Removal of viable and inactivated Cryptosporidium by dual- and tri-media filtration

The limited efficacy of disinfectants, other than ultraviolet irradiation and ozonation, as a barrier against Cryptosporidium parvum in drinking water treatment has underscored the increased importance of oocyst removal by filtration. Currently, no reliable surrogates have been identified for C. parvum removal by filtration. As a result, evaluations of the Cryptosporidium removal by treatment operations have been performed using oocysts. It has typically been assumed that chemically inactivated oocysts are suitable surrogates for viable oocysts. Measurements of electrophoretic mobility, however, have shown that chemical inactivation changes the surface charge of Cryptosporidium oocysts. The present bench-scale research indicated that formalin-inactivated oocysts are reliable surrogates for viable oocysts during both stable filter operation and periods where filtration processes are challenged, such as coagulation failure. This finding is important because of the practical difficulties associated with using viable oocysts in filtration investigations. Poor coagulation conditions severely compromised removal of viable and inactivated oocysts by dual- and tri-media filters compared to stable operating conditions and filter ripening, emphasizing the importance of optimized chemical pre-treatment (coagulation) for the successful removal of oocysts during filtration. The treatment optimization experiments also indicated that tri-media filters offered only marginally higher oocyst removals than dual-media filters.     

Role of predation by zooplankton in transport and fate of protozoan (oo)cysts in granular activated carbon filtration

The significance of zooplankton in the transport and fate of pathogenic organisms in drinking water is poorly understood, although many hints of the role of predation in the persistence of microorganisms through water treatment processes can be found in literature. The objective of this study was to assess the impact of predation by natural zooplankton on the transport and fate of protozoan (oo)cysts in granular activated carbon (GAC) filtration process. UV-irradiated unlabelled Cryptosporidium parvum and Giardia lamblia (oo)cysts were seeded into two pilot-scale GAC filtration columns operated under full-scale conditions. In a two-week period after seeding, a reduction of free (oo)cysts retained in the filter bed was observed. Zooplankton was isolated from the filter bed and effluent water on a 30 μm net before and during the two-week period after seeding; it was enumerated and identified. Rotifers, which are potential predators of (oo)cysts, accounted for the major part of the isolated zooplankton. Analytical methods were developed to detect (oo)cysts internalized in natural zooplankton isolated from the filter bed and effluent water. Sample sonication was optimized to disrupt zooplankton organisms and release internalized microorganisms. (Oo)cysts released from zooplankton after sonication were isolated by IMS and stained (EasyStain™) for microscopic counting. Both Cryptosporidium and Giardia (oo)cysts were detected in association with zooplankton in the filter bed samples as well as in the effluent of GAC filters. The results of this study suggest that predation by zooplankton can play a role in the remobilization of persistent pathogens such as Cryptosporidium and Giardia (oo)cysts retained in GAC filter beds, and consequently in the transmission of these pathogens in drinking water.     

Genotyping of single Cryptosporidium oocysts in sewage by semi-nested PCR and direct sequencing

This study describes an approach for genotyping individual Cryptosporidium oocysts obtained from sewage. We isolated single immunofluorescent assay (IFA)-stained Cryptosporidium oocysts from sewage concentrate using glass capillary pipettes and inverted epifluorescence microscopy. Each isolated Cryptosporidium oocyst was analyzed by semi-nested PCR for the 18S rRNA gene and direct sequencing of the PCR products. A total of 74 of 107 oocysts isolated from sewage were genotyped successfully. Of the 74 genotyped isolates, 51% (38 oocysts) were identified as C. parvum genotype 1, 4% (3 oocysts) of C. parvum VF383 human isolates, 20% (15 oocysts) of C. parvum genotype 2, 14% (10 oocysts) of C. meleagridis, 7% (5 oocysts) of C. sp. Pig 1, 3% (2 oocysts) of C. sp PG1-26 pig isolates and 1% (1 oocyst) of C. parvum CPM1 isolated from mouse. The results of this study demonstrate that 18S rRNA-based semi-nested PCR and direct sequencing can be used to characterize individual Cryptosporidium oocysts and also to reveal the distribution of Cryptosporidium genotypes in environmental waters.     

