Fecal Pollution In Water Research Articles

How suitable is freshwater sponge Ephydatia fluviatilis (Linnaeus, 1759) for time-integrated biomonitoring of microbial water quality?

Faecal pollution of water by bacteria has a negative effect on water quality and can pose a potential health hazard. Conventional surveillance of microbial water quality relies on the analysis of low-frequency spot samples and is thus likely to miss episodic or periodic pollution. This study aimed to investigate the potential of filter-feeding sponges for time-integrated biomonitoring of microbial water quality. Laboratory trials tested the effects of different ratios of bacterial abundance and the sequence of exposure on bacterial retention by the freshwater sponge Ephydatia fluviatilis (Linnaeus, 1759) to establish its potential to indicate bacterial exposure. Gemmule grown sponges were simultaneously exposed to Escherichia coli and Enterococcus faecalis but at different ratios (Trial 1) or individually exposed to each bacterial species but in different sequential order (Trial 2). The E. coli and E. faecalis retained in each sponge was quantified by culture on selective agars. Data analysis was conducted using the Kruskal-Wallis test and/or the Mann-Whitney U test to compare between the numbers of bacteria retained in each treatment. Additionally, the Wilcoxon matched-paired signed-rank test was used for comparison of the different bacterial abundances retained within each individual sponge. Sponges from all trials retained E. coli and E. faecalis in small numbers relative to the exposure (<0.05 % Trial 1 and <0.07 % Trial 2) but exhibited higher retention of E. coli. Higher abundance of either bacterial species resulted in significantly lower (P<0.005) retention of the same species within sponges (Trial 1). An initial exposure to E. coli resulted in significantly higher (P=0.040) retention of both bacterial species than when sponges were exposed to E. faecalis first (Trial 2).Bacterial retention by sponges was neither quantitatively representative of bacterial abundance in the ambient water nor the sequence of exposure. This implies either selective filtration or an attempt by sponges to prevent infection. However, freshwater sponges may still be useful in biomonitoring as qualitative time-integrated samplers of faecal indicator bacteria as they detect different bacteria present in the water even if their quantities cannot be estimated.

The use of sterol profiles, supported with other faecal source tracking methods, to apportion septic tanks contamination in rural catchments

Identifying the origin of faecal pollution in water is needed for effective water management decisions to protect both human health and aquatic ecosystems. Traditionally used indicators of faecal contamination, such as E. coli, only indicate pollution from warm-blooded animals and not the specific source of contamination; hence, more source specific tracers are required. The study has focussed on separating the two main sources of contaminants within rural catchments in Ireland, agriculture and on-site wastewater treatment systems (predominantly septic tanks). While human-specific effluent tracers may assist in identifying potential pathways from individual septic tanks to surface waters, it is difficult to quantify the cumulative impact of such systems at a catchment scale. This study has investigated faecal sterols as a method to quantify such an impact on four small catchments in areas of low subsoil permeability with high densities of septic tanks. The results demonstrate the usefulness of faecal sterols which provide a quantitative evaluation of the respective impact between agricultural pasture inputs and on-site effluent showing differences between the four catchments. The study also highlights the need to derive more specific local reference sterol profile databases for specific countries or regions, using local source material of animal faeces and effluent. Two intensive sampling campaigns on the four catchments then used faecal sterols in parallel to fluorescent whitening compounds (FWCs), caffeine, artificial sweeteners and selected pharmaceuticals to gain further insights and confirmation about contamination hotspots as well as providing comparison between the different parameters. The combination of sterols, FWCs, caffeine, acesulfame and cyclamate has proven suitable to provide an estimate of the extent of human contamination in these rural catchments and has yielded additional information about potential pollution pathways and proximity of contamination. Overall, this methodology can help to facilitate a targeted and effective water management in such catchments.

