The occurrence of fungi in hospital and community potable waters

The prevalence of fungi was investigated in 126 potable water samples (84 hospital and 42 community samples), in parallel with the standard pollution indicator micro-organisms. Filamentous fungi were isolated from 104 of 126 (82.5%) samples and yeasts from 14 (11.1%), whereas their mean counts were 36.6 and 4.4, respectively. Fungi were isolated from 95.2% of community and 76.2% of hospital water samples, with the difference being statistically significant (P < 0.05), while yeasts were isolated from 9.5 and 11.9%, respectively. Prevailing genera were Penicillium spp., isolated from 64, Aspergillus spp., from 53, and Candida, from nine of the examined samples. Colony-forming units of yeasts were significantly correlated with those of total and faecal coliforms, whereas the counts of filamentous fungi were significantly correlated with total heterotrophic bacteria counts. These results suggest that tap water is a potential transmission route for fungi both in hospitals and the community in the examined region and may pose a health hazard mainly for the immunocompromised host.     

Magnitude of pollution indicator organisms in rural potable water.

A total of 460 water samples were randomly drawn from the potable water supply sources of rural communities in three counties of South Carolina. About 10% of the population, not incorporated in municipalities, was sampled. The samples were tested for total coliforms, Escherichia coli, and fecal streptococci. Significant levels of these pollution indicator organisms were detected in almost all the water supplies. Total coliforms were the most common, and only 7.5% of the water supplies were uncontaminated. E. coli, considered a reliable indicator of recent and dangerous pollution, was observed in 43% of the water supplies. Statistical analyses indicated that the bacterial populations, especially E. coli, were associated with the supply source depth and its distance from the septic tank. Total coliform counts were also weakly correlated to the pH of the water.     

Magnitude of pollution indicator organisms in rural potable water.

A total of 460 water samples were randomly drawn from the potable water supply sources of rural communities in three counties of South Carolina. About 10% of the population, not incorporated in municipalities, was sampled. The samples were tested for total coliforms, Escherichia coli, and fecal streptococci. Significant levels of these pollution indicator organisms were detected in almost all the water supplies. Total coliforms were the most common, and only 7.5% of the water supplies were uncontaminated. E. coli, considered a reliable indicator of recent and dangerous pollution, was observed in 43% of the water supplies. Statistical analyses indicated that the bacterial populations, especially E. coli, were associated with the supply source depth and its distance from the septic tank. Total coliform counts were also weakly correlated to the pH of the water.     

Failure of the most-probable-number technique to detect coliforms in drinking water and raw water supplies.

A procedure was developed to detect false-negative reactions (interference) in the standard most-probable-number (S-MPN) technique for coliform enumeration of untreated surface water and potable water supplies. This modified MPN (M-MPN) procedure allowed a quantitative assessment of the interference with coliform detection in untreated surface water and potable water supplies. Coliform interference was found to occur in the presumptive, confirmed, and completed tests of the S-MPN technique. When coliforms were present, interference with their detection occurred in over 80% of the samples. The inferior nature of the S-MPN was revealed by the 100% increase in the incidence of completed coliform-positive drinking water samples obtained with the M-MPN technique. The M-MPN procedure was also superior to the standard membrane filter technique. Eight different species of coliforms were recovered from false-negative tests, including Citrobacter, Enterobacter, Klebsiella, and Escherichia coli (in decreasing order of occurrence). The use of standard MPN techniques for monitoring potable water supplies may lead to a false security that the drinking water supply is potable, i.e., free from indicator bacteria.     

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.     

