The composition and persistence of faecal coliforms and enterococcal populations in sewage treatment plants

AIMS: The changes in structure and composition of faecal coliforms and enterococcal populations in sewage from different treatment plants, and the elimination of vancomycin- and erythromycin-resistant enterococci (VRE and ERE, respectively) in these treatment plants was analysed to determine any selective reduction. METHODS AND RESULTS: Faecal coliforms, enterococci, VRE, ERE and spores of sulphite-reducing bacteria were enumerated using standard methods. Samples were enriched where necessary in order to isolate antibiotic resistant strains. The structure and composition of these bacterial populations were determined by biochemical fingerprinting and clustering analysis. High diversity and similarity indexes were detected among all the bacterial populations in raw and treated sewage, independently of their origin and the treatment processes employed. Antibiotic resistant strains were detected in all sewage tested and no selective reduction was observed. CONCLUSIONS: The faecal coliforms and enterococci populations did not differ in the sewage samples studied. The vancomycin and erythromycin resistances of the enterococcal populations were similar in the sewage samples. Resistance to both antibiotics persisted after the treatment process independently of raw sewage flow, faecal origin or size of the human population contributing to sewage. However, sewage of mixed origin (human and animal) presented a lower similarity index for the two bacterial populations compared with that of the other human sewage analysed. SIGNIFICANCE AND IMPACT OF THE STUDY: Although a significant reduction in bacterial populations was observed, the persistence of VRE and ERE strains in the same proportions in sewage suggests that there is no selective elimination of bacterial populations during the treatment processes. The ability of antibiotic resistance strains to survive sewage treatment systems should be considered in certain water reuse programmes.     

Occurrence and densities of bacteriophages proposed as indicators and bacterial indicators in river waters from Europe and South America

AIMS: To evaluate the feasibility of bacteriophages as a complementary tool for water quality assessment in surface waters from different parts of the globe. METHODS AND RESULTS: Faecal coliform bacteria, enterococci, spores of sulphite-reducing clostridia, somatic coliphages, F-specific RNA bacteriophages and bacteriophages infecting Bacteroides fragilis were determined by standardized methods in raw sewage and in 392 samples of river water from 22 sampling sites in 10 rivers in Argentina, Colombia, France and Spain, which represent very different climatic and socio-economic conditions. The results showed that the indicators studied maintained the same relative densities in the raw sewage from the different areas. Classifying the river water samples according to the content of faecal coliform bacteria, it can be observed that the relative densities of the different bacterial indicators and bacteriophages changed according to the concentration of faecal coliform bacteria. There was a relative increase in the densities of all groups of bacteriophages and sulphite-reducing clostridia with respect to faecal coliforms and enterococci in the samples with low counts of faecal coliform bacteria. CONCLUSIONS: The numbers of bacterial indicators and bacteriophages were similar in the different geographical areas studied. Once released in rivers, the persistence of the different micro-organisms differed significantly. Bacteriophages and spores of sulphite-reducing clostridia persisted longer than faecal coliforms and enterococci. SIGNIFICANCE AND IMPACT OF THE STUDY: Bacteriophages in river water samples provide additional information to that provided by bacteria about the fate of faecal micro-organisms in river water. The easy, fast and cheap methods for phage determination are feasible both in industrialized and developing countries.     

Distribution and persistence of fecal bacterial populations in liquid and dewatered sludge from a biological treatment plant

The changes in composition and structure of fecal coliforms (FC) and enterococci (ENT) populations, as well as the elimination of spores of sulphite-reducing bacteria (SRB), were compared between municipal sewage and their derived sludge in a biological treatment plant in order to determine any selective reduction or adsorption to sludge during the treatment process. Additionally, the persistence of antibiotic-resistant enterococcal populations in two kinds of sludge was also considered to evaluate their potential elimination in the treatment process. Microbial indicators, vancomycin-resistant and erythromycin-resistant enterococci were enumerated. The structure and composition of FC and ENT populations were determined by biochemical fingerprinting and clustering analyses. Raw and treated sewage showed a concentration of FC 1 log unit higher than ENT and nearly 2 log units higher than spores of SRB. However, the three studied indicators showed similar concentrations in both types of sludge. Consequently, FC were eliminated in higher proportion than ENT and spores of SRB in sludge. FC and ENT populations showed high diversity and similarity population indexes for all kinds of samples. Antibiotic-resistant enterococci persisted in a similar proportion in respect to total enterococci not only in treated sewage but also in sludge. The persistence of antibiotic-resistant strains in sludge as well as in treated sewage should be considered if they are used for land disposal or for water reutilization, respectively.     