Digital metabolic activity assay enables fast assessment of 2D materials bactericidal efficiency

BackgroundThe identification and quantification of viable Escherichia coli (E. coli) are important in multiple fields including the development of antimicrobial materials, water quality, food safety and infections diagnosis. However, the standard culture-based methods of viable E. coli detection suffer from long detection times (24 h) and complex operation, leaving the unmet requirement for fast assessing the efficiency of antimicrobial materials, early alerting the contamination of water and food, and immediately treatment of infections. ResultsWe present a digital β-d-glucuronidase (GUS) assay in a self-priming polydimethylsiloxane (PDMS) microfluidic chip for rapid E. coli identification and quantification. The GUS expression in viable bacteria was investigated to develop a fast GUS assay at the single-cell level. Single E. coli were stochastically discretized in picoliter chambers and identified by specific GUS activity. The digital GUS assay enabled identifying E. coli within 3 h and quantifying within 4 h for different E. coli subtypes. The specificity of our method was confirmed by using blended bacteria including E. coli, Bacillus, Shigella and Vibrio. We utilized digital GUS assay to enumerate viable E. coli after incubated with antibacterial materials for assessing the antibacterial efficiency. Moreover, the degassed chip can realize automatic sample distribution without external instruments. SignificanceThe results demonstrated the functionality and practicability of digital GUS assay for single E. coli identification and quantification. With air-tight packaging, the developed chip has the potential for on-site E. coli analysis and could be deployed for diagnosis of E. coli infections, antimicrobial susceptibility testing, and warning the fecal pollution of water. Digital GUS assay provides a paradigm, examining the activity of metabolic enzyme, for detecting the viable bacteria other than E. coli.

Marine sediments are identified as an environmental reservoir for Escherichia coli: comparing signature-based and novel amplicon sequencing approaches for microbial source tracking

Viable Escherichia coli were detected in sediments near a point of wastewater discharge in a marine coastal environment in Sweden. Since high concentrations were found in the sediments nearest the pipe, this suggested that treated wastewater effluent was the source of the microbes. In order to examine this hypothesis, different bioinformatics approaches were applied using 16S rRNA gene V3-V4 amplicon sequences from the sediments. Both signature-based source tracking using sequence libraries describing known sources of fecal water pollution (SourceTracker); and, a curated source tracking method, indicated that sediments were contaminated with wastewater. The results from the curated approach were independently confirmed using differential abundance analysis (DESeq2). A number of taxa originating from wastewater were identified which can be used to describe contamination of the sediments, and examine the spread of these specific taxa, even at low relative abundance, along the urban coast. Sequences of phylum Bacteroidetes (such as Bacteroides and Prevotella) and Firmicutes (such as Romboutsia) increased in sediments with higher concentrations of E. coli. In addition, sequences from Trichococcus are proposed as an indicator for treated wastewater. All three source tracking approaches, and the detection of viable E. coli, suggest that urban sediments can be a reservoir for indicator bacteria.

An Overview of Microbial Source Tracking Using Host-Specific Genetic Markers to Identify Origins of Fecal Contamination in Different Water Environments

Fecal contamination of water constitutes a serious health risk to humans and environmental ecosystems. This is mainly due to the fact that fecal material carries a variety of enteropathogens, which can enter and circulate in water bodies through fecal pollution. In this respect, the prompt identification of the polluting source(s) is pivotal to guiding appropriate target-specific remediation actions. Notably, microbial source tracking (MST) is widely applied to determine the host origin(s) contributing to fecal water pollution through the identification of zoogenic and/or anthropogenic sources of fecal environmental DNA (eDNA). A wide array of host-associated molecular markers have been developed and exploited for polluting source attribution in various aquatic ecosystems. This review is intended to provide the most up-to-date overview of genetic marker-based MST studies carried out in different water types, such as freshwaters (including surface and groundwaters) and seawaters (from coasts, beaches, lagoons, and estuaries), as well as drinking water systems. Focusing on the latest scientific progress/achievements, this work aims to gain updated knowledge on the applicability and robustness of using MST for water quality surveillance. Moreover, it also provides a future perspective on advancing MST applications for environmental research.

