Prevalence and genetic diversity of waterborne pathogenic viruses in surface waters of tropical urban catchments

Aims: To study the virological quality of surface water from highly urbanized tropical water catchment areas and to determine predominant enteric viral genotypes in surface water. Methods and Results: A wide range of human pathogenic viruses in urban surface waters was screened by nested PCR assays after concentration by ultrafiltration. Among the 84 water samples collected, at least one virus was detected in 70 (83·3%) of these samples. Noroviruses were determined to be the most prevalent enteric viruses detected in urban surface water samples, followed by astroviruses, enteroviruses, adenoviruses and hepatitis A viruses. The molecular characterization of environmental viral isolates suggested co‐circulation of multiple genotypes of both noroviruses GI and GII, astroviruses and enteroviruses in urban surface waters. Conclusions: Human enteric viruses with great genetic diversity were detected in surface waters, indicating the presence of human origin of faecal contamination in highly urbanized water catchment areas. Significance and Impact of the Study: The present study identifies and characterizes potential viral hazards of source waters for drinking water supply and recreational activities. This will enable scientific decisions to be made regarding the selection and prioritization of human pathogenic viruses to be included in the future risk assessment and treatment evaluation for water and wastewater.     

Sources and species of cryptosporidium oocysts in the Wachusett Reservoir watershed

Understanding the behavior of Cryptosporidium oocysts in the environment is critical to developing improved watershed management practices for protection of the public from waterborne cryptosporidiosis. Analytical methods of improved specificity and sensitivity are essential to this task. We developed a nested PCR-restriction fragment length polymorphism assay that allows detection of a single oocyst in environmental samples and differentiates the human pathogen Cryptosporidium parvum from other Cryptosporidium species. We tested our method on surface water and animal fecal samples from the Wachusett Reservoir watershed in central Massachusetts. We also directly compared results from our method with those from the immunofluorescence microscopy assay recommended in the Information Collection Rule. Our results suggest that immunofluorescence microscopy may not be a reliable indicator of public health risk for waterborne cryptosporidiosis. Molecular and environmental data identify both wildlife and dairy farms as sources of oocysts in the watershed, implicate times of cold water temperatures as high-risk periods for oocyst contamination of surface waters, and suggest that not all oocysts in the environment pose a threat to public health.     

Genotypes of Cryptosporidium from Sydney water catchment areas

AIMS: Currently cryptosporidiosis represents the major public health concern of water utilities in developed nations and increasingly, new species and genotypes of Cryptosporidium are being identified in which the infectivity for humans is not clear. The complicated epidemiology of Cryptosporidium and the fact that the majority of species and genotypes of Cryptosporidium cannot be distinguished morphologically makes the assessment of public health risk difficult if oocysts are detected in the raw water supplies. The aim of this study was to use molecular tools to identify sources of Cryptosporidium from the Warragamba catchment area of Sydney, Australia. METHODS AND RESULTS: Both faecal and water samples from the catchment area were collected and screened using immunomagnetic separation (IMS) and immunofluorescence microscopy. Samples that contained Cryptosporidium oocysts were genotyped using sequence and phylogenetic analysis of the 18S rDNA, and the heat-shock (HSP-70) gene. Analysis identified five Cryptosporidium species/genotypes including C. parvum (cattle genotype), C. suis, pig genotype II, the cervid genotype and a novel goat genotype. CONCLUSIONS: Monitoring and characterization of the sources of oocyst contamination in watersheds will aid in the development and implementation of the most appropriate watershed management policies to protect the public from the risks of waterborne Cryptosporidium. SIGNIFICANCE AND IMPACT OF THE STUDY: This study has shown that quantification by IMS analysis can be combined with the specificity of genotyping to provide an extremely valuable tool for assessing the human health risks from land use activities in drinking water catchments.     

