Bacteriological and virological analysis of water from four fresh water beaches

Water samples from four beaches were analysed for the presence of total coliforms, fecal coliforms, fecal streptococci, salmonellae and enteric viruses. Analysis of the data did not reveal any correlation between the presence of viruses in 20 l. of water and the presence of the enteric bacteria. There was also no correlation between the presence of salmonellae and the presence of the other bacterial indicators. A positive correlation between virus isolation and water turbidity was demonstrated when turbidity was greater than 10 N.T.U.     

Distribution of enteroviruses in sediments contiguous with a deep marine sewage outfall

The distribution of enteroviruses, coliforms, fecal coliforms and fecal streptococcus discharged from a raw sewage outfall pipe 3.6 km from shore at a depth of 44 m was studied. Enteroviruses and indicator bacteria were only isolated from water in an area within 200 m of the outfall. However, viruses were isolated from sediments as far as 3.6 km from the outfall at recreational bathing beaches. The concentration of enteroviruses in the sediment at the outfall boil ranged from 112 to 78 PFU l⁻¹ and 0 to 30 PFU l⁻¹ at the bathing beach. Indicator bacteria were not detected in the water or sediment at the bathing beach. Analysis of sediments for viruses may give a better idea of the long term distribution of sewage discharged from the outfall pipe.     

Some effects of shoreline cottage development on lake bacteriological water quality

Eighteen cottaged lakes and one uncottaged lake in Southern Ontario, and their inflowing streams were surveyed for bacterial indicators of pollution by membrane filter techniques. Water samples were taken on 5 consecutive days in the spring, summer, and fall of the year. Inputs of fecal indicator bacteria were determined through an analysis of variance. Fecal coliform and fecal streptococcus levels in lakewater varied in a seemingly erratic manner. When the bacterial densities at different points on the lake were examined by an analysis of variance, the locations with significantly higher densities of fecal coliforms and fecal streptococci were most often found along the cottaged shoreline. Total coliform densities did not show this relationship with the cottaged shoreline. In addition, after rainfall, a large increase in numbers of fecal coliforms were found in the cottaged lakes but not in the uncottaged lake. These findings indicated that the cottaged shoreline was a greater source of fecal indicator bacteria than the uncottaged shoreline. However, the highest concentrations of fecal indicator bacteria in these lakes were found in the inflowing streams. The bacteria in these streams arose from stormwater from undeveloped forested watersheds and were probably, therefore, of soil or animal origin.     

The enterovirus test in the assessment of recreational water-associated gastroenteritis

This study reports the results of a series of analyses designed to identify possible mathematical relationships between the numbers of fecal streptococci vs the numbers of enteroviruses present (as measured by the enterovirus assay) in marine recreational waters in order to indirectly assess the relationship between enteroviral densities and subsequent risk of gastroenteritis among bathers exposed to marine waters contaminated with domestic sewage. A database consisting of 2066 parallel fecal streptococci and enterovirus enumerations taken from 416 different marine water locations throughout the United Kingdom was used in the analyses. Polychotomous logistic regression was used to model the probabilities of zero enterovirus, a possible infectious dose and a probable infectious dose of enterovirus being present over increasing fecal streptococci densities. The results of the polychotomous logistic regression analyses showed that the probability of the absence of enterovirus (i.e., zero enteroviruses being present) remained higher than the probability of the presence of either a possible or probable infectious dose in 101 of seawater up to a fecal streptococci density of 1000/100 ml of sample (actual estimated probabilities for the three groupings of enterovirus densities described above, at a fecal streptococci density of 1000 equaled 0.44, 0.44, and 0.12, respectively). In addition, the probability of zero enteroviruses being present remained higher than the probability of one or more enteroviruses being present in 101 of seawater up to a fecal streptococci density of 450/100 ml of sample. Since previously published epidemiological studies have shown an excess risk of gastroenteritis to occur among bathers exposed to as few as 32 fecal streptococci per 100 ml of sample, these results suggest that the actual viruses enumerated by the enterovirus assay are not etiologically related to recreational water-associated gastroenteritis. Since gastroenteritis has been the most consistently reported bathing water associated illness, these findings suggest the enterovirus assay to be of limited use in assessing marine recreational water quality given the increasing tendency to formulate water quality standards based on increased risk of gastroenteritis among bathers relative to non-bathers.     