QT-AMP: Sequencing PCR amplicons from Quanti-Tray wells to analyze enterococci communities

Enterococcus is ubiquitous in human feces and has been adopted as a useful indicator of human fecal pollution in water. Although regular enterococci monitoring only examines their numbers, identifying human-specific Enterococcus species or genotypes could help discriminate human fecal contamination from other environmental sources. We documented a new approach to characterize enterococci using a high-throughput 16S rRNA gene amplicon sequencing platform from Quanti Trays after following the counting of the most probable numbers of enterococci. We named this method QT-AMP (Quanti-Tray-based amplicon sequencing). We tested surface water samples collected from three rivers in southwest Florida. We detected 11 Enterococcus species from 45 samples in 1.1 million sequence reads. The method detected three rare species and eight cosmopolitan species (Enterococcus faecalis, E. faecium, E. casseliflavus, E. hirae, E. mundtii, E. gallinarum, E. avium, and E. durans) which have been commonly documented in previous studies. The approximate detection level of QT-AMP was four orders of magnitude higher than regular 16S rRNA gene amplicon sequencing. The current Enterolert MPN method only provides quantitative information but now we can look into the relative abundance of Enterococci species composition by accompanying Illumina sequencing. This QT-AMP could be a useful tool to streamline the quantification and identification of enterococci and could be used in various water management projects and human health risk assessment.

Monitoring SARS-CoV-2 in the Wastewater and Rivers of Tapachula, a Migratory Hub in Southern Mexico.

The COVID-19 pandemic has been monitored by applying different strategies, including SARS-CoV-2 detection with clinical testing or through wastewater-based epidemiology (WBE). We used the latter approach to follow SARS-CoV-2 dispersion in Tapachula city, located in Mexico’s tropical southern border region. Tapachula is a dynamic entry point for people seeking asylum in Mexico or traveling to the USA. Clinical testing facilities for SARS-CoV-2 monitoring are limited in the city. A total of eighty water samples were collected from urban and suburban rivers and sewage and a wastewater treatment plant over 4 months in Tapachula. We concentrated viral particles with a PEG-8000-based method, performed RNA extraction, and detected SARS-CoV-2 particles through RT-PCR. We considered the pepper mild mottle virus as a fecal water pollution biomarker and analytical control. SARS-CoV-2 viral loads (N1 and N2 markers) were quantified and correlated with official regional statistics of COVID-19 bed occupancy and confirmed cases (r > 91%). Our results concluded that WBE proved a valuable tool for tracing and tracking the COVID-19 pandemic in tropical countries with similar water temperatures (21–29 °C). Monitoring SARS-CoV-2 through urban and suburban river water sampling would be helpful in places lacking a wastewater treatment plant or water bodies with sewage discharges.Supplementary InformationThe online version contains supplementary material available at 10.1007/s12560-022-09523-2.

Modeling human pollution in water bodies using somatic coliphages and bacteriophages that infect Bacteroides thetaiotaomicron strain GA17

The ability to detect human fecal pollution in water is of great importance when assessing the associated health risks. Many microbial source tracking (MST) markers have been proposed to determine the origin of fecal pollution, but their application remains challenging. A range of factors, not yet sufficiently analyzed, may affect MST markers in the environment, such as dilution and inactivation processes. In this work, a statistical framework based on Monte Carlo simulations and non-linear regression was used to develop a classification procedure for use in MST studies. The predictive model tested uses only two parameters: somatic coliphages (SOMCPH), as an index of general fecal pollution, and human host-specific bacteriophages that infect Bacteroides thetaiotaomicron strain GA17 (GA17PH). Taking into account bacteriophage dilution and differential inactivation, the threshold concentration of SOMCPH was calculated to be around 500 PFU/100 mL for a limit of detection of 10 PFU/100 mL. However, this threshold can be lowered by increasing the analyzed volume sample, which in turn lowers the limit of detection. The resulting model is sufficiently accurate for application in practical cases involving MST and could be easily used with markers other than those tested here.