Multi-scale Cryptosporidium/sand interactions in water treatment

Owing to its widespread occurrence in drinking water supplies and its significant resistance to environmental stresses, Cryptosporidium parvum is regarded as one of the most important waterborne microbial parasites. Accordingly, a substantial research effort has been aimed at elucidating the physical, chemical and biological factors controlling the transport and removal of Cryptosporidium oocysts in natural subsurface environments and drinking water treatment facilities. In this review, a multi-scale approach is taken to discuss the current state-of-knowledge on Cryptosporidium-sand interactions at a nano-scale, bench-scale and field-scale relevant to water treatment operations. Studies conducted at the nano-scale and bench-scale illustrate how techniques based on the principles of colloid and surface chemistry are providing new insights about oocyst-sand interactions during transport of Cryptosporidium oocysts in granular porous media. Specifically, atomic force microscopy and impinging jet experiments reveal the importance of oocyst surface biomolecules in controlling Cryptosporidium/sand interactions by a mechanism of steric hindrance. Traditional bench-scale column transport studies conducted over a broad range of experimental conditions highlight the role of physicochemical filtration and physical straining in the removal of oocysts from the pore fluid. Such experiments have also been used to evaluate the influence of biofilms formed on grain surfaces and the presence of natural organic matter on oocyst-sand interactions. Whilst filtration studies conducted at the plant-scale have been useful for evaluating the effectiveness of various materials as surrogates for Cryptosporidium oocysts, at this macro-scale, little could be learnt about the fundamental mechanisms controlling oocyst-sand interactions. This review of the literature on Cryptosporidium-sand interactions at different length scales points to the importance of combining studies at the plant-scale with well-controlled investigations conducted at the nano- and bench-scales. Furthermore, because oocyst surface properties play an important role in controlling the extent of interaction with sand surfaces, a thorough discussion of Cryptosporidium oocyst characteristics and electrical properties is presented.     

Effect of NOM and biofilm on the removal of Cryptosporidium parvum oocysts in rapid filters

Laboratory experiments were performed to evaluate the effects of biofilm and natural organic matter (NOM) on removal of Cryptosporidium parvum oocysts from water by filtration. The bench-scale rapid filters consisted of 2.54 cm ID x 30.5 cm polycarbonate plastic columns packed with 0.55 mm spherical glass beads to a depth of 25 cm and a porosity of 40%. Calcium chloride (0.01 M) served as the coagulant in most of the experiments. The oocyst removal efficiency decreased from 51 +/- 6% for a clean bed to 23 +/- 3% for the biofilm-coated bed and to 14 +/- 1% in the presence of 5 ppm of NOM. The oocyst removal for an experiment with a combination of biofilm-coated filter media and NOM was similar to that for the experiment with NOM alone (15 +/- 1%). The zeta potential values for the oocysts pre-equilibrated with NOM were significantly more negative than those obtained for untreated oocysts. This suggests that NOM enhanced the electrostatic repulsion between the oocysts and the negatively charged glass beads. Fortunately, use of alum as coagulant at a dosage sufficient to neutralize the surface charge of the NOM-coated oocysts resulted in a high removal efficiency (73 +/- 6%). Pre-equilibration of the oocysts with NOM also increased the hydrophobicity of the oocysts, but this was deemed to have a negligible effect on deposition onto the glass beads. The results of these experiments suggest that water treatment facilities treating source waters with moderate organic matter concentrations and/or employing biologically active filters have a greater potential for oocyst breakthrough and proper coagulation is critical for effective removal of oocysts in the filters.     

A stochastic model of an ozonation reactor

Disinfection of some microorganisms is characterized by a lag-phase (a minimum required ozone exposure until disinfection occurs). This phenomenon is easy to model in laboratory batch reactors but not in continuous flow mixed reactors. This paper introduces a stochastic disinfection model where individual microorganisms are followed on their paths through full-scale reactors. Combining exponentially distributed transport processes with delayed exponential disinfection kinetics for large populations of microorganisms (up to 10,000 individuals) yields predictions which can be evaluated statistically. It could be shown that deterministic models work well for systems with good disinfection performance (more than 2 log units reduction of active microorganisms), for reactors with poor performance stochastic models have to be applied. It could be demonstrated for real reactors that Bacillus subtilis spores are poor surrogates for Cryptosporidium parvum oocysts. The differences between the two microorganisms are large for reactors that deviate significantly from plug-flow behaviour.     

