Biofilms containing pathogenic organisms from the water supply are a potential source of protozoan parasite outbreaks and a significant public health concern. The aim of the present study was to demonstrate the simultaneous and multi-spatial occurrence of waterborne protozoan pathogens (WBPP) in substrate-associated biofilms (SAB) and compare it to surface water (SW) and sediments with bottom water (BW) counterparts using manual filtration and elution from low-volume samples. For scenario purposes, simulated environmental biofilm contamination was created from in-situ grown one-month-old SAB (OM-SAB) that were spiked with Cryptosporidium parvum oocysts. Samples were collected from the largest freshwater reservoirs in Luzon, Philippines and a University Lake in Thailand. A total of 69 samples (23 SAB, 23 SW, and 23 BW) were evaluated using traditional staining techniques for Cryptosporidium, and Immunofluorescence staining for the simultaneous detection of Cryptosporidium and Giardia. WBPP were found in 43% SAB, 39% SW, and 39% BW of the samples tested in the present study with SAB results reflecting SW and BW results. Further highlights were demonstrated in the potential of using low-volume samples for the detection of parasites in source water. Scanning electron microscopy of OM-SAB samples revealed a naturally-associated testate amoeba shell, while Cryptosporidium oocysts spiked samples provided a visual profile of what can be expected from naturally contaminated biofilms. This study provides the first evidence for the simultaneous and multi-spatial occurrence of waterborne protozoan pathogens in low-volume aquatic matrices and further warrants SAB testing along with SW and BW matrices for improved water quality assessment strategies (iWQAS).

Cryptosporidium and Giardia are important parasites due to their zoonotic potential and impact on human health, often causing waterborne outbreaks of disease. Detection of (oo)cysts in water matrices is challenging and few countries have legislated water monitoring for their presence. The aim of this study was to investigate the presence and origin of these parasites in different water sources in Northern Greece and identify interactions between biotic/abiotic factors in order to develop risk-assessment models. During a 2-year period, using a longitudinal, repeated sampling approach, 12 locations in 4 rivers, irrigation canals, and a water production company, were monitored for Cryptosporidium and Giardia, using standard methods. Furthermore, 254 faecal samples from animals were collected from 15 cattle and 12 sheep farms located near the water sampling points and screened for both parasites, in order to estimate their potential contribution to water contamination. River water samples were frequently contaminated with Cryptosporidium (47.1%) and Giardia (66.2%), with higher contamination rates during winter and spring. During a 5-month period, (oo)cysts were detected in drinking-water (<1/litre). Animals on all farms were infected by both parasites, with 16.7% of calves and 17.2% of lambs excreting Cryptosporidium oocysts and 41.3% of calves and 43.1% of lambs excreting Giardia cysts. The most prevalent species identified in both water and animal samples were C. parvum and G. duodenalis assemblage AII. The presence of G. duodenalis assemblage AII in drinking water and C. parvum IIaA15G2R1 in surface water highlights the potential risk of waterborne infection. No correlation was found between (oo)cyst counts and faecal-indicator bacteria. Machine-learning models that can predict contamination intensity with Cryptosporidium (75% accuracy) and Giardia (69% accuracy), combining biological, physicochemical and meteorological factors, were developed. Although these prediction accuracies may be insufficient for public health purposes, they could be useful for augmenting and informing risk-based sampling plans.

Giardia is a protozoan parasite of primary concern for the drinking water industry. High contact times are required for Giardia inactivation by chlorination, while ozonation may be effective at much lower Ct products. In this study, we have assessed the occurrence of Giardia cysts in raw water, and in chlorinated or ozonated water from a drinking water treatment plant (DWTP) in Brazil, over a 16-month period. Moreover, we analyzed the effects of primary disinfection on cysts, and calculated the infection risk caused by the occurrence of Giardia cysts in raw water, chlorinated or ozonated water. Furthermore, we assessed the correlation of Giardia cysts with indicator bacteria in raw water. Data referring to concentration of Giardia cysts in raw water showed adherence to a gamma distribution at a significance level α = 0.05. The detection frequency and the mean concentration of Giardia cysts were higher in raw water (86.6%, 26 cysts∙L⁻¹), than in chlorinated (46.1%, 15.7 cysts·L⁻¹) or ozonated water (43.5%, 11.1 cysts·L⁻¹). Overall, Giardia non-viable cysts were detected more frequently in ozonated water (80%) than in chlorinated water (68.2%) or raw water (37.7%). Ozonation and chlorination resulted, respectively, in ≈27.5- and ≈13- fold reduction of Giardia infection risk, when compared to the risk calculated for raw water. Total coliform and Escherichia coli proved to be suitable surrogates to predict the occurrence of Giardia cysts in raw surface water, however, the indicator bacteria may not be suitable surrogates to predict the disinfection of Giardia cysts, as no correlation was found between indicator bacteria and Giardia cysts in treated water. To our knowledge, this is the first study reporting the efficacy of chlorine and ozone at Ct products actually applied at a full-scale drinking water treatment plant against Giardia cysts naturally occurring in the source water, i.e. real situation. Ozonation has proven more efficient than chlorination against Giardia cysts in surface water. Escherichia coli proved to be suitable surrogate to predict Giardia cysts in raw surface water.