Development of a Microbiological Paper-Based Analytical Device to Detect Fecal Contamination of Water in Resource-Limited Settings

Microbial contamination of water is a significant burden to public health, particularly in low- and middle-income countries. To develop an affordable and user-friendly method for detecting microbial water contamination in resource-limited settings, a business card-sized device, named the MicroBioPAD, was developed using filter paper saturated with the chromogenic sugar X-Gal. This device was compared to the standard most probable number (MPN) test and a plate culture-based test to assess fecal pollution of water in Nepal. A total of 120 water samples were collected from the Central (Kathmandu) and Southern (Chitwan) regions; these included 36 samples from three major rivers and 84 samples from various drinking water sources. The MPN and plate culture tests revealed fecal contamination in 93.1 and 27.3% of the samples collected in Kathmandu and Chitwan, respectively, with the levels of E. coli ranging from < 3 to ≥ 1100 CFU/100 ml in the Kathmandu water samples and < 3 to 160 CFU/100 ml in the Chitwan water samples. Compared to the standard culture-based tests, the MicroBioPAD produced positive results within a shorter time (24 h) and showed a 100% accuracy of detection for samples containing ≥ 3 E. coli CFU/ml, suggesting a detection limit of 3 E. coli CFU/ml when using only 100 μl of the sample. The MicroBioPAD will be useful as a relatively faster semi-quantitative device to determine water suitability for bathing, cleaning, and recreational purposes.

Relation between Water Source and Rotaviral infection: An Observational Study

Fecal pollution of water is a major concern in river based countries as it is associated with transmition of the pathogens. Rotavirus is one of them which is the major cause of acute severe dehydrating diarrhoea in children below five years. This cross-sectional observational study was designed to determine the association of contaminated water source with Rotaviral infection among under five children admitted in the Department of Paediatrics, Sylhet MAG Osmani Medical College Hospital, Sylhet and was carried out in the Department of Microbiology during the period from 1st January to 31st December, 2018. Total 184 under five children with acute watery diarrhoea were enrolled in this study by convenient sampling. Stool samples were obtained and assayed for rotavirus antigens by enzyme linked immunosorbent assay (ELISA). Rotaviral antigen was found positive in 86 cases. The Rotavirus infection was found highest in age group of 7 to 12 months (50.56%) and in male (59.30%) children. It was found significantly higher in patients from lower socio-economic condition (64.00%), in rural area (48.75%) and those who consumed unboiled pond water (94.44%). It was found higher among those children who were not exclusively breastfed (83.87%). Bottle feeding, lower educational level of mother and improper hand washing may serve as predisposing factors of rotavirus disease in these children. Using of contaminated drinking and household water has an important relation with Rotaviral infection. Ignorance of using boiled water can play a significant role for increased prevalence of Rotaviral diarrhoea.

Pepper mild mottle virus in different water matrices

Abstract Background Pepper mild mottle virus (PMMoV), a plant virus belonging to Virgoviridae, has recently been suggested as a potential viral indicator for faecal pollution in aquatic environments, since it has been found to be abundantly excreted from healthy human subjects. Methods The occurrence, amount and diversity of PMMoV was investigated in water environments by nested RT-PCR and TaqMan based quantitative PCR. During 2017-2019, 251 water samples (92 urban wastewaters, 32 treated effluents, 16 surface water samples, 9 estuarine samples, 20 seawater samples, 67 groundwater samples, and 15 drinking waters) were analysed. Results PMMoV was detected in 73/92 (79%) wastewater samples, 22/32 (69%) treated sewages, 11/16 (69%) river samples, 6/9 (67%) estuarine samples, 5/20 (25%) bathing waters, and 9/67 (13%) groundwaters, whilst drinking water samples tested always negative. Mean viral concentrations (genome copies/L) were: raw sewage 4.2 × 106, treated sewage 7.4 × 105, river 3.2 × 103, estuarine waters 9.6 × 102, seawaters 3.0 × 102, groundwaters 7.7 × 101. Conclusions This study highlights the significant occurrence of PMMoV in aquatic environment in Italy, and a clear gradient of viral prevalence and concentrations from polluted to clean waters (wastewaters to drinking waters). Key messages PMMoV is ubiquitous throughout the water cycle in Italy with different concentrations. Studies are needed to evaluate the suitability of PMMoV as a viral indicator for human fecal pollution in waters.