Inactivation and potential repair of Cryptosporidium parvum following low- and medium-pressure ultraviolet irradiation

This study investigated the level of inactivation and the potential for Cryptosporidium parvum to repair following low doses (1 and 3mJ/cm(2)) of ultraviolet (UV) irradiation from both low- and medium-pressure UV lamps. Cryptosporidium parvum oocysts suspended in phosphate buffered saline were exposed to UV using a bench-scale collimated beam apparatus. Oocyst suspensions were incubated at 5 degrees C or 25 degrees C under light and dark conditions up to 120 h (5 days) following exposure to UV irradiation, to examine photoreactivation and dark repair potential, respectively. Cryptosporidium parvum infectivity was determined throughout the incubation period using an HCT-8 cell culture and an antibody staining procedure for detection. No detectable evidence of repair was observed after incubation under light or dark conditions following either LP or MP UV lamp irradiation.     

Impact of bathers on levels of Cryptosporidium parvum oocysts and Giardia lamblia cysts in recreational beach waters

Recreational beach water samples collected on weekends and weekdays during 11 consecutive summer weeks were tested for potentially viable Cryptosporidium parvum oocysts and Giardia lamblia cysts using the multiplexed fluorescence in situ hybridization (FISH) method. The levels of oocysts and cysts on weekends were significantly higher than on the weekdays (P<0.01). Concentrations of oocysts in weekend samples (n=27) ranged from 2 to 42 oocysts/L (mean: 13.7 oocysts/L), and cyst concentration ranged from 0 to 33 cysts/L (mean: 9.1 cysts/L). For the samples collected on weekdays (n=33), the highest oocyst concentration was 7 oocysts/L (mean: 1.5 oocysts/L), and the highest cyst concentration was 4 cysts/L (mean: 0.6 cysts/L). The values of water turbidity were significantly higher on weekends than on weekdays, and were correlated with the number of bathers and concentration of C. parvum oocysts and G. lamblia cysts (P<0.04). The study demonstrated positive relationships between number of bathers and levels of waterborne C. parvum oocysts and G. lamblia cysts in recreational beach water. It is essential to test recreational waters for Cryptosporidium and Giardia when numbers of bathers are greatest, or limit the number of bathers in a recreational beach area.     

Detection of enteric viruses, Giardia and Cryptosporidium in two different types of drinking water treatment facilities

In this study, two types of drinking water treatment facilities (two conventional drinking water treatment plants (DWTPs) and two compact units (Cus)) were compared referring to their production capacity. Water samples were collected from three main points: (a) different water treatment steps (b) washings of sand filters and (c) distribution system at different distances from the water treatment plants. Both viruses and protozoa were concentrated from each water sample by adsorption and accumulation on the same nitrocellulose membrane filters (0.45 microm pore size). Enteroviruses were detected by plaque infectivity assay in BGM cells and HAV, HEV and Norovirus were detected by RT-PCR. Giardia and Cryptosporidium were detected by conventional staining methods and PCR. The results revealed that enterovirus load at the intake ranged between 10-15 PFU/L for the two compact units and between 4.5 and 75 PFU/L for the two conventional DWTPs. The virus load in distribution system of the first type DWTPs at 1 km from the plant was the same as that of the intake. Viruses in the other type of treatment plants CUs at 1, 5 and 7 km, were much reduced. Investigation of raw water sediments of the two DWTPs showed enterovirus counts between 12 and 17.5 PFU/L. Virus count was reduced in sand of filters after washing. Giardia cysts were equally detected by microscopy and PCR in only intake samples of EL-Hawamdia CU (33.3%) and Meet Fares DWTP (50%). Cryptosporidium oocysts were equally detected by microscopy and PCR in intake samples of Abo EL-Nomros CU (100%), EL-Hawamdia CU (66.7%) and Fowa DWTP (50%). At Meet Fares DWTP three positive intake samples for Cryptosporidium were detected by PCR, compared with only two positive samples by microscopy. Giardia cysts and Cryptosporidium oocysts were detected in raw water sediment and sand of filters before washing. Only one sample from Meet Fares DWTP sand of filters after washing was positive for both Giardia and Cryptosporidium. It can be concluded that the poor microbial quality of the water may be due to improper operational skills and management of the various water treatment plants (especially at the two high capacity treatment plants).     