Land use/Land cover (LULC) associated with Cryptosporidium sp. and Giardia sp. quantification and distribution can provide identification of the environmental circulation patterns of these parasites. The aim of this research was to relate the occurrence and circulation of these parasites to the LULC watershed with poor sanitation infrastructure and livestock as important economic activity. The study involved 11 municipalities in the state of São Paulo, located in southeastern Brazil. Sampling was carried out at the catchment sites of each water supply on a monthly basis, starting in December 2014 and lasting until November 2015, totalizing 128 samples. Protozoans were quantified according to the 1623.1 US. EPA Method. For watershed delimitation, the hydrographic network was extracted from the hydrology tool of ArcGIS 10.1. The frequency of occurrence of these pathogens and the high concentrations were evidenced in the municipality with the largest urban area (16.2%) and intense livestock activity (39%) near the catchment site. The municipality that showed the lowest frequency of occurrence presented the smallest urban area (0.87%) and absence of livestock activity near the catchment site. The high concentration of pathogens suggests a correlation between the impact on water supply networks and river basin degradation caused by urban activity and livestock.

The Tula Valley receives untreated wastewater from Mexico City for agricultural irrigation, half of which infiltrates to aquifers from where drinking water is extracted. Samples of wastewater and infiltrated water from three areas of the valley were analyzed for microorganisms, organic micropollutants, and some basic parameters. Concentrations of microorganisms in the infiltrated water were generally very low but the incidence of fecal coliforms (present in 68% of samples), somatic bacteriophages (36%), Giardia spp. (14%), and helminth eggs (8%) suggested a health risk. Organic micropollutants, often present at high concentrations in the wastewater, were generally absent from the infiltrated water except carbamazepine which was in 55% of samples (up to 193 ng/L). There was no correlation between carbamazepine concentrations and the presence of microorganisms but highest concentrations of carbamazepine and boron coincided. A treatment such as nanofiltration would be necessary for the infiltrated water to be a safe potable supply.

Sewage is a major contributor to pollution problems involving human pathogens in tropical coastal areas. This study investigated the occurrence of intestinal protozoan parasites (Giardia and Cryptosporidium) in tropical recreational marine waters contaminated with sewage. The potential risks of Cryptosporidium and Giardia infection from recreational water exposure were estimated from the levels of viable (oo) cysts (DIC+, DAPI+, PI−) found in near-shore swimming areas using an exponential dose response model. A Monte Carlo uncertainty analysis was performed in order to determine the probability distribution of risks. Microbial indicators of recreational water quality (enterococci, Clostridium perfringens) and genetic markers of sewage pollution (human-specific Bacteroidales marker [HF183] and Clostridium coccoides) were simultaneously evaluated in order to estimate the extent of water quality deterioration associated with human wastes. The study revealed the potential risk of parasite infections via primary contact with tropical marine waters contaminated with sewage; higher risk estimates for Giardia than for Cryptosporidium were found. Mean risks estimated by Monte Carlo were below the U.S. EPA upper bound on recreational risk of 0.036 for cryptosporidiosis and giardiasis for both children and adults. However, 95th percentile estimates for giardiasis for children exceeded the 0.036 level. Environmental surveillance of microbial pathogens is crucial in order to control and eradicate the effects that increasing anthropogenic impacts have on marine ecosystems and human health.

Water is a vital resource to every living thing on the earth. Once the water is contaminated (physically, chemically, biologically, or radiologically), it brought negative impacts to the living thing. This paper provides a brief review of the characterization of biological pollutants in drinking water and their effects on human health. Some biological contamination was detected in water resources such as pathogenic bacteria (Escherichia coli, Vibrio cholerae, Salmonella, etc.), viruses (hepatitis A virus, hepatitis E virus, rotavirus, etc.), parasites (Giardia, Entamoeba, Cyclospora, etc.), and parasitic worm (Ascaris lumbricoides, Ancylostoma duodenale, Strongyloides stercoralis, etc.). The diseases were significantly prevalent in developing countries due to limited access to clean water and poor sanitation. Most of the diseases had common symptoms such as diarrhea, fever, and body and muscle aches that were transmitted to humans through the fecal–oral route. About 1.7 billion children were affected by diarrhea each year and about 525,000 of the children died each year. Besides, nearly 1 million adults were killed by diarrhea every year. Some treatment was implemented to remove the biological contamination in drinking water, such as oxidation treatment, ultraviolet radiation, distillation, biologically active carbon filtration, electrochemical, and nanotechnology.