Antibiotic resistance of bacteria in raw and biologically treated sewage and in groundwater below leaking sewers

More than 750 isolates of faecal coliforms (>200 strains), enterococci (>200 strains) and pseudomonads (>340 strains) from three wastewater treatment plants (WTPs) and from four groundwater wells in the vicinity of leaking sewers were tested for resistance against 14 antibiotics. Most, or at least some, strains of the three bacterial groups, isolated from raw or treated sewage of the three WTPs, were resistant against penicillin G, ampicillin, vancomycin, erythromycin, triple sulfa and trimethoprim/sulfamethoxazole (SXT). Only a few strains of pseudomonads or faecal coliforms were resistant against some of the other tested antibiotics. The antibiotic resistances of pseudomonads, faecal coliforms and enterococci from groundwater varied to a higher extent. In contrast to the faecal coliforms and enterococci, most pseudomonads from all groundwater samples, including those from non-polluted groundwater, were additionally resistant against chloramphenicol and SXT. Pseudomonads from sewage and groundwater had more multiple antibiotic resistances than the faecal coliforms or the enterococci, and many pseudomonads from groundwater were resistant to more antibiotics than those from sewage. The pseudomonads from non-polluted groundwater were the most resistant isolates of all. The few surviving faecal coliforms in groundwater seemed to gain multiple antibiotic resistances, whereas the enterococci lost antibiotic resistances. Pseudomonads, and presumably, other autochthonous soil or groundwater bacteria, such as antibiotic-producing Actinomyces sp., seem to contribute significantly to the gene pool for acquisition of resistances against antibiotics in these environments.     

Evidence of septic system failure determined by a bacterial biochemical fingerprinting method

AIMS: To provide evidence of septic system failure by comparing two faecal indicator bacteria, enterococci and Escherichia coli, from defective septic tanks and adjacent creeks. METHODS AND RESULTS: A biochemical fingerprinting method was used to type and compare enterococci and E. coli strains from 39 septic tanks with creek water samples. Phenotypic diversity of enterococci (0.5 +/- 0.3) and E. coli (0.5 +/- 0.3) in septic tanks were significantly lower than those found in water samples (0.8 +/- 0.1, P < 0.0001 for enterococci and 0.9 +/- 0.1, P < 0.0001 for E. coli). Among 1072 enterococci isolates tested from septic tanks, 203 biochemical phenotypes (BPTs) were found of which 98 BPTs from 33 septic tanks were identical to several water samples. Similarly, among 621 E. coli isolates tested from septic tanks, 159 BPTs were found of which 53 BPTs from 26 septic tanks were also identical to water samples. The number of the latter bacteria was significantly (P = 0.01) higher in water samples collected from downstream compared with that of upstream in the study area. A high similarity between the populations of both indicator bacteria was also found between defective septic tanks and downstream water samples further indicating the contamination of both creeks by defective septic systems. CONCLUSIONS: Biochemical fingerprinting of faecal indicator bacteria is a useful and rapid method to provide direct evidence for septic system failure. Combination of both faecal indicator bacteria (enterococci and E. coli) provides a better judgement of the performance of a septic system. SIGNIFICANCE AND IMPACT OF THE STUDY: This study is the first to provide direct evidence of septic system failure by identifying the presence of specific bacterial types in septic tanks and surface waters. Based on our findings, we suggest that the performance evaluation of a septic system should be accompanied by direct analysis of faecal indicator bacteria.     

Survival of faecal indicator bacteria in tropical estuarine waters (Bangpakong River,Thailand)

AIMS: To investigate the survival of cultivable bacteria in the tropical Bangpakong estuary (Eastern Thailand) under different salinities and light conditions. METHODS AND RESULTS: Dark and light microcosm experiments using membrane diffusion chambers were carried out under three different experimental conditions, namely (i) low salinity, (ii) progressive mixing with brackish water and (iii) fast mixing with high salinity water spiked with raw urban sewage. Faecal coliforms declined faster than faecal enterococci, as shown by survival T90 values ranging from 82.2 +/- 4.2 to 14.5 +/- 0.8 h and 97.5 +/- 0.4-20.6 +/- 1.2 h, respectively. The survival of freshwater heterotrophic bacteria was higher but variable (121.2 +/- 5.0-30.1 +/- 14.3 h), whereas that of heterotrophic marine bacteria was rather stable (81.5 +/- 4.2-44.6 +/- 2.5 h). CONCLUSIONS: Overall survival was higher in low salinities. Light had a further deleterious effect, since it accelerated the decay of faecal indicators, particularly in high salinities. Faecal enterococci had a higher resistance to environmental conditions compared with faecal coliforms. SIGNIFICANCE AND IMPACT OF THE STUDY: This study is relevant to the understanding of the behaviour of different faecal indicator bacteria and the optimization of sewage treatment plants aimed at the reduction and/or elimination of faecal load discharged into estuarine waters submitted to salinity variations.     