Development of a nested-PCR assay for the detection of Cryptosporidium parvum in finished water

A nested-PCR assay, incorporating an internal positive control, was developed for Cryptosporidium monitoring in finished water. This assay was capable of reproducibly detecting 8 oocysts in spiked-filtered water samples collected from 5 South Australian water treatment plants. The RT-PCR assay of Kaucner and Stinear (Appl. Environ. Microbiol. 64(5) (1998) 1743) was also evaluated for the detection of Cryptosporidium parvum. Initially, under our experimental conditions, a detection level of 27 oocysts was achieved for spiked reagent water samples. This level was improved to 5 oocysts by modification of the method. Untreated South Australian source waters concentrated by calcium carbonate flocculation were found to be highly inhibitory to the RT-PCR assay. Concentration of similar samples using Envirochek filters appeared to eliminate PCR inhibition. While both methods possessed similar sensitivities the nested-PCR assay was more reproducible, more cost effective, simpler to perform and could detect both viable and non-viable intact Cryptosporidium parvum oocysts, which is an important consideration for plant operators. These factors make the nested-PCR assay the method of choice for screening large numbers of potable water samples, where a reliable low level of detection is essential.     

Capture of water-borne colloids in granular beds using external electric fields: improving removal of Cryptosporidium parvum

Suboptimal coagulation in water treatment plants often results in reduced removal efficiency of Cryptosporidium parvum oocysts by several orders of magnitude (J. AWWA 94(6) (2002) 97, J. AWWA 93(12) (2001) 64). The effect of external electric field on removal of C. parvum oocysts in packed granular beds was studied experimentally. A cylindrical configuration of electrodes, with granular media in the annular space was used. A negative DC potential was applied to the central electrode. No coagulants or flocculants were used and filtration was performed with and without application of an electric field to obtain improvement in removal efficiency. Results indicate that removal of C. parvum increased from 10% to 70% due to application of field in fine sand media and from 30% to 96% in MAGCHEM media. All other test particles (Kaolin and polystyrene latex microspheres) used in the study also exhibited increased removal in the presence of an electric field. Single collector efficiencies were also computed using approximate trajectory analysis, modified to account for the applied external electric field. The results of these calculations were used to qualitatively explain the trends in the experimental observations.     

Biology, persistence and detection of Cryptosporidium parvum and Cryptosporidium hominis oocyst

Cryptosporidium parvum and Cryptosporidium hominis are obligate enteric protozoan parasites which infect the gastrointestinal tract of animals and humans. The mechanism(s) by which these parasites cause gastrointestinal distress in their hosts is not well understood. The risk of waterborne transmission of Cryptosporidium is a serious global issue in drinking water safety. Oocysts from these organisms are extremely robust, prevalent in source water supplies and capable of surviving in the environment for extended periods of time. Resistance to conventional water treatment by chlorination, lack of correlation with biological indicator microorganisms and the absence of adequate methods to detect the presence of infectious oocysts necessitates the development of consistent and effective means of parasite removal from the water supply. Additional research into improving water treatment and sewage treatment practices is needed, particularly in testing the efficiency of ozone in oocyst inactivation. Timely and efficient detection of infectious C. parvum and C. hominis oocysts in environmental samples requires the development of rapid and sensitive techniques for the concentration, purification and detection of these parasites. A major factor confounding proper detection remains the inability to adequately and efficiently concentrate oocysts from environmental samples, while limiting the presence of extraneous materials. Molecular-based techniques are the most promising methods for the sensitive and accurate detection of C. parvum and C. hominis. With the availability of numerous target sequences, RT-PCR will likely emerge as an important method to assess oocyst viability. In addition, a multiplex PCR for the simultaneous detection of C. parvum, C. hominis and other waterborne pathogens such as Giardia lamblia would greatly benefit the water industry and protect human health.     