Sources and Species of Cryptosporidium Oocysts in the Wachusett Reservoir Watershed

Understanding the behavior of Cryptosporidium oocysts in the environment is critical to developing improved watershed management practices for protection of the public from waterborne cryptosporidiosis. Analytical methods of improved specificity and sensitivity are essential to this task. We developed a nested PCR-restriction fragment length polymorphism assay that allows detection of a single oocyst in environmental samples and differentiates the human pathogen Cryptosporidium parvum from other Cryptosporidium species. We tested our method on surface water and animal fecal samples from the Wachusett Reservoir watershed in central Massachusetts. We also directly compared results from our method with those from the immunofluorescence microscopy assay recommended in the Information Collection Rule. Our results suggest that immunofluorescence microscopy may not be a reliable indicator of public health risk for waterborne cryptosporidiosis. Molecular and environmental data identify both wildlife and dairy farms as sources of oocysts in the watershed, implicate times of cold water temperatures as high-risk periods for oocyst contamination of surface waters, and suggest that not all oocysts in the environment pose a threat to public health.     

Nitrogen and oxygen isotopes in nitrate in the groundwater and surface water discharge from two rural catchments: implications for nitrogen loading to coastal waters

In Prince Edward Island, Canada, widespread intensive potato production has contributed to elevated nitrate concentrations in groundwater and streams, and eutrophic or anoxic conditions occur regularly in several estuarine systems. In this research, the stable isotopes of nitrogen and oxygen in nitrate in intertidal groundwater discharge and stream water were used, in conjunction with water quality and quantity data and land use information, to better understand the characteristics of nitrate delivered to two small estuaries with contrasting land use in their contributory catchments. Most of the water samples collected during the two-year study had isotopic signatures that fell in the range expected for nitrate derived from ammonium-based fertilizers (26.5 % of the samples) or in the overlapping range formed between ammonium-based fertilizers and nitrate derived from soil (64 % of the samples). Overall, isotopic signatures spanned over relatively narrow ranges, and correlations with other water quality parameters, or catchment characteristics, were weak. Nitrate in groundwater discharge and surface water in the Trout River catchment exhibited significantly different isotopic signatures only for the nitrogen isotope, while in the McIntyre Creek catchment groundwater discharge and surface water had similar isotopic signatures. When the isotopic results for the waters from the two catchments were compared, the surface waters were found to be similar, while the isotopic signatures of nitrate in groundwater were distinct only for the nitrogen isotope. Denitrification in the two study catchments was not evident based on the isotopic results for nitrate; however, in the case of the Trout River catchment, where a small freshwater pond exists, an average nitrate load reduction of 14 % was inferred based on a comparison of nitrate loads entering and leaving the pond. Overall, it appears that natural attenuation processes, occurring either in the streams or groundwater flow systems, do not significantly reduce nitrate loading to these estuaries.     

Mycobacterium avium subsp. paratuberculosis in lake catchments, in river water abstracted for domestic use, and in effluent from domestic sewage treatment works: diverse opportunities for environmental cycling and human exposure

Mycobacterium avium subsp. paratuberculosis from infected animals enters surface waters and rivers in runoff from contaminated pastures. We studied the River Tywi in South Wales, United Kingdom, whose catchment comprises 1,100 km2 containing more than a million dairy and beef cattle and more than 1.3 million sheep. The River Tywi is abstracted for the domestic water supply. Between August 2002 and April 2003, 48 of 70 (68.8%) twice-weekly river water samples tested positive by IS900 PCR. In river water, the organisms were associated with a suspended solid which was depleted by the water treatment process. Disposal of contaminated slurry back onto the land established a cycle of environmental persistence. A concentrate from 100 liters of finished water tested negative, but 1 of 54 domestic cold water tanks tested positive, indicating the potential for these pathogens to access domestic outlets. In the separate English Lake District region, with hills up to 980 m, tests for M. avium subsp. paratuberculosis in the high hill lakes and sediments were usually negative, but streams and sediments became positive lower down the catchment. Sediments from 9 of 10 major lakes receiving inflow from these catchments were positive, with sediment cores indicating deposition over at least 40 to 50 years. Two of 12 monthly 1-liter samples of effluent and a single 100-liter sample from the Ambleside sewage treatment works were positive for M. avium subsp. paratuberculosis. Since Lake Ambleside discharges into Lake Windermere, which is available for domestic supply, there is a potential for these organisms to cycle within human populations.     

