Determining potential indicators of Cryptosporidium oocysts throughout the wastewater treatment process

Most research on wastewater treatment efficiency compliance focuses on physicochemical and microbial indicators; however, very little emphasis has been placed so far on determining suitable indicator organisms to predict the discharge level of pathogens from treatment plants. In this study, raw wastewater, activated sludge, and the resulting final effluents and biosolids in four municipal wastewater treatment plants (WWTPs A, B, C and D) were seasonally investigated for human-virulent water-borne pathogens Cryptosporidium parvum/hominis and Giardia duodenalis, and microsporidia (e.g. Encephalitozoon hellem, E. intestinalis, and Enterocytozoon bieneusi) between 2008 and 2009. A suite of potential microbial indicators for human-virulent protozoa and microsporidia was also determined. A combination of multiple fluorescent in situ hybridization and immunofluorescent antibody assays were applied to detect Cryptosporidium oocysts, Giardia cysts, and microsporidian spores. Escherichia coli, enterococci and Clostridium perfringens spores were cultivated in selective media. Positive correlations were found between the abundance of enterococci and E. coli and abundance of Cryptosporidium oocysts (r(s) > 0.47, p < 0.01) and Giardia cysts (r(s) > 0.44, p < 0.01) at WWTPs A-D. Cryptosporidium perfringens spores were positively correlated to Cryptosporidium oocysts (r(s) = 0.40, p < 0.01) and Giardia cysts (r(s) = 0.46, p < 0.01). There was a strong positive correlation between abundance of Giardia cysts and that of Cryptosporidium oocysts (r(s) > 0.89, p < 0.01). To sum up, a suite of faecal indicator bacteria can be used as indicators for the presence of Cryptosporidium oocysts and Giardia cysts in these activated-sludge systems (WWTPs A, B and C). Overall, Giardia duodenalis was noted to be the best Cryptosporidium indicator for human health in the community-based influent wastewater and throughout the treatment process.     

上海市浦东给水厂两虫去除研究

采用EPA1623法(免疫磁分离及荧光染色法)对上海市几家典型的给水厂的水源水、出厂水、管道水中贾第鞭毛虫和隐孢子虫的分布状况进行了测定分析,并对给水厂净水过程中不同操作单元对两虫的去除效果进行了研究.研究表明:上海浦东给水厂水源水中存在贾第鞭毛虫与隐孢子虫,但其密度分别维持在0～6个/10 L和0～8个/10 L的低水平,而出厂水与管道水中均未检出两虫;给水厂净水过程中,混凝沉淀对两虫具有明显的去除效果.     

Inactivation of enteric microbes in water by electro-chemical oxidant from brine (NaCl) and free chlorine.

Oxidant solutions of mostly free chlorine can be electrochemically produced on-site from brine (NaCl) solution and used to disinfect water at the household or community level. In this study electrochemical oxidant (ECO) from brine and free chlorine were evaluated under laboratory conditions for inactivation of test microbes. Purified suspensions of Escherichia coli, the rugose strain of Vibrio cholerae, Clostridium perfringens spores, MS2 coliphage and Cryptosporidium parvum oocysts were treated with 2 mg/L or 5 mg/L solutions of ECO or free chlorine at 5 degrees C and 25 degrees C and pH 6, 8, and 10 (pH 7 and 25 degrees C only for C. parvum oocysts) for contact times <60 min. Under nearly all conditions, inactivation kinetics were more rapid for E. coli, V. cholerae, C. perfringens spores and MS2 coliphage with ECO than with free chlorine. ECO reduced E. coli, V. cholerae and MS2 by >4 log10 within 30 min and C. perfringens spores by >2 log10 within 10 min at pH 8 and 25 degrees C. Contrary to previous results, however, C. parvum oocysts were not inactivated by ECO, and the reasons for this difference are uncertain. The on-site electrolytic generation of oxidants from brine provided a convenient and inexpensive disinfectant containing free chlorine that was effective against many enteric microbes, for the treatment of household and community drinking-water supplies worldwide. However, the effectiveness of such oxidants for inactivating C. parvum oocysts was variable and sometimes ineffective.     

Genotyping of single Cryptosporidium oocysts isolated from sewage and river water.