A quantitative microbial risk assessment method can be used to evaluate infections probabilities for microorganisms in a specific place. The methodology provides suitable information to generate strategies focusing on health problems. Giardia cysts (GC) and Cryptosporidium oocysts (CO) are considered emerging pathogens that can infect human and animals by ingesting contaminated food or water, where food and water are transport vehicles for these parasites. Studies for GC and CO have reported occurrences for these parasites in water up to 100%, and some of these studies documented a number of cases, about 403,000 people, infected worldwide. This review is focused on compiling the most relevant works assessing the risk for GC and CO and their presence in different water samples that are susceptible for direct and indirect human consumption. The annual risk infection probability for these parasites has been reported from different water sources, with a range between 1 × 10⁻⁶ and 1, while the world standard regulation is 1 × 10⁻⁴. The infection probability depends not only on water quality but also on water treatment implementations.

Knowledge gaps in the exposure parameters for recreational water activities make quantitative risk assessment related to water recreation difficult. Therefore, the annual exposure frequency and single exposure duration for the recreational water activities of residents from ten cities in the North and South of China were investigated. Questionnaire interviews were carried on recreational water activities comprising swimming (SW), boating (BA), playing in interactive fountains (PF), and watching fountains (WF). Quantitative microbial risk assessment for the exposure of urban residents to Cryptosporidium and Giardia was also performed. For the four recreational water activities, the participation rates of urban residents in SW and WF were higher than the others. For SW and BA, the mean annual exposure frequency and single exposure duration for males were significantly higher than those for females. PF and WF showed the opposite. The annual exposure frequency for above 35-year-old residents was higher than that for young residents (18–35 years). However, the single exposure duration for young residents was highest in SW, BA, and PF. The mean annual exposure frequency and single exposure duration for North China residents were higher than those for South China residents in all recreational water activities, except for SW. Overall, the annual exposure frequency and single exposure duration in recreational water activities for all urban residents followed a lognormal distribution. In the four recreational water activities, the total annual infection risk of male exposure to Cryptosporidium was 1.0 × 10⁻², with the confidence intervals between 95 and 5% of [4.3 × 10⁻⁴, 3.7 × 10⁻²], whereas that for females was 6.8 × 10⁻³ and [4.2 × 10⁻⁴, 2.4 × 10⁻²]. Also, the annual infection risk of males to Giardia was 8.8 × 10⁻³ and [5.1×10⁻⁴, 3.2×10⁻²], and that of females was 5.3 × 10⁻³ and [4.0 × 10⁻⁴, 1.8 × 10⁻²]. These results demonstrated that SW and PF made the highest contribution to the total annual infection risk. Sensitivity analysis highlighted that the characterization of exposure parameters plays a critical role in health risk assessment, which may provide a scientific basis for recreational water quality standards formulation.

Treatments using ozone for filter backwash water (FBW) and calcium oxide for floated residue (FR) were evaluated adopting bench-scale testing for the inactivation of Giardia spp. cysts and Cryptosporidium parvum oocysts. The protocol chosen for protozoa detection involved following the concentration step by direct centrifugation (adding ICN 7X cleaning solution at 1.0%) and purification by immunomagnetic separation (IMS). The FR treatment with calcium oxide (dosage of 23 mg CaO 100 mL⁻¹ and 3-day contact time at 25 °C) proved to be efficient, as no parasites were detected after the treatment. The reduction of calcium oxide dosage (16 mg CaO 100 mL⁻¹ and 3-day contact time at 25 °C) was insufficient to inactivate the protozoa, since potentially viable organisms were identified using propidium iodide (PI). Concerning the disinfection conditions with ozone (5-min and 10-min contact time and dosage of 10 mg O₃ L⁻¹ and 7.5 mg O₃ L⁻¹, respectively), there was complete removal of the target organisms, as no protozoa were detected after the FBW treatment. From the results obtained, the tested treatments can be considered promising alternatives for water treatment plants (WTPs). However, the costs incurred from these treatments have to be considered.