Establishing relative release kinetics of faecal indicator organisms from different faecal matrices

Aims: A laboratory assay for comparative characterization of various faecal matrices with respect to faecal indicator organism (FIO) release using, artificial rain water. Methods and Results: Fresh sheep and beef‐cattle faeces, dairy cattle slurry and beef cattle farm yard manure (FYM) were collected from commercial units in south‐west England and applied to 20 randomized 1 m² plots established on permanent grassland. Representative samples from each faecal matrix (n = 5) were collected on four occasions over 16 days. One gram of each sample was transferred to a sterile vial to which 9 ml of standard local rain was carefully pipetted. The vial was then rotated through 360°, 20 times in 60 s to ‘simulate’ a standardized interaction of the faecal material with rainfall, providing an assay of comparative release potential. Appropriate decimal dilutions were prepared from the eluent. Following agitation, with a sterile spatula, the remaining faecal material and eluent in the vials were vortex mixed for 60 s before decimal dilutions were prepared from the resulting mixture, providing a quantitative assessment of the total FIO in the sample from which percentage release could be determined. Bacterial concentrations were enumerated in duplicate by membrane filtration following standard methods for FIO. Significant differences in release kinetics of Escherichia coli and enterococci from each of the faecal matrices were determined. Conclusions: Differences in release from each faecal substrate and between FIO type (E. coli and intestinal enterococci) were observed in this laboratory study. The order of release of E. coli from the faecal matrices (greatest to least, expressed as a percentage of the total present) was dairy cattle slurry > beef cattle FYM > beef‐cattle faeces > sheep faeces. For intestinal enterococci the order of percentage release was dairy cattle slurry > beef‐cattle faeces > beef cattle FYM > sheep faeces. Significance and Impact of the Study: This laboratory‐based method provides the first data on the relative release kinetics of FIO from different faecal matrices in rain water. This is fundamental information needed to parameterize laboratory‐based microbial models and inform approaches to field and catchment risk assessment.     

Pathogen Sources Estimation and Scenario Analysis Using the Soil and Water Assessment Tool (SWAT)

Concerns over water quality in Ireland have increased in recent years, in part due to the more frequent contamination of drinking water by pathogens such as Escherichia coli and Cryptosporidium. The objective of this study was to assess the use of SWAT for pathogen source estimation and to analyze the effects of various source scenarios on pathogen outputs in Irish catchments. Two agricultural catchments in Ireland susceptible to pathogen contamination of source water were the center of the SWAT model development with the primary focus on levels of E. coli in surface water. Model simulations used site and source specific information which was analyzed considering the total E. coli count for the simulation period (Fergus: January 2005–October 2006; Kilshanvey: January 2006–July 2007). Pathogen source estimation identified point sources as the most significant contributors to E. coli output with direct deposition the primary contributor (95%) in Kilshanvey and wastewater treatment plant outflow (89%) the main contributor in the Fergus catchment. A scenario analysis evaluated possible situations that may occur in study locations. The analysis indicated that restriction of livestock access to water sources and improved wastewater treatment would represent effective methods of improving water quality in both catchments.     

Mixed Giardia duodenalis assemblage A and E infections in calves

A molecular epidemiological study was conducted on 100 dairy (499 calves) and 50 beef (333 calves) farms in Belgium to estimate the prevalence of different Giardia duodenalis assemblages in calves younger than 10 weeks of age. Positive samples from the epidemiological study and from a previous clinical study were selected and genotyped based on the amplification of the beta-giardin gene. To investigate the occurence of mixed assemblage A and E infections in calves, a novel assemblage-specific PCR was developed based on the triose-phosphate isomerase gene. The prevalence was 22% (95% Probability Interval (PI): 12-34%) in dairy calves and 45% (95% PI: 30-64%) in beef calves. In total, 120 Giardia-positive samples from dairy and beef calves collected in the epidemiological study and from clinically affected calves were identified based on the amplification of the beta-giardin gene. Overall G. duodenalis assemblage E was more prevalent (in 64% of the samples), although the majority (59%) of the dairy calves were infected with G. duodenalis assemblage A. Furthermore, mixed G. duodenalis assemblage A and E infections were identified in 31% of the calf samples (n=101) using the assemblage-specific PCR. We believe this is the first report of mixed infections in calves, and the results of the present study indicate that calves, although mainly infected with the host-specific G. duodenalis assemblage E, are frequently infected with the zoonotic assemblage A, either as a mixed or mono-infection, suggesting that calves might be underestimated as a potential zoonotic reservoir for human infections.     