The study was designed to genotype individual Cryptosporidium oocysts using an 18S rRNA gene-based semi-nested PCR and direct sequencing procedure. Positive PCR amplification was observed in all single C. parvum HNJ-1 oocyst samples tested. Semi-nested PCR and direct sequencing was applied to Cryptosporidium oocysts isolated from sewage and river water. The procedure could genotype 54% of FITC-stained single oocysts isolated from sewage and 32% from river water. The predominant genotype in both sewage and river water was C. parvum genotype 1, accounting for 33 and 25%, respectively, of all the FITC-stained intact Cryptosporidium oocysts present.     

Quantification and genotyping of Cryptosporidium spp. in river water by quenching probe PCR and denaturing gradient gel electrophoresis.

A new detection method was developed for the simultaneous quantification and genotyping of Cryptosporidium spp. in river water. Several modifications made to the US EPA Method 1623 enabled high and stable recovery of Cryptosporidium from 40 L of river water (geometric mean = 35%, standard deviation = 8.7%). Quenching probe PCR (QProbe PCR) was used to quantify the 18S rRNA gene of Cryptosporidium spp. This method could successfully detect single oocysts in a sample, and the lower quantitation limit was as low as 2.5 oocysts/sample. In addition, denaturing gradient gel electrophoresis (DGGE) followed by DNA sequencing was used to identify the genotypes. These methods were applied to detect Cryptosporidium spp. in the Koyama River, Japan. The positive ratio was 69% (11/16) with the maximum concentration of 59 oocysts/100 L. Seven genotypes including two novel ones were identified. These results showed that this detection method could provide valuable information on Cryptosporidium in river water, both in the concentration and in the genotypes, which is essential for the precise assessment of waterborne risk to human health.     

Evidence for the existence of Cryptosporidium oocysts as single entities in surface runoff.

There is uncertainty whether Cryptosporidium oocysts attach to particles or to each other under ambient water conditions. Particle size distributions of Cryptosporidium oocyst suspensions were determined over a range of ionic strengths and pHs to determine under those environmental conditions that may promote oocyst aggregation. Cryptosporidium oocysts were shown to only aggregate in high ionic strength solutions (>0.45 M) and remain largely as single entities at ionic strengths and pHs that were likely to be encountered in surface runoff. Similarly, in loam soil suspensions, rather than attaching to the soil particles the majority of oocysts also remained as single entities. Overall, oocysts are expected to remain largely unattached to either themselves or soil particles in overland runoff. This has implications for pathogen transport and modelling since oocysts that are freely suspended are more likely to be transported in runoff to surface waters than if attached to more dense soil/faecal particles.     

Elimination of viruses, bacteria and protozoan oocysts by slow sand filtration.

The decimal elimination capacity (DEC) of slow sand filters (SSF) for viruses, bacteria and oocysts of Cryptosporidium has been assessed from full-scale data and pilot plant and laboratory experiments. DEC for viruses calculated from experimental data with MS2-bacteriophages in the pilot plant filters was 1.5-2 log10. E. coli and thermotolerant coliforms (Coli44) were removed at full-scale and in the pilot plant with 2-3 log10. At full-scale, Campylobacter bacteria removal was 1 log10 more than removal of Coli44, which indicated that Coli44 was a conservative surrogate for these pathogenic bacteria. Laboratory experiments with sand columns showed 2-3 and >5-6 log10 removal of spiked spores of sulphite-reducing clostridia (SSRC; C. perfringens) and oocysts of Cryptosporidium respectively. Consequently, SSRC was not a good surrogate to quantify oocyst removal by SSF. Removal of indigenous SSRC by full-scale filters was less efficient than observed in the laboratory columns, probably due to continuous loading of these filter beds with spores, accumulation and retarded transport. It remains to be investigated if this also applies to oocyst removal by SSF. The results additionally showed that the schmutzdecke and accumulation of (in)organic charged compounds in the sand increased the elimination of microorganisms. Removal of the schmutzdecke reduced DEC for bacteria by +/-2 log10, but did not affect removal of phages. This clearly indicated that, besides biological activity, both straining and adsorption were important removal mechanisms in the filter bed for microorganisms larger than viruses.     

粒状活性炭在管道分质供水口感改善中的作用研究

口感是评价管道分质供水质量的一项关键指标,它直接影响消费者对管道分质供水的接受程度。研究了影响管道分质供水口感的主要因素及粒状活性炭对口感改善所起的作用。结果表明,净水中余氯浓度、TOC含量及VOCs含量均会对净水口感产生影响;粒状活性炭过滤可有效降低管道分质供水中净水余氯浓度、TOC与VOCS含量,明显改善净水口感,而对其他水质指标无不良影响。结合试验结果与工程实例,提出了应用粒状活性炭改善管道分质供水口感的技术参数,并建议采用粒状活性炭过滤器作为管道分质供水的终端过滤。     

Using ultraviolet light for disinfection of finished water.