Correlation between bacterial indicators and bacteriophages in sewage and sludge

The use of bacteriophages as potential indicators of faecal pollution has recently been studied. The correlation of the number of bacterial indicators and the presence of three groups of bacteriophages, namely somatic coliphages (SOMCPH), F-RNA-specific phages (FRNAPH) and phages of Bacteroides fragilis (BFRPH), in raw and treated wastewater and sludge is presented in this study. Raw and treated wastewater and sewage sludge samples from two wastewater treatment plants in Athens were collected on a monthly basis, over a 2-year period, and analysed for total coliforms, Escherichia coli, intestinal enterococci and the three groups of bacteriophages. A clear correlation between the number of bacterial indicators and the presence of bacteriophages was observed. SOMCPH may be used as additional indicators, because of their high densities and resistance to various treatment steps.     

Application of microbial source tracking methods in a Gulf of Mexico field setting

Aims:  Microbial water quality and possible human sources of faecal pollution were assessed in a Florida estuary that serves shellfishing and recreational activities.
Methods and Results:  Indicator organisms (IO), including faecal coliforms, Escherichia coli and enterococci, were quantified from marine and river waters, sediments and oysters. Florida recreational water standards were infrequently exceeded (6–10% of samples); however, shellfishing standards were more frequently exceeded (28%). IO concentrations in oysters and overlaying waters were significantly correlated, but oyster and sediment IO concentrations were uncorrelated. The human-associated esp gene of Enterococcus faecium was detected in marine and fresh waters at sites with suspected human sewage contamination. Lagrangian drifters, used to determine the pathways of bacterial transport and deposition, suggested that sediment deposition from the Ochlockonee River contributes to frequent detection of esp at a Gulf of Mexico beach.
Conclusions:  These data indicate that human faecal pollution affects water quality in Wakulla County and that local topography and hydrology play a role in bacterial transport and deposition.
Significance and Impact of the Study:  A combination of IO enumeration, microbial source tracking methods and regional hydrological study can reliably inform regulatory agencies of IO sources, improving risk assessment and pollution mitigation in impaired waters.     

Evaluation of Selected Membrane Filtration and Most Probable Number Methods for the Enumeration of Faecal Coliforms,Escherichia coli and Enterococci in Environmental Waters

Selected methods recommended in national and international water quality guidelines were compared in tests on environmental waters with different levels of faecal pollution. The following methods yielded no statistically significant differences in counts of faecal coliforms and Escherichia coli in raw sewage, semi-treated effluent, polluted urban run-off and stored potable water: Membrane filtration (MF) using MFc Agar or Chromocult Coliform Agar containing X-glucuronide, or a miniaturised microtitre-plate Most Probable Number (MPN) assay using a liquid growth medium containing chromogenic 4-methyl-umbelliferyl--D-glucuronide. Significant differences were, however, found between the Chromocult and the other methods for unpolluted river water. Counts of faecal enterococci in raw sewage, semi-treated effluent and polluted urban run-off, obtained by the following methods did not differ significantly: MF using M-Enterococcus Agar, Bile-Esculin Agar or Enterococcus Selective Agar, or a microtitre-plate MPN method with a liquid growth medium containing chromogenic 4-methyl-umbelliferyl--D-glucoside. Significant differences were, however, found between the MPN and the other methods for unpolluted river water and stored potable water. MF using Chromocult Coliform Agar had useful benefits for the simultaneous enumeration of coliforms and E coli. However, in view of cost and practical considerations, MF using MFc Agar or M-Enterococcus Agar proved the methods of choice for respectively enumerating faecal coliforms and E coli, or faecal enterococci, in analyses for general water quality surveillance purposes.     