One-year monthly quantitative survey of noroviruses, enteroviruses, and adenoviruses in wastewater collected from six plants in Japan

Sewerage systems are important nodes to monitor human enteric pathogens transmitted via water. A quantitative virus survey was performed once a month for a year to understand the seasonal profiles of noroviruses genotype 1 and genotype 2, enteroviruses, and adenoviruses in sewerage systems. A total of 72 samples of influent, secondary-treated wastewater before chlorination and effluent were collected from six wastewater treatment plants in Japan. Viruses were successfully recovered from 100ml of influent and 1000ml of the secondary-treated wastewater and effluent using the acid rinse method. Viruses were determined by the RT-PCR or PCR method to obtain the most probable number for each sample. All the samples were also assayed for fecal coliforms (FCs) by a double-layer method. The seasonal profiles of noroviruses genotype 1 and genotype 2 in influent were very similar, i.e. they were abundant in winter (from November to March) at a geometric mean value of 190 and 200 RT-PCR units/ml, respectively, and less frequent in summer (from June to September), at 4.9 and 9.1 RT-PCR units/ml, respectively. The concentrations of enteroviruses and adenoviruses were mostly constant all the year round, 17 RT-PCR units/ml and 320 PCR units/ml in influent, and 0.044 RT-PCR units/ml and 7.0 PCR units/ml in effluent, respectively.     

In situ persistence of indicator bacteria in streams polluted with acid mine drainage

Persistence of indicator organisms (total coliforms, fecal coliforms, and fecal streptococci) associated with natural samples of raw sewage was studied following in situ exposure to five aquatic environments. Three of these streams contained significant amounts of acid mine water (AMW) while the other two were relatively uncontaminated. Indicator organisms were rapidly killed upon exposure to the acid mine systems, whereas little reduction in numbers was observed in the uncontaminated streams. Seasonal changes affected survival of indicators as reflected by prolonged persistence at colder in situ water temperatures. The fecal coliform group was most susceptible to the AMW stress, while the fecal streptococci were most persistent. An enrichment technique resulted in substantially enhanced recovery of certain species of sublethally injured survivors of acid stress. Enrichment was particularly beneficial for recovery of AMW-injured fecal coliforms. Relatively little improvement in recovery of fecal streptococci was afforded by the enrichment technique.     

Intermittent filtration of wastewater--removal of fecal coliforms and fecal streptococci

Removal of fecal coliforms and fecal streptococci was monitored over a period of 13 months in 14 buried pilot scale filters, treating septic tank effluent. The effects of grain size, hydraulic dosing rate and distribution method were investigated. Two different natural sands (sorted sand and unsorted sand) and three different types of light weight aggregates (LWA 0-4 mm, LWA 2-4 mm and crushed LWA 0-3 mm) were used. Intermittent dosing rates from 20 to 80 mm/day in 12 doses per day were applied to the filters by uniform pressure distribution or point application by gravity dosing. Removal of fecal coliforms was more than three orders of magnitude higher in the media with the finest grain sizes (unsorted sand) as compared to the coarsest media (LWA 0-4 mm and LWA 2-4 mm) operated under same conditions. Fecal streptococci were determined only in effluent from filters with LWA 0-4 mm and LWA 2-4 mm. Higher removal of fecal coliforms was observed in pressure dosed filters compared to gravity dosed filters. A lower removal was observed by increasing the hydraulic dosing rate. Minimum retention time was found to be a key parameter for predicting removal of bacteria in unsaturated, aerobic filters. At minimum retention times lower than about 50 h, there was a correlation of 0.96 between retention time and removal of fecal coliforms. Retention times longer than 50 h gave almost complete removal of fecal coliforms.     