Preliminary study of microbiological parameters in eight inland recreational waters

A pilot survey of the counts of total coliform bacteria, thermotolerant coliform bacteria, Escherichia coli and faecal streptococci was carried out at eight inland recreational waters at weekly intervals during July 1991. The aims were to assess the feasibility of determining candidate indicators of recreational water quality and to assess the possible scale of variability of these parameters. The numbers of total coliforms were difficult to determine reliably because of interference from the background bacterial flora. There was a strong correlation between thermotolerant coliforms and E. coli and faecal streptococci. The average counts of the indicator organisms varied between and within the eight recreational waters by up to 10000-fold. The greatest variation was between the eight recreational waters. At any one water, the greatest source of variation was time but there was substantial variation between sample points at one time. Counts in samples collected 1 m apart exhibited greater than random variation. Counts from surface samples tended to be higher than those at 30 cm or 100 cm depth. The proportion of thermotolerant coliforms confirmed to be E. coli varied from water to water between 60% and 96%.     

Incidence of coliphage in potable water supplies.

Samples of drinking water from different sources in greater Cairo, Egypt, and bottled drinking water were tested for total coliform, fecal coliform, and coliphage populations. Of the 147 samples tested, 4 samples were positive for both total coliforms and coliphage, 65 samples were negative for total coliforms, fecal coliforms, and coliphage, and 78 samples were positive for coliphage and negative for total coliforms and fecal coliforms. The incidence of coliphage in these potable water supplies reflects the probability of human pathogenic virus survival in these waters also.     

Incidence of coliphage in potable water supplies

Samples of drinking water from different sources in greater Cairo, Egypt, and bottled drinking water were tested for total coliform, fecal coliform, and coliphage populations. Of the 147 samples tested, 4 samples were positive for both total coliforms and coliphage, 65 samples were negative for total coliforms, fecal coliforms, and coliphage, and 78 samples were positive for coliphage and negative for total coliforms and fecal coliforms. The incidence of coliphage in these potable water supplies reflects the probability of human pathogenic virus survival in these waters also.     

Long-term variations in abundance and distribution of sewage pollution indicator and human pathogenic bacteria along the central west coast of India

Safe water quality criteria on the load and types of microbial populations are important for human use from fishery, tourism and navigational viewpoints. To understand the variations in sewage pollution indicator and certain human pathogenic bacteria, data collected from various locations along central west coast of India during 2002–2007 were analyzed. Water and sediment samples were examined for total viable counts (TVC), pollution indicator bacteria (total coliforms – TC, fecal coliforms – FC and Escherichia coli – EC) and potential pathogens (Vibrio cholerae – VC, Shigella – SH, and Salmonella spp. – SA). In both Mandovi and Zuari estuaries, where fishing and tourist-related activities are sizable and long-term data collection was regular, we observed high counts of TC, FC, VC, SH and SA in particular during monsoon due to increased land runoff. Further, the abundance of TC and FC has increased significantly over the years in the water column to much above either USEPA or India permissible limits. The concentrations of Vibrio cholerae, and Shigella correlated with those of coliforms. Pathogenic bacteria were detected even 20 km and/or 25 km offshore mainly due to dumping of raw or improperly treated sewage effluents either from land, fishing trawlers and/or ships in the anchorage. Higher concentrations of fecal coliforms and pathogenic bacteria in neretic waters signify threats to environmental and human health.     

The fate of stormwater-associated bacteria in constructed wetland and water pollution control pond systems

The performances of a constructed wetland and a water pollution control pond were compared in terms of their abilities to reduce stormwater bacterial loads to recreational waters. Concentrations of thermotolerant coliforms, enterococci and heterotrophic bacteria were determined in inflow and outflow samples collected from each system over a 6-month period. Bacterial removal was significantly less effective in the water pollution control pond than in the constructed wetland. This was attributed to the inability of the pond system to retain the fine clay particles (< 2 microm) to which the bacteria were predominantly adsorbed. Sediment microcosm survival studies showed that the persistence of thermotolerant coliforms was greater in the pond sediments than in the wetland sediments, and that predation was a major factor influencing bacterial survival. The key to greater bacterial longevity in the pond sediments appeared to be the adsorption of bacteria to fine particles, which protected them from predators. These observations may significantly affect the choice of treatment system for effective stormwater management.     