Salmonellae and pollution indicator bacteria in municipal and food processing effluents and the Cornwallis River

A study was conducted to determine the incidence of fecal coliforms, fecal streptococci, Aeromonas hydrophilia, and Salmonella spp. in the waste discharges of seven sewage treatment plants, four fruit and vegetable canneries, a meat packing plant, a poultry processing plant, and a potato processing plant located along the Cornwallis River in Nova Scotia, Canada. Surface water samples were also collected from 13 locations in the river to assess the impact of these waste discharges on the receiving water quality. The results showed that the final effluents from most of the sewage treatment and processing plants were of very poor bacteriological quality, with the number of indicator bacteria comparable with those found in raw sewage. Salmonellae were isolated from the effluents of the meat and poultry plants and five of the seven sewage treatment plants surveyed. No salmonellae were detected in the effluents of the fruit and vegetable canneries. The impact of the discharge of untreated municipal and food processing wastes on the Cornwallis River water quality was evidenced by the recovery from river water of five Salmonella serotypes, and the high fecal coliform counts which exceeded recommended limit for bathing and shellfish harvesting.     

Peracetic acid as an alternative wastewater disinfectant to chlorine dioxide

AIMS: The aim of this study was to compare the efficiency of peracetic acid with that of chlorine dioxide in the disinfection of wastewater from a sewage treatment plant (serving about 650 000 inhabitants) that has been using peracetic acid as a disinfectant since 1998. METHODS AND RESULTS: A total of 23 samplings were made, each consisting of three samples: from secondary effluent, effluent disinfected with 2 mg l(-1) of peracetic acid and effluent disinfected with 2.2 mg l(-1) of chlorine dioxide (contact time 20 min). For each sample, measurements were made of the heterotrophic plate count at 36 degrees C, total and faecal coliforms, Escherichia coli, enterococci, pH, suspended solids and chemical oxygen demand (COD). During the first phase of the experiment the peracetic acid was seen to be less efficient than chlorine dioxide. To improve the disinfectant action a system of mechanical agitation was added which led to a greater efficiency in the inactivation of bacteria of faecal origin. CONCLUSIONS: Both products were found to be influenced by the level of microbial contamination, the amount of suspended solids and COD but not by the pH of the effluent before disinfection. The immediate mixing of the wastewater and disinfectant caused a greater reduction in enterococci. SIGNIFICANCE AND IMPACT OF THE STUDY: Since peracetic acid was seen to produce a high abatement of micro-organisms, it can be considered as a valid alternative to chlorine dioxide in the disinfection of wastewaters.     

The microbiological quality of drinking water sold on the streets in Kumasi,Ghana

AIM: The aim of this study was to assess the microbiological quality of Ghanaian bottled and plastic-bagged drinking water sold on the streets of Metropolitan Kumasi, Ghana. METHODS AND RESULTS: Eight bottled, 88 factory-filled plastic sachet and 40 hand-filled hand-tied polythene-bagged drinking waters were examined for the presence of heterotrophic bacteria total viable counts (TVCs), indicators of faecal contamination (total coliforms, faecal coliforms and enterococci) and for lead, manganese and iron. Heterotrophic bacteria were found in all three types of water with TVCs per millilitre ranging from 1 to 460 for bottled water, 2-6.33 x 10(5) for factory-bagged sachet water and 2.33 x 10(3)-7.33 x 10(12) for hand-filled hand-tied bagged water. None of the microbial indicators of faecal contamination were detected in bottled water, whereas 4.5% of the factory-bagged sachets contained total coliforms and 2.3% faecal coliforms, and 42.5% of the hand-filled hand-tied bags contained total coliforms, 22.5% faecal coliforms and 5% enterococci. Iron was found in all three types of drinking water but at concentrations well within the WHO recommendations. Lead and manganese were not detected. CONCLUSION: Ghanaian bottled water is of good microbiological quality but some factory-bagged sachet and hand-filled hand-tied polythene-bagged drinking water are of doubtful quality. SIGNIFICANCE AND IMPACT OF THE STUDY: Factory-bagged sachets and hand-filled hand-tied bags of drinking water sold in Ghana should be monitored for microbiological contamination, with the aim of raising standards in the industry and re-assuring the public.     