Transport and fate of Cryptosporidium parvum oocysts in intermittent sand filters

The transport potential of Cryptosporidium parvum (C. parvum) through intermittent, unsaturated, sand filters used for water and wastewater treatment was investigated using a duplicated, 2³ factorial design experiment performed in bench-scale, sand columns. Sixteen columns (dia=15 cm, L=60 cm) were dosed eight times daily for up to 61 days with 65,000 C. parvum oocysts per liter at 15°C. The effects of water quality, media grain size, and hydraulic loading rates were examined. Effluent samples were tested for pH, turbidity, and oocyst content. C. parvum effluent concentrations were determined by staining oocysts on polycarbonate filters and enumerating using epifluorescent microscopy. At completion, the columns were dismantled and sand samples were taken at discrete depths within the columns. These samples were washed in a surfactant solution and the oocysts were enumerated using immunomagnetic separation techniques.The fine-grained sand columns (d₅₀=0.31 mm) effectively removed oocysts under the variety of conditions examined with low concentrations of oocysts infrequently detected in the effluent. Coarse-grained media columns (d₅₀=1.40 mm) yielded larger numbers of oocysts which were commonly observed in the effluent regardless of operating conditions. Factorial design analysis indicated that grain size was the variable which most affected the oocyst effluent concentrations in these intermittent filters. Loading rate had a significant effect when coarse-grained media was used and lesser effect with fine-grained media while the effect of feed composition was inconclusive. No correlations between turbidity, pH, and effluent oocyst concentrations were found. Pore-size calculations indicated that adequate space for oocyst transport existed in the filters. It was therefore concluded that processes other than physical straining mechanisms are mainly responsible for the removal of C. parvum oocysts from aqueous fluids in intermittent sand filters used under the conditions studied in this research.     

Filter backwash and start-up strategies for enhanced particulate removal

A significant consideration in forward planning for water treatment works design and operation concerns the effectiveness of a filtration plant in providing a barrier to particulates in the low micrometre size range, including Cryptosporidium oocysts. The performance of rapid gravity filtration plants is believed to be dependent on backwash and start-up regimes. It was the aim of this study to optimize direct sand filtration by identifying optimum filter backwash and start-up conditions which minimized the passage of particulates into the filtrate. The filter ripening period has long been identified as a cause for concern with respect to particulate passage into the filtrate; this work has shown that up to 40% of all particles that pass into supply during a 48 h run, do so in the first hour of operation. Optimum combined air water “collapse-pulsing” backwash durations were identified that reduced the number of 2–5 μm particles entering the filtrate, especially during the ripening period. Slow start-up was also found to reduce the number of 2–5 μm particles in the filtrate during the ripening period. The reductions in particulate passage resulting from a slow start was found to be media dependent, with smaller media requiring a longer slow start duration than coarser media.     

Prevalence of Cryptosporidium oocysts and giardia cysts in the drinking water supply in Japan

A one-year monitoring of Cryptosporidium oocysts and Giardia cysts was conducted at a water purification plant. A total of 13 samples of 50 L river source water and 26 samples of 2,000 L-filtered water, treated by coagulation flocculation, sedimentation and rapid filtration, were tested. Prior to conducting a survey of a water purification plant, we developed a method for concentrating Cryptosporidium oocysts from a large volume of raw or filtered water using a hollow fiber ultrafiltration (UF) membrane, and this procedure was adapted to survey a water purification plant. Cryptosporidium oocysts were detected in all of the 13 raw water samples. The geometric mean concentration was 40 oocysts 100 L. Giardia cysts were detected in 12 of 13 raw water samples (92%) and the geometric mean concentration was 17 cysts/100 L. Probability distributions of both Cryptosporidium oocyst and Giardia cyst concentration in raw water were nearly lognormal. In filtered water samples, Cryptosporidium oocysts were detected in 9 of the 26 samples (35%) with the geometric mean concentration of 1.2 oocysts /1,000 L and Giardia cysts in 3 samples (12%) with 0.8 cysts/1,000 L. The estimated log10 removal efficiency of Cryptosporidium oocysts and Giardia cysts by rapid-sand filtration was 2.47 and 2.53, respectively. Empty particles were removed at a higher log10 than intact oocysts and cysts. The efficiency of particle removal in the rapid sand filtration process tends to be reduced under cold-water conditions. Close management is necessary in the winter when the water temperature is low.     