Effect of noncoliforms on coliform detection in potable groundwater: improved recovery with an anaerobic membrane filter technique

A total of 529 well and distribution potable water samples were analyzed for total coliforms by the most-probable-number and membrane filter (MF) techniques. Standard plate count bacteria and MF noncoliform bacteria were also enumerated. Frequency of coliform detection, turbidity in most-probable-number tubes, and extensive overgrowth by noncoliforms on MF filters were directly proportional to standard plate counts. Recovery of coliforms was greatest by the MF method at low (less than 100 CFU/ml) standard plate count densities and better by the most-probable-number method (confirming gas and turbid tube) at high (greater than 500 CFU/ml) standard plate count densities. In the latter case, overgrowth by noncoliforms on MF filters suppressed sheen development and, in turn, masked coliform detection. Of 341 atypical (no sheen) MF colonies verified by parallel inoculation of lauryl sulfate broth and billiant green-bile broth, 156 were aerogenic in the latter medium. Of atypical isolates, 84% were identified as either Citrobacter or Enterobacter species. A 4.3-fold reduction in numbers of overgrown MF filters and an 2.2-fold increase in numbers of coliforms recovered from 127 water samples was accomplished by anaerobic incubation of MF cultures. This anaerobic modification of the standard MF technique significantly reduced overgrowth and enhanced recovery of coliforms from potable groundwater. This technique is simple, cost effective, and suitable for monitoring of untreated ground water common to some small water systems and private water supplies.     

Effect of noncoliforms on coliform detection in potable groundwater: improved recovery with an anaerobic membrane filter technique.

A total of 529 well and distribution potable water samples were analyzed for total coliforms by the most-probable-number and membrane filter (MF) techniques. Standard plate count bacteria and MF noncoliform bacteria were also enumerated. Frequency of coliform detection, turbidity in most-probable-number tubes, and extensive overgrowth by noncoliforms on MF filters were directly proportional to standard plate counts. Recovery of coliforms was greatest by the MF method at low (less than 100 CFU/ml) standard plate count densities and better by the most-probable-number method (confirming gas and turbid tube) at high (greater than 500 CFU/ml) standard plate count densities. In the latter case, overgrowth by noncoliforms on MF filters suppressed sheen development and, in turn, masked coliform detection. Of 341 atypical (no sheen) MF colonies verified by parallel inoculation of lauryl sulfate broth and billiant green-bile broth, 156 were aerogenic in the latter medium. Of atypical isolates, 84% were identified as either Citrobacter or Enterobacter species. A 4.3-fold reduction in numbers of overgrown MF filters and an 2.2-fold increase in numbers of coliforms recovered from 127 water samples was accomplished by anaerobic incubation of MF cultures. This anaerobic modification of the standard MF technique significantly reduced overgrowth and enhanced recovery of coliforms from potable groundwater. This technique is simple, cost effective, and suitable for monitoring of untreated ground water common to some small water systems and private water supplies.     

Distribution and Significance of Fecal Indicator Organisms in the Upper Chesapeake Bay

Total viable aerobic, heterotrophic bacteria, total coliforms, fecal coliforms, and fecal streptococci were enumerated in samples collected at five stations located in the Upper Chesapeake Bay, December 1973 through December 1974. Significant levels of pollution indicator organisms were detected at all of the stations sampled. Highest counts were observed in samples collected at the confluence of the Susquehanna River and the Chesapeake Bay. The indicator organisms examined were observed to be quantitatively distributed independently of temperature and salinity. Counts were not found to be correlated with concentration of suspended sediment. However, significant proportions of both the total viable bacteria (53%) and fecal indicator organisms (>80%) were directly associated with suspended sediments. Correlation coefficients (r) for the indicator organisms examined in this study ranged from r = 0.80 to r = 0.99 for bottom water and suspended sediment, respectively. Prolonged survival of fecal streptococci in most of the sediment samples was observed, with concomitant reduction of the correlation coefficient from r = 0.99, fecal streptococci to total coliforms in water, to r = 0.01, fecal streptococci to fecal coliforms in sediments. The results of this study compared favorably with fecal coliforms: fecal streptococci ratios for the various sample types. Characterization of organisms beyond the confirmed most-probable-number procedure provided good correlation between bacterial indicator groups.     