Characterization of bacterial coliform occurrences in different zones of a drinking water distribution system

AIMS: To compare the bacterial coliforms detected from occurrences in three zones of a water distribution system supplied by two separate water sources. METHODS AND RESULTS: Conventional and standardized protocols for identifying enterobacterial populations were applied. Additional tests to confirm isolates were included. Analyses of diversity and population similarity were performed using the Phene Plate System, a miniaturized biochemical phenotyping method. Isolates were identified by the API 20E system in tandem with biochemical phenotyping. A total of 16 576 samples were taken from the water distribution system, with 1416 isolates analysed. A low number of coliform occurrences were observed (2%). Escherichia coli was not detected in either water origin or in Zone 2 samples; however, in Zones 1 and 3 a low number of cases of E. coli were recorded. The percentages of E. coli depended on the identification criteria. Eight biochemical profiles for coliform populations were defined according to the results of the confirmative tests. There was a high diversity among these populations in the three zones studied, although no significant variations in their composition (associated with occurrences in the different zones) were observed. Klebsiella oxytoca was the most commonly detected species irrespective of zone, although seven other enterobacterial genera were also found. CONCLUSIONS: Analysis of the enzymatic activity of beta-glucuronidase or application of the criteria established in the norm ISO 9308-1, in tandem with thermotolerance was needed to evaluate the occurrence of E. coli in the distribution systems. Detected occurrences of bacterial coliforms could be associated with re-growth patterns for specific sampling points in the distribution system. Seasonal differences, independent of the studied zones, were observed. SIGNIFICANCE AND IMPACT OF THE STUDY: Biochemical phenotyping of bacterial coliforms was shown to be a useful method on the characterization of occurrences in water distribution systems.     

Influence of wastewater treatment on the microbial ecology of a large, temperate river system — the Danube River

In an attempt to assess the influence of tertiary-treated sewage influx on bacterial metabolism in the Danube River, bacterial abundance, ectoenzymatic activity, faecal coliforms, faecal streptococci, chlorophyll a (chl a), concentrations of dissolved (DOC) and total organic carbon (TOC) and inorganic nutrients were measured upstream of the sewage influx and compared with sampling sites downstream. Additional samples were taken near the outlet of the sewage treatment plant. Bacterial abundance as determined by epifluorescence microscopy was compared with plate counts of total heterotrophic bacteria. Significantly higher values were obtained at the stations downstream from the sewage influx only for faecal coliforms and faecal streptococci, for glucuronidase activity and bacterial biovolume. All the other parameters were not significantly different from values obtained at the upstream sampling site. Strong seasonal dependence was detectable for nitrate with high concentrations during the winter ( 250 µM) and low concentrations during summer ( 100 µM). A distinct spring phytoplankton bloom was noticeable in the river reaching chl a concentrations of 70 µg 1^–1; during the remaining seasons chl a concentrations were <20 µg 1^–1. Highly significant correlations were found between faecal coliform counts and glucuronidase activity. C:N:P ratios of 13.9:10.7:1 (for the upstream station) and 11.7:9.2:1 (for the downstream stations) of dissolved nutrients are rather low indicating severe carbon limitation of bacterioplankton. Based on our results we conclude that the heterotrophic bacterial community is not significantly effected by the input of treated sewage but faecal contamination is readily detectable over a comparatively long reach of 30 km.     

Enumeration and speciation of enterococci found in marine and intertidal sediments and coastal water in southern California

AIMS: To determine the levels and species distribution of enterococci in intertidal and marine sediments and coastal waters at two beaches frequently in violation of bacterial water standards. METHODS AND RESULTS: Faecal indicator bacteria were extracted from sediment and enumerated using membrane filtration. High levels of enterococci were detected in intertidal sediments in a seasonal river and near a storm drain outlet. Low levels were found in marine sediments at 10 m depths and in surf zone sand. Bacterial isolates presumptively identified as Enterococcus on mEI media were speciated. The predominant species found in both water and sediment included Enterococcus faecalis, Enterococcus faecium, Enterococcus hirae, Enterococcus casseliflavus and Enterococcus mundtii. A number of isolates (11-26%) from regulatory water samples presumptively identified as enterococci on mEI media were subsequently identified as species other than Enterococcus. At both study sites, the distribution of species present in water was comparable with those in sediments and the distribution of species was similar in water samples passing and exceeding bacterial indicator standards. CONCLUSIONS: High levels of Enterococcus in intertidal sediments indicate retention and possible regrowth in this environment. SIGNIFICANCE AND IMPACT OF THE STUDY: Resuspension of enterococci that are persistent in sediments may cause beach water quality failures and calls into question the specificity of this indicator for determining recent faecal contamination.     