Evaluation of microbiological accumulation capability of the commercial sponge Spongia officinalis var. adriatica (Schmidt) (Porifera, Demospongiae)

This study was carried out to evaluate the microbiological accumulation capability of the demosponge Spongia officinalis var. adriatica. Six microbiological parameters were researched in two sampling periods in the water and in reared sponge samples coming from sites with different degrees of microbial contamination: an off-shore fish farm displaced off the Apulian coast (Southern Adriatic Sea) and a no-impacted area displaced into the Marine Protected Area of Porto Cesareo (Apulian coast-Ionian Sea). We detected the density of culturable heterotrophic bacteria by spread plate on marine agar, total culturable bacteria at 37 degrees C on Plate Count Agar and vibrios on thiosulphate-citrate-bile-sucrose-salt (TCBS) agar. Total and fecal coliforms as well as fecal streptococci concentrations were detected by the MPN method. Bacterial densities were always higher in the sponge homogenates compared with the corresponding seawater in the sampling points and in both sampling periods. As regard vibrios, total culturable bacteria at 37 degrees C and fecal streptococci concentrations, the highest values were observed in the sponge samples coming from the off-shore fish farm during the summer period. The ability of Spongia officinalis var. adriatica to accumulate the microbial pollution indicators suggests that this species can be employed as a bioindicator for monitoring water quality.     

Relationships between Bacteroides 16S rRNA genetic markers and presence of bacterial enteric pathogens and conventional fecal indicators

Occurrence and prevalence of different bacterial enteric pathogens as well as their relationships with conventional (total and fecal coliforms) and alternative fecal indicators (host-specific Bacteroides 16S rRNA genetic markers) were investigated for various water samples taken from different sites with different degrees of fecal contamination. The results showed that a wide range of bacterial pathogens could be detected in both municipal wastewater treatment plant samples and in surface water samples. Logistic regression analysis revealed that total and human-specific Bacteroides 16S rRNA genetic markers showed significant predictive values for the presence of Escheriachia coli O-157, Salmonella, heat-labile enterotoxin (LT) of enterotoxigenic E. coli (ETEC), and heat-stable enterotoxin for human (STh) of ETEC. The probability of occurrence of these pathogenic bacteria became significantly high when the concentrations of human-specific and total Bacteroides 16S rRNA genetic markers exceeded 10(3) and 10(4) copies/100 mL. In contrast, Clostridium perfringens was detected at high frequency regardless of sampling sites and levels of Bacteroides 16S rRNA genetic markers. No genes related to Shigella spp., Staphylococcus aureus and Vibrio cholerae were detected in any samples analyzed in this study. Conventional indicator microorganisms had low levels of correlation with the presence of pathogens as compared with the alternative fecal indicators. These results suggested that real-time PCR-based measurement of alternative Bacteroides 16S rRNA genetic markers was a rapid and sensitive tool to identify host-specific fecal pollution and probably associated bacterial pathogens. However, since one fecal indicator might not represent the relative abundance of all pathogenic bacteria, viruses and protozoa, combined application of alternative indicators with conventional ones could provide more comprehensive pictures of fecal contamination, its source and association with pathogenic microorganisms.     

Use of in-stream reservoirs to reduce bacterial contamination of rural watersheds

An investigation into bacterial water quality problems was conducted on an interconnected stream and irrigation system within the Oldman River Basin of southern Alberta, Canada. Levels of indicator bacteria, including fecal coliforms, generic Escherichia coli and fecal streptococci, were repeatedly measured in streams and irrigation return canals of this river basin during the summer of 2001. Bacterial-loading segments of the irrigation/stream system were identified through a comparison of indicator bacteria levels in pairs of upstream and downstream sites. Mann-Whitney U-tests indicated that reservoirs significantly reduced bacterial counts. A temporal comparison of E. coli counts and river discharges suggested that these indicator bacteria do not originate from within in-stream sediments. Site-specific as well as cumulative inputs from a variety of non-point sources are likely to be responsible for the high downstream levels of indicator bacteria in this water system. The use of management practices such as in-stream reservoirs may significantly reduce contamination, and increase the quality of limited rural water supplies to allow their reuse and safe discharge into downstream water sources. The identification of bacteria-loading river/canal segments could also be used to prioritize restoration projects.     

Virus survival in solid waste leachates

Disposal of domestic solid waste either in open dumps or in sanitary landfills results in leachates which may find their way into groundwater aquifers. This study is concerned with the survival of microorganisms of sanitary significance, with particular emphasis on enteric viruses, in leachates produced by model disposal systems. Data are presented that indicate the presence of considerable amounts of human and animal feces in fresh domestic refuse. Significant numbers of coliforms, fecal coliforms, and fecal streptococci are present in refuse and also persist for weeks in the leachates produced.A method for the concentration and recovery of virus from relatively large volumes of leachate is evaluated. Viruses were sporadically recovered for periods up to 20 weeks from leachates generated by systems to which poliovirus was purposely introduced as well as from control systems. The leachates produced were shown not to be acutely toxic to poliovirus.     