Group A Rotavirus as a Cause of Neonatal Calf Enteritis in Sweden

Faeces samples were collected during outbreaks of neonatal calf diarrhoea in 14 beef and dairy herds. Samples from 33 calves were taken at the onset of diarrhoea as well as from 30 calves with no signs of enteritis. No vaccines or medical treatment had previously been given. The mean age of the calves was 16.8 days (SD 8.2). The clinical evaluation of faeces consistency corresponded well to the dry matter content of the faeces (p<0.001). The samples were analyzed for rotavirus, Cryptosporidium species and Escherichia coli K99+. Group A rotavirus was detected by ELISA and RNA Polyacrylamide gel electrophoresis (RNA-PAGE) in 14/33 (43.8%) of the samples from scouring calves and 1/30 (3.7%) of the samples from non-scouring calves. The correlation between group A rotavirus and diarrhoea was statistically significant (p<0.001). No non-group A rotaviruses were found by RNA-PAGE. Cryptosporidium species were detected through demonstration of oocysts in smears from 12/63 (19.0%) of the faecal samples, but no statistically significant correlation between diarrhoea and detection of oocysts was demonstrated. Escherichia coli K99+ was not detected in any faeces sample. The clear association between group A rotavirus and diarrhoea is suggested to be due to low pathogenic load in the herds.     

Protection of calves against cryptosporiosis by oral inoculation with gamma-irradiated Cryptosporidium parvum oocysts

The purpose of this study was to determine whether gamma-irradiated Cryptosporidium parvum oocysts could elicit protective immunity against cryptosporidiosis in dairy calves. Cryptosporidium parvum Iowa strain oocysts (1 x 10(6) per inoculation) were exposed to various levels of gamma irradiation (350-500 Gy) and inoculated into 1-day-old dairy calves. The calves were examined daily for clinical signs of cryptosporidiosis, and fecal samples were processed for the presence of C. parvum oocysts. At 21 days of age, the calves were challenged by oral inoculation with 1 x 10(5) C. parvum oocysts and examined daily for oocyst shedding and clinical cryptosporidiosis. Calves that were inoculated with C. parvum oocysts exposed to 350-375 Gy shed C. parvum oocysts in feces. Higher irradiation doses (450 or 500 Gy) prevented oocyst development, but the calves remained susceptible to C. parvum challenge infection. Cryptosporidium parvum oocysts exposed to 400 Gy were incapable of any measurable development but retained the capacity to elicit a protective response against C. parvum challenge. These findings indicate that it may be possible to protect calves against cryptosporidiosis by inoculation with C. parvum oocysts exposed to 400-Gy gamma irradiation.     

A survey of the macro-invertebrate riffle fauna of the rivers Ystwyth and Rheidol, Wales

SUMMARY. A total of 155 taxa was collected from the Ystwyth (111) and Rheidol (134) catchments and total macro-invertebrate densities ranged from 435 to 9105 m⁻² and 155 to 4290 m⁻² in the two catchments, respectively. The fauna in each catchment was dominated by Insecta, principally Plecoptera, Ephemeroptera and Diptera. Trichoptera and Oligochaeta were proportionally more abundant in the Rheidol catchment. There was no evidence that the distribution or relative abundance of the faunas of the catchments were related simply to heavy metal contamination (mean zinc concentration up to 2.0 mg I⁻¹ and 0.34 mg I⁻¹ in the Ystwyth and Rheidol, respectively). Average linkage clusters of Spearman rank correlation coefficients suggested that community affinities were probably related principally to location within the catchment. There was no evidence in the Rheidol catchment that irregular discharges (up to c . 15 m³ s⁻¹) of water for hydroelectric purposes had any substantial effect on the invertebrate fauna although total invertebrate density below the discharge was significantly higher than at an upstream site.     