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.     

Comparison of four membrane filter methods for fecal coliform enumeration in tropical waters.

Four membrane filter methods for the enumeration of fecal coliforms were compared for accuracy, specificity, and recovery. Water samples were taken several times from 13 marine, 1 estuarine, and 4 freshwater sites around Puerto Rico, from pristine waters and waters receiving treated and untreated sewage and effluent from a tuna cannery and a rum distillery. Differences of 1 to 3 orders of magnitude in the levels of fecal coliforms were observed in some samples by different recovery techniques. Marine water samples gave poorer results, in terms of specificity, selectivity, and comparability, than freshwater samples for all four fecal coliform methods used. The method using Difco m-FC agar with a resuscitation step gave the best overall results; however, even this method gave higher false-positive error, higher undetected-target error, lower selectivity, and higher recovery of nontarget organisms than the method using MacConkey membrane broth, the worst method for temperate waters. All methods tested were unacceptable for the enumeration of fecal coliforms in tropical fresh and marine waters. Thus, considering the high densities of fecal coliforms observed at most sites in Puerto Rico by all these methods, it would seem that these density estimates are, in many cases, grossly overestimating the degree of recent fecal contamination. Since Escherichia coli appears to be a normal inhabitant of tropical waters, fecal contamination may be indicated when none is present. Using fecal coliforms as an indicator is grossly inadequate for the detection of recent human fecal contamination and associated pathogens in both marine and fresh tropical waters.     

Marine bacteria cause false-positive results in the Colilert-18 rapid identification test for Escherichia coli in Florida waters

The Colilert-18 system for enumeration of total coliforms and Escherichia coli is approved by the U.S. Environmental Protection Agency for use in drinking water analysis and is also used by various agencies and research studies for enumeration of indicator organisms in fresh and saline waters. During monitoring of Pinellas County, Fla., marine waters, estimates of E. coli numbers (by Colilert-18) frequently exceeded fecal coliform counts (by membrane filtration) by 1 to 3 orders of magnitude. Samples from freshwater sites did not display similar discrepancies. Fecal coliforms, including E. coli, could be cultured from 100% of yellow fluorescent wells (denoting E. coli-positive results) inoculated with freshwater samples but could be cultured from only 17.1% of the "positive" wells inoculated with marine samples. Ortho-nitrophenyl-beta-D-galactopyranoside (ONPG)-positive or 4-methylumbelliferyl-beta-D-glucuronide (MUG)-positive noncoliform bacteria were readily cultured from Colilert-18 test wells inoculated with marine samples. Filtered cell-free seawater did not cause false positives. Coculture preparations of as few as 5 CFU of Vibrio cholerae (ONPG positive) and Providencia sp. (MUG positive) ml(-1) inoculated into Colilert-18 caused false-positive E. coli results. Salinity conditions influenced coculture results, as the concentration of coculture inoculum required to cause false positives in most wells increased from about 5 CFU ml(-1) in seawater diluted 1:10 with freshwater to approximately equal to 5,000 CFU ml(-1) in seawater diluted 1:20 with freshwater. Estimated E. coli numbers in various marine water samples processed at the 1:10 dilution ranged from 10 to 7,270 CFU.100 ml(-1), while E. coli numbers in the same samples processed at the 1:20 dilution did not exceed 40 CFU.100 ml(-1). The lower estimates of E. coli numbers corresponded well with fecal coliform counts by membrane filtration. This study indicates that assessment of E. coli in subtropical marine waters by Colilert-18 is not accurate when the recommended 1:10 sample dilution is used. The results suggest that greater dilution may diminish the false-positive problem, but further study of this possibility is recommended.     

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.     