Reduction of Salmonella, E. Coli, coliforms and fecal streptococci by chlorination of sewage treatment plant effluents

Salmonellae are reduced by chlorination of effluents in three purification plants, when 0.10 mg mgl⁻¹ residual chlorine or more (as measured by the ortholidine method) is found. With 0.25 mgl⁻¹ the reduction factor of E. coli, coliforms and fecal streptococci amount to approx. 4 deciine concentrations do not increase the reduction.     

Occurrence of rotaviruses and enteroviruses in recreational waters of Oak Creek, Arizona

Recent epidemiological studies have shown a relationship between swimming in recreational waters meeting bacteriological standards and gastroenteritis with a suggested viral etiology. No previous studies have been conducted in the United States on the occurrence of human pathogenic enteric viruses in freshwater recreational areas. The presence of enteroviruses and rotaviruses was investigated in Oak Creek, Arizona, a heavily used recreational area. Water samples were filtered through positively charged filters (168–1555 I.), eluted with beef extract, and assayed for human enteroviruses and rotaviruses. Eighteen of the 41 recreational water samples were positive for enterovirus or rotavirus. Of these, nine samples exceeded the Arizona State recommended limit of 1 PFU 40 l⁻¹ for full body contact in effluent dominated recreational waters. Several virus positive samples met the recommended fecal coliform standards (200 CFU 100 ml⁻¹) for recreational waters indicating the inadequacy of bacterial standards for monitoring viral water quality. The isolation of the pathogenic enteric viruses (i.e., poliovirus 1, echovirus 1, coxsackievirus B1 and B6 and rotavirus) from this popular recreational water demonstrates the potential for transmission of viral disease.     

Mytilus galloprovincialis filter feeding on the bacterial community in a Mediterranean coastal area (Northern Ionian Sea, Italy)

This study was carried out seasonally, throughout a year, to evaluate the filtering activity on bacteria of Mytilus galloprovincialis. Six microbiological parameters were researched in the water and mussels samples collected along the coastal area of the Northern Ionian Sea in three stations, S. Vito, Lido Gandoli and Lido Silvana. We detected the densities of culturable heterotrophic bacteria by spread plate on Marine Agar, total culturable bacteria at 37 degrees C on Plate Count Agar and vibrios abundance on thiosulphate-citrate-bile-sucrose-salt (TCBS) agar. Total and fecal coliforms as well as fecal streptococci were determined by the Most Probable Number. Bacterial concentrations at 20 and 37 degrees C as well as vibrios concentrations were higher in the mussel samples compared to the corresponding seawater throughout the year. The results obtained could contribute to improve the information relatively either to the natural processes existing between bacteria and mussels or to the risk of human infections related to the consumption of mussels.     

Enterovirus removal from sewage: The effectiveness of four different treatment plants

Human enterovirus removal from effluents at four different sewage treatment plants was tested, an oxidation pond system, two utilising trickling filtration and the fourth incorporating sedimentation and chlorination. Virus numbers were not significantly reduced by any of the treatment plants.No correlation was found between virus numbers and faecal coliforms, suspended solids, pH or conductivity.Seasonal fluctuations in virus numbers indicated that more enteroviruses were present in the summer months than in the winter.     

Relationship of viruses and indicator bacteria in water and wastewater of Israel

Different aquatic resources of Israel were monitored to obtain information on the occurrence of enteroviruses and their relationship to the bacteriological indicators of water quality. A total of 155 samples of groundwater, surface water, potable water and swimming pool water were examined, of which 45 (29.0%) yielded virus. Echovirus type 7 was the most predominant, followed by poliovirus 1, echovirus 6 and coxsackieviruses B2, B5 and B6. Virus was isolated on several occasions when the water in question met current bacteriological standards. Viruses were isolated from water samples with no detectable fecal or total coliform bacteria. No significant statistical correlation could be determined between the occurrence of bacterial indicators and the presence of viruses. This study raises serious doubts about the validity of the indicator bacteria concept to predict the virological quality of water, particularly in countries with a high incidence of enteric viral disease.     