Bacteriological quality of water samples from Kochi, Southwest Coastal India and its implications

BACKGROUND: Coliforms and other bacterial indicators are known to be present in Community water samples causing faecal pollution of water. There is also a growing body of evidence demonstrating that the aquatic environmental microbes are drug resistant. OBJECTIVE: The objective of this study was to evaluate the microbial quality of the water from tap, well and pond in urban households of Kochi, Kerala state. MATERIALS AND METHODS: A total of 100 water samples (46 from well water, 45 from tap water and 9 from lakes and ponds) were analysed by the multiple fermentation tube method to determine the presumptive coliform count or the most probable number (MPN) of coliforms, and the isolates were identified using standard procedures, followed by susceptibility testing. RESULTS: Eighty samples were positive for growth (76 grew coliforms, four samples grew non-pathogens). A total of 105 coliform isolates were grown in culture. The MPN numbers were noted to be >100 for over 50% of all the samples (n = 56), thus making the water unsatisfactory for drinking purposes. The highest number of organism isolated was Klebsiella pneumonia (n = 55), followed by Enterobacter spp (n = 34) and Escherichia coli (n = 16). Well water had the highest proportion of all three of these organisms. Up to 26%–40% resistance was seen to amoxicillin–clavulanic acid and 5.4%–5.8% of resistance was noted to third-generation cephalosporins, while two isolates were extended-spectrum beta-lactamase-producing Enterobacteriaceae (CTX-M and TEM types). CONCLUSION: The city municipal water authorities need to adopt more aggressive treatment/disinfection practices to combat high coliform contamination. The bacteriological quality of well water at the source needs to be monitored. Meanwhile, continued adequate home purification is suggested for drinking water. The city water does not pose a threat of antibiotic resistance for now. Environmental sampling should be given equal priority as clinical sampling in the coming days.

A study of isolation and identification of bacteria from lake water in and around Udaipur, Rajasthan.

Introduction:Water is an essential nutrient which plays an important role in digestion, absorption of food and elimination of waste products by urine. Udaipur also known as ‘City of Lakes’ or even the ‘Venice of East’ due to its unique and beautiful lake system has an exceptional importance at national and international levels. The aquatic systems are mostly dominated by bacteria and fungi and in the natural environments micro-organisms have very specific roles with regard to the recycling of materials and purification of water. Presence of coliform bacteria in the water indicates the fecal pollution of water.Methods:Water from all the three lakes were included for the assessment of their water quality parameters. Biological parameters like MPN of coliforms and presence of bacterial and cyanobacterial contaminants were read. The MPN (Most Probable Number) of Coliform was determined by the Presumptive Coliform Count (Multiple Tube Method) in 100 ml of sample water.Conclusion:The coliform bacteria is the primary bacterial indicator for fecal pollution in water. The purpose of study was to make people aware about the bacterial load in water bodies. Based on this, categorical representation of water was unacceptable for the purpose of swimming and bathing.

Improving the identification of the source of faecal pollution in water using a modelling approach: From multi-source to aged and diluted samples

The last decades have seen the development of several source tracking (ST) markers to determine the source of pollution in water, but none of them show 100% specificity and sensitivity. Thus, a combination of several markers might provide a more accurate classification. In this study Ichnaea® software was improved to generate predictive models, taking into account ST marker decay rates and dilution factors to reflect the complexity of ecosystems. A total of 106 samples from 4 sources were collected in 5 European regions and 30 faecal indicators and ST markers were evaluated, including E. coli, enterococci, clostridia, bifidobacteria, somatic coliphages, host-specific bacteria, human viruses, host mitochondrial DNA, host-specific bacteriophages and artificial sweeteners. Models based on linear discriminant analysis (LDA) able to distinguish between human and non-human faecal pollution and identify faecal pollution of several origins were developed and tested with 36 additional laboratory-made samples. Almost all the ST markers showed the potential to correctly target their host in the 5 areas, although some were equivalent and redundant. The LDA-based models developed with fresh faecal samples were able to differentiate between human and non-human pollution with 98.1% accuracy in leave-one-out cross-validation (LOOCV) when using 2 molecular human ST markers (HF183 and HMBif), whereas 3 variables resulted in 100% correct classification. With 5 variables the model correctly classified all the fresh faecal samples from 4 different sources. Ichnaea® is a machine-learning software developed to improve the classification of the faecal pollution source in water, including in complex samples. In this project the models were developed using samples from a broad geographical area, but they can be tailored to determine the source of faecal pollution for any user.