Settling velocity of Cryptosporidium parvum and Giardia lamblia

Understanding transport behavior of Cryptosporidium parvum oocysts and Giardia lamblia cysts (together referred to as (oo)cysts) in overland flow is important for beneficial uses of receiving water bodies. Like sediment, (oo)cysts are subjected to deposition once they are present in overland flow or low flow environments like reservoirs, wetlands and sedimentation basins. The objectives of this paper are to present the theory and experiment to determine the free settling velocity (v(s)) of (oo)cysts and to compare experimental settling velocities to estimates using Stokes' law. A settling experiment was designed to quantify the v(s) of (oo)cysts in an aqueous column. C. parvum oocysts used were spherical with average diameter (+/-1SD) of 6.6+/-1.1 microm. G. lamblia cysts were oval shaped (average eccentricity = 1.48+/-0.19) with average size of 11.8 +/-1.3 microm. Average densities were 1009 kg m(-3) for C. parvum oocysts and 1013 kg m(-3) for G. lamblia cysts. Observed experimental settling velocities are 0.27 microm s(-1) and 0.67 microm s(-1) for C. parvum and G. lamblia, respectively. Estimated average settling velocities using Stokes' law were 0.36 microm s(-1) for C. parvum and 0.84 microm s(-1) for G. lamblia. R-squared values of the observations from the settling experiments with the Stokes' law estimation are 0.87 and 0.88 for G. lamblia and C. parvum, respectively. Our results suggest that Stokes' law can be used to estimate settling velocities of (oo)cysts. Qualitatively, the low settling velocities indicate that (oo)cysts will very slowly settle out of suspension.     

Surface plasmon resonance-based inhibition assay for real-time detection of Cryptosporidium parvum oocyst

A surface plasmon resonance (SPR)-based inhibition assay method using a polyclonal anti-mouse IgM arrayed Cryptosporidium sensor chip was developed for the real-time detection of Cryptosporidium parvum oocysts. The Cryptosporidium sensor chip was fabricated by subsequent immobilization of streptavidin and polyclonal anti-mouse IgM (secondary antibody) onto heterogeneous self-assembled monolayers (SAMs). The assay consisted of the immunoreaction step between monoclonal anti-C. parvum oocyst (primary antibody) and oocysts, followed by the binding step of the unbound primary antibody onto the secondary antibody surface. It enhanced not only the immunoreaction yield of the oocysts by batch reaction but also the accessibility of analytes to the chip surface by antibody-antibody interaction. Furthermore, the use of optimum concentration of the primary antibody maximized its binding response on the chip. An inversely linear calibration curve for the oocyst concentration versus SPR signal was obtained in the range of 1x10(6)-1x10(2)oocystsml(-1). The oocyst detection was also successfully achieved in natural water systems. These results indicate that the SPR-based inhibition assay using the Cryptosporidium sensor chip has high application potential for the real-time analysis of C. parvum oocyst in laboratory and field water monitoring.     

Transport and fate of Cryptosporidium parvum oocysts in intermittent sand filters.

The transport potential of Cryptosporidium parvum (C. parvum) through intermittent, unsaturated, sand filters used for water and wastewater treatment was investigated using a duplicated, 2³ factorial design experiment performed in bench-scale, sand columns. Sixteen columns (dia=15 cm, L=60 cm) were dosed eight times daily for up to 61 days with 65,000 C. parvum oocysts per liter at 15°C. The effects of water quality, media grain size, and hydraulic loading rates were examined. Effluent samples were tested for pH, turbidity, and oocyst content. C. parvum effluent concentrations were determined by staining oocysts on polycarbonate filters and enumerating using epifluorescent microscopy. At completion, the columns were dismantled and sand samples were taken at discrete depths within the columns. These samples were washed in a surfactant solution and the oocysts were enumerated using immunomagnetic separation techniques.