Ultraviolet light is now recognised to be very effective for inactivation of Cryptosporidium parvum oocysts; however, its application for disinfection of finished water necessitates validation of UV reactors prior to their installation. Although reactor performance will likely be assessed using non-pathogenic microorganisms as biodosimetry surrogates, it would be prudent for the water industry to simultaneously measure Cryptosporidium oocysts inactivation in controlled bench-scale studies using the water matrix intended for disinfection. The likelihood of that occurring is dependent upon the availability of infectivity measurement procedures that are more user-friendly than the mouse infectivity assays currently used. This study describes a modified cell culture procedure that would enable reliable measurement of changes in oocysts' infectivity following their UV treatment. Also, a number of different biodosimetry surrogates were examined and one selected for comparing the UV doses delivered between bench-scale and full-scale biodosimetry studies. Impacts of UV disinfection on production of disinfection byproducts, effects of lamp ageing on effectiveness of disinfection and the costs associated with employing this technology were also examined.     

Evaluating ozone dose for AOC removal in two-step GAC filters

Upgrading an existing post-ozonation plant with two-step granular activated carbon (GAC) filtration for assimilable organic carbon (AOC) removal was studied. The effects of ozone dose on AOC formation and its removal in the subsequent two-step GAC filtration was studied using chemically pretreated 2 to 14° C humic lake water. Two parallel pilot-plant trains with different ozone doses (0 to 1.2 mgO₃/mgTOC) and a shortterm ozonation study were performed. The optimum ozone dose for maximum AOC formation was 0.4–0.5 mgO₃/mgTOC. The AOC-P17 of ozonated water was three-fold higher and AOC-NOX over ten-fold higher than in non-ozonated water, while the following biofiltration (first step) removed 51 % and 72 % of AOC-P17 and AOC-NOX, respectively. The adsorber (second step) contributed to less than 10% of the overall AOC reduction. It appeared that biofiltration is a feasible method in upgrading water treatment plants for AOC removal even when treating cold humic waters, while the subsequent adsorber seems to have less significance for AOC removal.     

Evaluation of activated sludge treatment and the efficiency of the disinfection of Giardia species cysts and Cryptosporidium oocysts by UV at a sludge treatment plant in Campinas, south-east Brazil.

Among many waterborne diseases the giardiasis and cryptosporidiosis are of particular public health interest, because Giardia cysts and Cryptosporidium oocysts can persist for long periods in the environment, and both pathogenic protozoa have been implicated as the cause of many outbreaks of gastroenteritis in the last 25 years. In order to evaluate the efficiency of cysts and oocysts' removal by the activated sludge process, and by UV reactor in inactivating cysts and oocysts in one wastewater treatment plant (WWTP) of Campinas, three sampling points were selected for study: (1) influent, (2) treated effluent without UV disinfection and (3) treated effluent with UV disinfection. Giardia spp. cysts prevailed with higher density in the three different sample types. Cryptosporidium spp. oocysts were observed in only two samples of influent and just one sample of treated sewage with UV disinfection. In the animal infectivity assay for Giardia spp, one mouse of the UV treated group revealed trophozoites in intestinal scrapings. The results of the present study indicate that treatment by activated sludge process delivered a reduction of 98.9% of cysts and 99.7% of oocysts and UV disinfection was not completely efficient regarding the inactivation of Giardia cysts in the case of the WWTP studied.     

Evaluation of the purification capacity of nine portable, small-scale water purification devices.

A test was performed to evaluate the microbial and chemical purification capacity of nine portable, small-scale water purification filter devices with production capacity less than 100 L/h. The devices were tested for simultaneous removal capacity of bacteria (cultured Escherichia coli, Clostridium perfringens, Klebsiella pneumoniae and Enterobacter cloacae), enteric protozoans (formalin-stored Cryptosporidium parvum oocysts), viral markers (F-RNA bacteriophages) and microcystins produced by toxic cyanobacterial cultures. In general, the devices tested were able to remove bacterial contaminants by 3.6-6.9 log10 units from raw water. Those devices based only on filtration through pores 0.2-0.4 microm or larger failed in viral and chemical purification. Only one device, based on reverse osmosis, was capable of removing F-RNA phages at concentrations under the detection limit and microcystins by 2.5 log10. The present study emphasised the need for evaluation tests of water purification devices from the public safety and HACCP (Hazard Analysis and Critical Control Point) points of view. Simultaneous testing for various pathogenic/indicator microbes and microcystins was shown to be a useful and practical way to obtain essential data on actual purification capacity of commercial small-scale drinking-water filters.     