Investigation of human sewage pollution and pathogen analysis at Florida Gulf coast beaches

Aims: Water quality at two Florida beaches was compared using faecal indicator bacteria measurements, microbial source tracking (MST) methods for detecting human source pollution and the assessment of pathogen presence. These values were also compared before and after remediation of wastewater infrastructure at one beach. Methods and Results: Faecal coliforms, Escherichia coli and enterococci were enumerated in estuarine water and sediment samples. PCR assays for the human‐associated esp gene of Enterococcus faecium and human polyomaviruses (HPyVs) were used to detect human sewage. Culturable Salmonella and enteric viruses were also analysed. MST identified human sewage contamination at one beach, leading to repair of a sewer main and relocation of portable restrooms. Exceedances of Florida recreational water regulatory standards were significantly reduced after remediation (by 52% for faecal coliforms and 39% for enterococci), and the frequency of detection of MST markers decreased. Coxsackie virus B4 and HPyVs were codetected following a major sewage spill, but Salmonella was not detected during the study. Conclusions: These data indicate that infrastructure remediation significantly reduced pollution from human sewage at the impacted beach. Significance and Impact of the Study: A comprehensive microbial water quality study that can identify contamination sources through the use of MST markers and close collaboration with local/and state agencies can result in tangible actions to improve recreational water quality and safety.     

Evaluation of the efficiency of peracetic acid in the disinfection of sewage effluents

AIMS: Evaluation of the efficiency of peracetic acid in the disinfection of wastewater in a large treatment plant. METHODS AND RESULTS: Over a period of 18 months 30 sample collections were made, each consisting of three samples taken from: raw incoming sewage, secondary effluent (after 10-12 h) and secondary effluent disinfected with 1.5-2 mg l(-1) of peracetic acid (contact time: 20 min). Total coliforms and Escherichia coli declined from 10(7) MPN 100 ml(-1) in the raw sewage to 10(2) in the disinfected effluent and the enterococci fell from 10(6) MPN 100 ml(-1) to 702 MPN 100 ml(-1). The reduction of bacteria increased with the rise in temperature and decreased with the rise in BOD5. CONCLUSIONS: Disinfection with peracetic acid reduced levels of faecal contamination by 97%, thus attaining the limit recommended by current Italian law (Escherichia coli 相似文献   

Microbe removal in secondary effluent by filtration

A study was carried out to assess the efficiency of filtration in reducing microbial contamination in municipal secondary effluent. After primary and secondary treatments, the wastewater underwent filtration through sand/hydroanthracite filters. A total of 20 samplings were made, each consisting of two instant samples (secondary effluent and filtered effluent). Each of the 40 samples was tested for: total and faecal coliforms,Escherichia coli, enterococci and somatic coliphages. The mean concentrations detected in the secondary effluent were in the order of 5 log for the total and faecal coliforms, 4 log for enterococci andEscherichia coli, and approx. 3 log for the coliphages. The filtration showed a higher efficacy in the reduction of feacal coliforms,Escherichia coli and in particular total coliforms. The results obtained for enterococci and coliphages were significanty lower. Filtration alone was not enough to reduce the bacterial indicators to within Italian legal limits, and showed a poor capacity to abate coliphages. However, by performing the filby-products and a consequent reduction in the chemical risk for the general population.     

Removal and inactivation of indicator bacteriophages in fresh waters

AIMS: The removal and inactivation of faecal coliform (FC) bacteria, enterococci (ENT), sulphite-reducing clostridia (SRC), somatic coliphages, F-specific RNA bacteriophages and bacteriophages infecting Bacteroides fragilis in fresh waters. METHODS AND RESULTS: Removal was studied in two areas of a river. The results showed different removal of each group of microbes. Faecal coliform bacteria were removed faster than any other, whereas SRC and bacteriophages infecting Bact. fragilis were the most persistent. Inactivation was measured by 'in situ' experiments, which showed significant differences in survival of the different groups of bacterial and bacteriophage indicators. The SRC and bacteriophages were more resistant than faecal coliforms and enterococci, with the exception of F-specific RNA bacteriophages in the summer. Inactivation experiments with pure cultures of bacteriophages confirmed that phage B40-8 of Bact. fragilis was the most resistant. CONCLUSIONS: Bacteria and bacteriophages show different resistance to natural inactivation. The use of phages allows information to be obtained in addition to that provided by bacterial indicators. Somatic coliphages and phages infecting Bact. fragilis might supply that indicator function. SIGNIFICANCE AND IMPACT OF THE STUDY: Confirmation was obtained that bacteriophages provided additional information to that provided by bacterial indicators to monitor the natural inactivation of viruses and/or pathogens.