Mycobacterium avium subsp. paratuberculosis in Lake Catchments, in River Water Abstracted for Domestic Use, and in Effluent from Domestic Sewage Treatment Works: Diverse Opportunities for Environmental Cycling and Human Exposure

Mycobacterium avium subsp. paratuberculosis from infected animals enters surface waters and rivers in runoff from contaminated pastures. We studied the River Tywi in South Wales, United Kingdom, whose catchment comprises 1,100 km² containing more than a million dairy and beef cattle and more than 1.3 million sheep. The River Tywi is abstracted for the domestic water supply. Between August 2002 and April 2003, 48 of 70 (68.8%) twice-weekly river water samples tested positive by IS900 PCR. In river water, the organisms were associated with a suspended solid which was depleted by the water treatment process. Disposal of contaminated slurry back onto the land established a cycle of environmental persistence. A concentrate from 100 liters of finished water tested negative, but 1 of 54 domestic cold water tanks tested positive, indicating the potential for these pathogens to access domestic outlets. In the separate English Lake District region, with hills up to 980 m, tests for M. avium subsp. paratuberculosis in the high hill lakes and sediments were usually negative, but streams and sediments became positive lower down the catchment. Sediments from 9 of 10 major lakes receiving inflow from these catchments were positive, with sediment cores indicating deposition over at least 40 to 50 years. Two of 12 monthly 1-liter samples of effluent and a single 100-liter sample from the Ambleside sewage treatment works were positive for M. avium subsp. paratuberculosis. Since Lake Ambleside discharges into Lake Windermere, which is available for domestic supply, there is a potential for these organisms to cycle within human populations.     

Modeling of Pathogen Indicator Organisms in a Small-Scale Agricultural Catchment Using SWAT

Degradation of water quality from microbial contaminants associated with agricultural activities has significant implications for source protection of potable water. Novel environmental approaches must be adopted to assess risks from waterborne pathogenic microbes. The objective of this study was to evaluate applicability of the Soil and Water Assessment Tool (SWAT) to predict daily concentrations of E. coli in a small-scale agricultural catchment in Ireland. The study area is based on the Kilshanvey catchment located in the west of Ireland. E. coli data (n = 25) from June 2006 to June 2007 were utilized for comparison with the model's predictions. Statistical analysis indicates an unsatisfactory to fair level of correlation for the model's predictions (R² = 0.03–0.35, NSE = –0.42–0.29). A sensitivity analysis identified direct stream deposition and die-off rates for E. coli as having a significant impact on the model's predictions. Our results suggest that the model is adequate to assess the magnitude of various microbial sources within catchments but capability to replicate daily observations is uncertain. However, model outputs could provide adequate data to develop a human exposure assessment to pathogen indicator organisms in surface water and assist policy-makers in developing appropriate risk management strategies.     

Assessing the impact of the spatial arrangement of agricultural practices on pesticide runoff in small catchments: Combining hydrological modeling and supervised learning

Pesticide runoff is an important contributor to surface-water contamination. Transfer conditions at the field scale are well known, but our understanding is still quite limited in assessing water contamination by pesticides at the catchment scale. In this study, we focused on the spatial arrangement of agricultural practices in small catchments (<1 km²) and its impact on surface-water quality with respect to pesticide losses. An innovative method, the conjugation of mechanistic modeling and supervised-learning, was used to simplify this complex situation of pesticide transfer at the catchment scale.At the catchment level, the connectivity between each field and the hydraulic network is a major factor in determining the transfer of pesticides from fields to the outlet. We propose a connectivity indicator that assesses the contribution of each field to the hydraulic network runoff flow, which can be interpreted as the impact of each field on the catchment water quality. The connectivity indicator was inferred from the results of a mechanistic model simulation. First, we constructed a database from simulations using a proven hydrological model (MHYDAS) with defined scenarios. Then, two-thirds of the data were processed by the REPtree method to yield a regression tree. The rules obtained by this tree formed the basis for the connectivity indicator. The tree was then validated using the last third of the simulated data and the connectivity indicator was applied to a small wine-growing catchment in Roujan, France. This indicator required several types of data for its calculation: a spatial mapping of the studied watershed (topography, field characteristics, and network distribution) and an estimation of the spatial distribution of saturated hydraulic conductivity. The indicator developed here can be used to carry out risk assessments for diffuse pollution in diagnostics as well as test-mitigation strategies. The assessment results can be represented as a risk map to facilitate their interpretation.     

Water-borne Cryptosporidium: a perspective from the USA

In the USA, Cryptosporidium parvum has attracted considerable interest due to recent outbreaks of cryptosporidiosis attributed to water supplies contaminated with oocysts. Because of its unprecedented magnitude, the outbreak which occurred in Milwaukee in April 1993 was widely publicized and studied. Information collected during this and other recent outbreaks is being evaluated by regulatory agencies and the water industry with the aim of designing cost-effective measures that will reduce the risk of future water-borne outbreaks, as discussed here by Giovanni Widmer, Margaretha Carraway and Saul Tzipori.     

