranes.MS2 bacteriophage is widely used as a surrogate for pathogenic waterborne viruses in Europe and the United States. In this study, the choice of MS2 bacteriophage as the best surrogate to be used for assessment of the effectiveness of drinking water treatment in removal of pathogenic waterborne viruses in worst conditions is clearly challenged. It was shown that GA bacteriophage is potentially a better surrogate as a worst case than MS2. Considering GA bacteriophage as the best surrogate in this study, a chlorine disinfection step could guaranteed a complete removal of this model and ensure the safety character of drinking water plants.