Possible health impacts due to animal and human fecal pollution in water intended for drinking water supply of Rio de Janeiro, Brazil

AbstractFecal matter is considered as one of the worst pollutants in waterbodies due to the potential spread of waterborne diseases. This study aimed to determine the host-specific fecal contamination in two Brazilian watersheds and to predict the possible impacts on human health. Fecal sources were enumerated using host-specific genetic markers to swine (16S rRNA), human and bovine (archaeal nifH), and equine (archaeal mcrA). A single cycling condition was established for four markers aiming to decrease the analysis time. Fifteen samples from São João watershed (75%) and 25 from Guandu (62.5%) presenting Escherichia coli enumeration in compliance with Brazilian guidelines (&amp;lt;1,000 MPN/100 mL) showed the human marker. Furthermore, the bovine, swine, and equine markers were present in 92% (59/64), 89% (57/64), and 81% (52/64) of the water samples, respectively. The molecular markers proposed for qPCR in our study were sensitivity and specific enough to detect host-specific fecal pollution in all samples regardless of E. coli levels reaffirming the low correlation among them and supporting their use in water quality monitoring programs. To our knowledge, this is the first study using this approach for quantification of nifH, mcrA, and rrs gene-associated human and animal fecal pollution in waters intended for drinking water supply in Brazil.

Fungi Occurrence Assessment in Drinking Water Distribution Systems and Its Relationship with Fecal Indicator Bacteria

Introduction: As recommended by World Health Organization, consumption of the fungal contaminated water does not cause to serious infection, but may lead to healthy or aesthetic problems. The aim of this research was to assess the occurrence of fungi in water and its relationship with fecal indicator bacteria. Materials and Methods: 110 water samples were collected from different location of water distribution systems in Aliabad-e Katul City, North of Iran during April to November 2018. Enumeration of coliforms bacteria and fungi were performed by multiple tube fermentation and membrane filtration method, respectively; as described in Standard Methods. Results: Our results showed that fungi and total coliform were detected in 31 (28.4%) and 26 (23.9%) samples, respectively. Among samples of fungi positive, 22 (20.2%), 7 (6.4%), and 5 (4.6%) water samples were positive for Aspergillus sp., Rhizopus sp., and Penicillium sp., respectively. However, none faecal coliform and E. coli were observed in all examined samples, proposing the absence of faecal pollution in water. The mean and SD residual chlorine and pH were 0.55 ± 0.23 (mg/l) and 7.30 ± 0.30, respectively. The statistical analysis showed a remarkable difference between the prevalence of total coliforms and fungal species (P &lt; 0.001). Conclusion: Presence of potential opportunistic pathogens fungi in potable water can be considered as a health risk, especially for immuno-suppressed individuals. Therefore, cleaning the processes such as biofilm removal and addition of the free chlorine concentration can be effective to decrease fungi contamination and total coliform from water distribution system.

Bifidobacterias como indicadoras de contaminación fecal en aguas tropicales

Human fecal pollution in water constitute a serious risk to the public health, nevertheless the indicator microorganisms commonly used to detect the levels of fecal pollution does not identify the specific source. The detection of certain Bifidobacterium species as B. adolescentis and B. dentium have been proposed as an effective marker of human fecal contamination, but this has not yet been demonstrated in tropical environmental conditions. The aim of the present work was to determine the Bifidobacteria profile in one sample of water from the Mesolandia Swamp in the Colombian Caribbean and feces samples of 260 human and 94 domestic animals from a human settlement around the swamp. DNA from all the samples was amplified by PCR using specific specie primers based on the 16S rRNA gene sequence and separated by DGGE (Denaturing Gradient Gel Electrophoresis). DGGE bands were excised, reamplified, sequenced, and compared to GenBank database. Bifidobacterial DGGE profiles showed that eight species of Bifidobacterium that were found in the water sample, were also present in the human and animal feces. Bifidobacterium adolescentis and B. dentium described as potential human fecal pollution indicators were found in domestic animals. In this study under the environmental and experimental conditions evaluated was not possible to find a Bifidobacterium species as specific marker of human fecal contamination in tropical areas. However, the applied method in this study could be useful to detect fecal pollution in tropical waters allowing a nearest approximation to the origin of the fecal pollution compared with cultural traditional methods, since it was possible to find identical DNA sequences in the water and in the fecal samples.