The fine-grained sand columns (d₅₀=0.31 mm) effectively removed oocysts under the variety of conditions examined with low concentrations of oocysts infrequently detected in the effluent. Coarse-grained media columns (d₅₀=1.40 mm) yielded larger numbers of oocysts which were commonly observed in the effluent regardless of operating conditions. Factorial design analysis indicated that grain size was the variable which most affected the oocyst effluent concentrations in these intermittent filters. Loading rate had a significant effect when coarse-grained media was used and lesser effect with fine-grained media while the effect of feed composition was inconclusive. No correlations between turbidity, pH, and effluent oocyst concentrations were found. Pore-size calculations indicated that adequate space for oocyst transport existed in the filters. It was therefore concluded that processes other than physical straining mechanisms are mainly responsible for the removal of C. parvum oocysts from aqueous fluids in intermittent sand filters used under the conditions studied in this research.     

Contribution of treated wastewater to the contamination of recreational river areas with Cryptosporidium spp. and Giardia duodenalis

Samples of the influent and final effluent from 12 wastewater treatment plants from Galicia (NW, Spain) were analyzed for the presence of Cryptosporidium spp. oocysts and Giardia duodenalis cysts. All of the plants discharge effluent to a hydrographic basin in which there are numerous recreational areas and fluvial beaches. The samples (25-50 liters) were collected in spring, summer, autumn and winter of 2007. A total of 96 samples were analyzed using techniques included in the US Environmental Protection Agency Method 1623. To identify the genotypes present, the following genes were amplified and sequenced: 18S SSU rRNA (Cryptosporidium spp.) and beta-giardina (G. duodenalis). Both parasites were detected in influent and effluent samples from all treatment plants (100%) throughout the year, and G. duodenalis always outnumbered Cryptosporidium spp. The mean concentration of G. duodenalis per liter of influent was significantly higher (P<0.05) than the mean concentration of Cryptosporidium spp. per liter of influent. The mean concentrations of parasites in influent samples ranged from 6 to 350 Cryptosporidium spp. oocysts per liter and from 89 to 8305 G. duodenalis cysts per liter. In final treated effluent, the mean concentration of parasites ranged from 2 to 390 Cryptosporidium spp. oocysts per liter and from 79 to 2469 G. duodenalis cysts per liter. The distribution of results per season revealed that in all plants, the highest number of (oo)cysts were detected in spring and summer. Cryptosporidium parvum, Cryptosporidium andersoni, Cryptosporidium hominis and assemblages A-I, A-II, E of G. duodenalis were detected. The risk of contamination of water courses by Cryptosporidium spp. and G. duodenalis is therefore considerable. It is important that wastewater treatment authorities reconsider the relevance of the levels of contamination by both parasites in wastewater, and develop adequate countermeasures.     

Filtration behaviors of Giardia and Cryptosporidium--ionic strength and pH effects

The laboratory-scale filtration tests of Giardia cysts and Cryptosporidium oocysts in both 2 mm-phi glass beads and 2 mm-phi polystyrene beads filters were conducted to investigate their filtration behaviors. The protozoan parasites were used as target particles, while the chemical system altered by changing the electrolyte concentration and pH. The results significantly indicate that ionic strength have a positive effect on the removal efficiencies for Giardia cysts and Cryptosporidium oocysts. The removal efficiency of two filters for Giardia cysts slightly decreased from pH 2.4 to 8.7 and decreased significantly in pH as pH up to 8.7, while that for Cryptosporidium slightly rippled beyond pH 8.7, and with the decrease in pH up to pH 8.7. The experimental collision efficiencies from the interactions between colloids and the filter media were calculated with a semi-empirical approach of the single sphere model and clean-bed filtration theory. The results also indicated that experimental collision efficiencies for (oo)cysts corresponded to the (oo)cysts removal efficiencies in all trials, and oocysts exhibits higher collision efficiencies than cysts.