剩余臭氧影响活性炭净水效能原因分析

通过中试分析了长期运行条件下臭氧—活性炭工艺中溶解性臭氧对活性炭净水效能产生影响的原因。试验条件下活性炭对UV254的去除能力随剩余臭氧浓度的增加而下降。根据试验推测臭氧对活性炭的影响主要体现在3个方面:首先,在臭氧投加量较高时,NOM中亲水性有机物比例增加,此类物质在活性炭上的吸附性较差;其次,臭氧氧化降低了活性炭的吸附能力,同时臭氧可能与活性炭反应生成新的氧化产物;最后,臭氧可抑制活性炭上微生物的生长繁殖。因此建议在臭氧—活性炭工艺运行时严格控制进入活性炭吸附池的剩余臭氧浓度。     

Efficient taste and odour removal by water treatment plants around the Han River water supply system.

Seven major water treatment plants in Seoul Metropolitan Area, which are under Korea Water Resources Corporation (KOWACO)'s management, take water from the Paldang Reservoir in the Han River System for drinking water supply. There are taste and odour (T&O) problems in the finished water because the conventional treatment processes do not efficiently remove the T&O compounds. This study evaluated T&O removal by ozonation, granular activated carbon (GAC) treatment, powder activated carbon (PAC) and an advanced oxidation process in a pilot-scale treatment plant and bench-scale laboratory experiments. During T&O episodes, PAC alone was not adequate, but as a pretreatment together with GAC it could be a useful option. The optimal range of ozone dose was 1 to 2 mg/L at a contact time of 10 min. However, with ozone alone it was difficult to meet the T&O target of 3 TON and 15 ng/L of MIB or geosmin. The GAC adsorption capacity for DOC in the three GAC systems (F/A, GAC and O3 + GAC) at an EBCT of 14 min is mostly exhausted after 9 months. However, substantial TON removal continued for more than 2 years (>90,000 bed volumes). GAC was found to be effective for T&O control and the main removal mechanisms were adsorption capacity and biodegradation.     

Removal of geosmin and 2-methylisoborneol by biological filtration.

The quality of drinking water is sometimes diminished by the presence of certain compounds that can impart particular tastes or odours. One of the most common and problematic types of taste and odour is the earthy/musty odour produced by geosmin (trans-1, 10-dimethyl-trans-9-decalol) and MIB (2-methylisoborneol). Taste and odour treatment processes including powdered activated carbon, and oxidation using chlorine, chloramines, potassium permanganate, and sometimes even ozone are largely ineffective for reducing these compounds to below their odour threshold concentration levels. Ozonation followed by biological filtration, however, has the potential to provide effective treatment. Ozone provides partial removal of geosmin and MIB but also creates other compounds more amenable to biodegradation and potentially undesirable biological instability. Subsequent biofiltration can remove residual geosmin and MIB in addition to removing these other biodegradable compounds. Bench scale experiments were conducted using two parallel filter columns containing fresh and exhausted granular activated carbon (GAC) media and sand. Source water consisted of dechlorinated tap water to which geosmin and MIB were added, as well as, a cocktail of easily biodegradable organic matter (i.e. typical ozonation by-products) in order to simulate water that had been subjected to ozonation prior to filtration. Using fresh GAC, total removals of geosmin ranged from 76 to 100% and total MIB removals ranged from 47% to 100%. The exhausted GAC initially removed less geosmin and MIB but removals increased over time. Overall the results of these experiments are encouraging for the use of biofiltration following ozonation as a means of geosmin and MIB removal. These results provide important information with respect to the role biofilters play during their startup phase in the reduction of these particular compounds. In addition, the results demonstrate the potential biofilters have in responding to transient geosmin and MIB episodes.     