Comparison of Campylobacter jejuni PFGE and Penner subtypes in human infections and in water samples from the Taieri River catchment of New Zealand

AIMS: To determine the degree of overlap in strain types of Campylobacter jejuni isolated from clinical cases and water samples from the Taieri catchment in the South Island of New Zealand. METHODS AND RESULTS: Thermophilic Campylobacter were collected from human cases of infection, the main stem of the Taieri River and streams within distinct land-use types over a 1-year period. Campylobacter jejuni (187 isolates) and Campylobacter lari (four isolates) were identified using a multiplex polymerase chain reaction protocol. Isolates were typed by the Penner method and pulsed-field gel electrophoresis (PFGE) utilizing two restriction endonucleases. Several serotypes and PFGE types occurred in both water samples and clinical cases when the restriction profiles for each enzyme were considered separately. However, when PFGE profiles and serotyping were combined, there was no overlap between Camp. jejuni types from water and clinical cases. CONCLUSIONS: The results of this study indicate that recreational water in the Taieri catchment is not a major source of campylobacteriosis in the Dunedin area. SIGNIFICANCE AND IMPACT OF THE STUDY: This study suggests the risk of acquiring campylobacteriosis from surface waters in the Taieri catchment is considerably lower than previously predicted and highlights the necessity of using two endonucleases in PFGE typing.     

Changes in water colour between 1986 and 2006 in the headwaters of the River Nidd, Yorkshire, UK: a critique of methodological approaches and measurement of burning management

The effect of moorland management on the release of dissolved organic carbon (DOC) from blanket peat is currently a topic of great interest in the UK. A recent paper by Chapman et al. (Biogeochemistry, doi:10.1007/s10533-010-9474-x, 2010) reports on changes in humic colour/DOC concentrations in surface waters draining 15 upland peat catchments in the North Pennines (UK) over two decades, and examines the possible underlying drivers of those changes. Chapman et al. identify significant variation in water colour between adjacent catchments and over time. One potential driver of changes in DOC is managed moorland burning, and Chapman et al. state that their study provides evidence that burning has no effect on colour in upland catchment drainage waters. This observation counters a recent series of papers showing strong links between new moorland burn management on blanket peat soils and colour/DOC in catchment drainage waters. We have reviewed the methodological approach and results presented by Chapman et al. that relate to the assessment of managed burning, and show significant errors in the data used in their analysis. This has resulted in conclusions being drawn about the role of managed burning in DOC release that are not supported by evidence.     

鼎湖山地带性植被及其不同演替阶段水文学过程长期对比研究

分析鼎湖山3种植被类型生态系统水文的长期连续观测资料,采用时空互代的方法,得到如下一些结果:1)鼎湖山自然保护区东沟集水区产水量达到降水量的66.5%,日径流量高峰的出现相对降水的发生滞后1d左右。2)地下水位平均稳定在2.22m,最低为2.84m,最高为1.14m。1999、2000、2001、2002和2003年地下水位平均值分别为2.38,2.27,2.08,2.13和2.11m。鼎湖山东沟集水区每日地下水位与前16d每一天的降水量相关。3)随着时间推移3种不同的植物群落中土壤含水量都有减少的趋势。季风林(p<0.01)和混交林(p<0.05)的土壤含水量减少趋势具有统计上的显著性,松林除外。4)鼎湖山3个处于不同演替阶段的植物群落其穿透水量与大气降水皆呈线性相关,它们的R2值随演替的进展而减小。穿透水占大气降水的比例也随演替进展而减少,松林、混交林和季风林分别为83.4%、68.3%和59.9%。松林、混交林和季风林的树干茎流占大气降水的比例分别为1.9%、6.5%和8.3%。树干茎流和胸径的关系受控于整个群落整体的影响而不仅仅是某个单一物种,并且群落的郁闭程度和结构是影响降水在林内再次分配的关键因素。季风林2月大气降水28.7mm林冠截流率为83.3%,而在大气降水为297.8mm的6月林冠截留率仅为18.9%,并且随着植物群落的演替,从松林、混交林到季风林的过程中林冠截留逐渐增大。