Detection of human-derived fecal pollution in environmental waters by use of a PCR-based human polyomavirus assay

Regulatory agencies mandate the use of fecal coliforms, Escherichia coli or Enterococcus spp., as microbial indicators of recreational water quality. These indicators of fecal pollution do not identify the specific sources of pollution and at times underestimate health risks associated with recreational water use. This study proposes the use of human polyomaviruses (HPyVs), which are widespread among human populations, as indicators of human fecal pollution. A method was developed to concentrate and extract HPyV DNA from environmental water samples and then to amplify it by nested PCR. HPyVs were detected in as little as 1 microl of sewage and were not amplified from dairy cow or pig wastes. Environmental water samples were screened for the presence of HPyVs and two additional markers of human fecal pollution: the Enterococcus faecium esp gene and the 16S rRNA gene of human-associated Bacteroides. The presence of human-specific indicators of fecal pollution was compared to fecal coliform and Enterococcus concentrations. HPyVs were detected in 19 of 20 (95%) samples containing the E. faecium esp gene and Bacteroides human markers. Weak or no correlation was observed between the presence/absence of human-associated indicators and counts of indicator bacteria. The sensitivity, specificity, and correlation with other human-associated markers suggest that the HPyV assay could be a useful predictor of human fecal pollution in environmental waters and an important component of the microbial-source-tracking "toolbox."     

Elimination of viruses and indicator bacteria at each step of treatment during preparation of drinking water at seven water treatment plants.

Seven drinking water treatment plants were sampled twice a month for 12 months to evaluate the removal of indicator bacteria and cytopathogenic enteric viruses. Samples were obtained at each level of treatment: raw water, postchlorination, postsedimentation, postfiltration, postozonation, and finished (tap) water. Raw water quality was usually poor, with total coliform counts exceeding 105 to 106 CFU/liter and the average virus count in raw water of 3.3 most probable number of cytopathogenic units (MPNCU)/liter; several samples contained more than 100 MPNCU/liter. All plants distributed finished water that was essentially free of indicator bacteria as judged by analysis of 1 liter for total coliforms, fecal coliforms, fecal streptococci, coagulase-positive staphylococci, and Pseudomonas aeruginosa. The total plate counts at 20 and 35 degrees C were also evaluated as a measure of the total microbial population and were usually very low. Viruses were detected in 7% (11 of 155) of the finished water samples (1,000 liters) at an average density of 0.0006 MPNCU/liter the highest virus density measured being 0.2 MPNCU/liter. The average cumulative virus reduction was 95.15% after sedimentation and 99.97% after filtration and did not significantly decrease after ozonation or final chlorination. The viruses isolated from treated waters were all enteroviruses: poliovirus types 1, 2, and 3, coxsackievirus types B3, B4, and B5, echovirus type 7, and untyped picornaviruses.     

Elimination of viruses and indicator bacteria at each step of treatment during preparation of drinking water at seven water treatment plants

Seven drinking water treatment plants were sampled twice a month for 12 months to evaluate the removal of indicator bacteria and cytopathogenic enteric viruses. Samples were obtained at each level of treatment: raw water, postchlorination, postsedimentation, postfiltration, postozonation, and finished (tap) water. Raw water quality was usually poor, with total coliform counts exceeding 105 to 106 CFU/liter and the average virus count in raw water of 3.3 most probable number of cytopathogenic units (MPNCU)/liter; several samples contained more than 100 MPNCU/liter. All plants distributed finished water that was essentially free of indicator bacteria as judged by analysis of 1 liter for total coliforms, fecal coliforms, fecal streptococci, coagulase-positive staphylococci, and Pseudomonas aeruginosa. The total plate counts at 20 and 35 degrees C were also evaluated as a measure of the total microbial population and were usually very low. Viruses were detected in 7% (11 of 155) of the finished water samples (1,000 liters) at an average density of 0.0006 MPNCU/liter the highest virus density measured being 0.2 MPNCU/liter. The average cumulative virus reduction was 95.15% after sedimentation and 99.97% after filtration and did not significantly decrease after ozonation or final chlorination. The viruses isolated from treated waters were all enteroviruses: poliovirus types 1, 2, and 3, coxsackievirus types B3, B4, and B5, echovirus type 7, and untyped picornaviruses.