Control of Cryptosporidium: From reservoirs to clarifiers to filters

Control of Cryptosporidium in potable waters requires an integrated multiple barrier approach of source water protection of water supply reservoirs and treatment technology through clarification, filtration, and disinfection. Pristine lakes and protected reservoirs have far lower concentrations of oocysts (mean values less than 10 oocysts/100 L) than unprotected supplies. Coagulation is critical to the effective control of Cryptosporidium by clarification and filtration. Coagulation conditions that produce filtered waters of low turbidities, particle counts, and effectively remove natural organic matter also produce high removals of Cryptosporidium. Dissolved air flotation (DAF) can achieve oocyst removals of 3 log compared to about 1 log by sedimentation. DAF and filtration provide two effective barriers to Cryptosporidium with cumulative log removals of 4 to 5 compared to log removals of 3 to 4 by sedimentation and filtration.     

Occurrence and persistence of bacterial and viral faecal indicators in wastewater biofilms.

Biofilms within wastewater treatment plants can capture enteric microorganisms initially present in the water phase immobilising them either definitively or temporarily. Consequently, fates of microorganisms may totally change depending on whether they interact or not with biofilms. In this study, we assessed the stability of wastewater biofilms comparing the evolution of the concentrations of bacteria (heterotrophic plate count [HPC], thermotolerant coliforms [TC]) and viral (somatic coliphages [SC] and F-specific phages [F +]) indicators in the biofilms and in the corresponding wastewaters at 4 and 20 dgrees C. Additionally, we assessed the monthly occurrence of these bacterial and viral indicators as well as of pathogenic protozoa (Cryptosporidium oocysts and Giardia cysts) in three native wastewater biofilms for four months. Our results show that viral indicators (SC and F + ) persist longer in biofilms than in the corresponding wastewaters at 4 degrees C as well as at 20 degrees C. In contrast, persistence of bacterial indicators (TC and HPC) depends on both the temperature and the matrix. Differences between viral and bacterial persistence are discussed. Monthly analysis of native wastewater biofilms shows that bacterial and viral indicators, as well as Cryptosporidium oocysts and Giardia cysts, attach to wastewater biofilms to a concentration that remains stable in time, probably as a result of a dynamic equilibrium between attachment and detachment processes.     

Pathogen monitoring offers questionable protection against drinking-water risks: a QMRA (quantitative microbial risk analysis) approach to assess management strategies.

Risk mitigation provided by human monitoring and control over a water supply system has been consistently overlooked when estimating pathogen exposure to consumers. The Systems-Actions-Management (SAM) framework lends itself neatly to Quantitative Microbial Risk Assessment (QMRA) as one way to establish this link. The general premise is that an organisational protocol will influence how a human controller behaves, in turn influencing the system performance. For illustrative purposes, the framework was applied to a hypothetical water supply system to quantify the risk reduction offered by routine Cryptosporidium monitoring and the response to oocyst 'detects'. Our findings suggest that infrequent direct pathogen monitoring may provide a negligible risk barrier. The practice of sampling treated water to verify microbiological integrity is also dubious: oocyst densities were largely under-estimated, in part due to the spatial dispersion of oocysts in the waterbody, but predominantly from imperfect detection methods. The development of 'event-driven' monitoring schemes with barrier performance-based treatment verification methods, as promoted in new guidelines, is supported as a pressing issue to reduce the likelihood of undetected pathogen passage through a treatment plant.     

Biofiltration as pre-treatment to water harvesting and recycling

This paper presents the results of the long term biofilter experiments conducted with raw stormwater collected from a canal at Carlton, in Sydney. Anthracite and granular activated carbon (GAC) were used as a single filter media in biofilter columns. Media heights of 75 and 40 cm were used. The filter columns were operated at filtration velocities of 0.12 and 0.25 m/h. The removal efficiency for turbidity and DOC for the GAC filter media were found to be 75% and almost 100% respectively. The removal efficiency for the anthracite filter was much lower. Molecular weight distribution analysis showed an almost similar trend to the DOC removal. Compared with anthracite filter media, the GAC biofilter removed a much larger range of organic compounds present in the stormwater. The GAC biofilter removes organic matter earlier as compared to anthracite. Based on a limited sample of stormwater, the removal efficiency for phosphorus was upto 74% and that of nitrogen was up to 30%. In general GAC filter shows higher heavy metal removal efficiency than anthracite. The removal of zinc, iron, lead and nickel were good. However the concentration of heavy metal in the raw surface water sample was low.