Dynamics of crAssphage as a human source tracking marker in potentially faecally polluted environments

Recent studies have shown that crAssphage is abundant in human faecal samples worldwide. It has thus been postulated as a potential microbial source tracking (MST) marker to detect human faecal pollution in water. However, an effective implementation of crAssphage in water management strategies will depend on an understanding of its environmental dynamics. In this work, the abundance and temporal distribution of crAssphage was analysed in the effluent of wastewater treatment plants using different sewage treatments, and in two rivers (water and sediments) that differ in pollution impact and flow regime. Additionally, the influence of environmental conditions (temperature and rainfall) on the removal of the marker was studied along a river section, and natural inactivation was assessed by a mesocosms approach. Molecular and culture-based tools were used to compare crAssphage abundance and dynamics with those of bacteria and bacteriophages currently applied as global indicators (E. coli, somatic coliphages, Bacteroides GA17 bacteriophages, and the human-associated MST markers HF183 and HMBif). CrAssphage concentrations in sewage effluent and river samples were similar to those of HF183 and HMBif and higher than other general and/or culture-based indicators (by 2–3 orders of magnitude). Measurement of crAssphage abundance revealed no temporal variability in the effluent, although rainfall events affected the dynamics, possibly through the mobilisation of sediments, where the marker was detected in high concentrations, and an increase in diffuse and point pollution. Another factor affecting crAssphage inactivation was temperature. Its persistence was longer compared with other bacterial markers analysed by qPCR but lower than culturable markers. The results of this study support the use of crAssphage as a human source tracking marker of faecal pollution in water, since it has similar abundances to other molecular human MST markers, yet with a longer persistence in the environment. Nevertheless, its use in combination with infectious bacteriophages is probably advisable.

BIOLOGICAL AND MICROBIOLOGICAL MONITORING OF THE QALITY OF THE RIVER NISAVA

This paper presents the degree of disturbance of the localities on the Nišava River, which are located near Bela Palanka and Niš (after the output of untreated water from city collectors). Quality assessment parameters were macroinvertebrates and microbial indicators of faecal pollution of water. The results show that there is a continuous pollution of the river, especially at the site near Niš, which imposes the need to build a wastewater treatment plant.

New approach for the simultaneous detection of somatic coliphages and F-specific RNA coliphages as indicators of fecal pollution

Two groups of coliphages have been recently included in different water management policies as indicators of viral fecal pollution in water and food: somatic coliphages, which infect E. coli through cell wall receptors, and F-specific RNA coliphages, which infect through the F-pili. Somatic coliphages are more abundant in fecally contaminated waters, except reclaimed waters, those disinfected by UV irradiation, and some groundwater samples that show a higher level of F-specific coliphages. Somatic coliphages are morphologically similar to DNA enteric viruses while F-specific coliphages are similar to RNA viruses such as norovirus and hepatitis A viruses, which are the viral pathogens of concern in sewage. The use of strains sensitive to both types of phages has been proposed for total coliphage enumeration, thereby avoiding double analysis. The standardized methods available for coliphage detection are robust and cost-effective, but the introduction of ready-to-use methods would facilitate routine implementation in laboratories. The fastest available tool for somatic coliphage enumeration is the recently developed Bluephage, which uses a modified β-glucuronide-overexpressing E. coli strain unable to take up the glucuronide substrate. The overexpressed enzyme accumulates inside the bacterial cells until released by phage-induced cell lysis, whereupon it encounters its substrate and the medium changes from yellow to blue. The present method uses E. coli strain CB12, sensitive to somatic coliphages and F-specific coliphages due to the expression of the F-pili. The Bluephage approach incorporating CB12 detects both types of coliphages in a time range of 1:30 to 4:00 h, as assayed with coliphages from raw sewage, river water, sludge and mussels. This strategy can be applied to obtain qualitative and quantitative results and is applicable to microplates as well as to large sample volumes (100 ml). Moreover it can provide monitoring of water bodies at real time, as for example for ambient recreational beach monitoring.