Bacteriological, virological and chemical evaluation of a wastewater-aquaculture system

Levels of fecal coliforms (FC), fecal streptococci (FS), Salmonella spp and enteric viruses were monitored in the water, sediment and fish in experimental wastewater-fish ponds near Benton, Arkansas, U.S.A. Concentrations of five heavy metals were also monitored in the fish and wastewater. Concentrations of indicator bacteria were reduced by as much as 99.7% through the series of six ponds which had a calculated total retention time of 72 days. Two filter-feeding species of Chinese carp, Hypophthalmichthys molitrix (silver carp) and Aristichthys nobilis (bighead carp), grown in the last three ponds accumulated FC and FS in their digestive tracts and skin at levels as great or greater than in the surrounding water and sediment. Only low levels of FC and FS were found in the fish muscle tissue (maximum of 25 FS per 100 g) even when concentrations of bacteria in the gut exceeded 10⁵ per 100 g. Concentrations of bacteria in the water and sediment were not good predictors of concentrations in the fish. No Salmonella and no enteric viruses were isolated from the fish, but this lack of isolates was attributed to the extremely low levels which were present in the influent wastewater. Higher levels of copper and mercury were found in the fish flesh than in the surrounding water, with three of eleven fish samples containing higher than acceptable levels of mercury in the edible portion. Based on the efficiency of wastewater treatment, an aquaculture system using silver and bighead carp was judged to be a viable treatment system for domestic sewage resulting in a product suitable for animal or human consumption if proper precautions are taken in harvesting and processing the fish.     

North and south american studies on the potential of coliphage as a water quality indicator

Studies were undertaken to assess the potential of coliphages to be used universally as water quality indicators and more specifically as health hazard indicators. Data were obtained from three water bodies, a northern Canadian River, inshore water samples from Lake Ontario and from marine beaches in Brazil. Data from this two continent, three water body study indicated (a) that within location fecal coliform and coliphages are positively correlated, (b) coliphage values can be predicted by using fecal coliform MPN, fecal streptococci MF and E. coli MF data and (c) a water quality guideline of 20 coliphage/100 ml for recreational fresh waters is proposed.     

Identification of the sources of fecal coliforms in an urban watershed using antibiotic resistance analysis

Bacteria such as fecal coliforms are used as indicators of fecal pollution in natural waters. These bacteria are found in the feces of most wild and domestic animals and thus provide no information as to the source of fecal contamination, yet identification of indicator bacteria sources allows improved risk assessment, remediation, and total daily maximum load (TDML) assessment of environmental waters. This bacterial source tracking study was initiated in order to identify the dominant source(s) of fecal contamination in the urban watershed of Stevenson Creek in Clearwater, Florida. Five sites that represent areas where routine monitoring has previously shown high levels of fecal coliforms were sampled over 7 months. Fecal coliforms were enumerated by membrane filtration, and antibiotic resistance analysis was used to "fingerprint" a subset of randomly selected isolates and statistically match them to fingerprints of fecal coliforms from known sources (the library). A field test of the classification accuracy of the library was carried out by isolating fecal coliforms from the soil and waters surrounding a failing onsite wastewater treatment and disposal system (OSTDS). The vast majority of the isolates were classified into the human category. The major sources of fecal pollution in Stevenson Creek over the course of the study were wild animal, human, and, to a lesser extent, dog. Overall, wild animal feces were identified as the dominant source when fecal coliform levels were high, but when fecal coliform levels were low, the dominant source was identified as human. The results of this study demonstrate that the sources of fecal indicator bacteria within one urban watershed can vary substantially over temporal and spatial distances.     

The ecology of "fecal indicator" bacteria commonly found in pulp and paper mill water systems

Coliform bacteria have long been used to indicate fecal contamination of water and thus a health hazard. In this study, the in-mill water and external effluent treatment systems of seven typical Canadian pulp and paper mills were all shown to support the growth of numerous coliforms, especially Klebsiella Spp., Escherichia coli. Enterobacter spp., and Citrobacter spp. In all mills and most sampled locations, klebsiellas were the predominant coliforms. Although all but one of the mills had no sewage input and most disinfected their feed (input) water, all contained the most typical fecal indicator bacterium, E. coli. Many of the mill coliforms were classified as fecal coliforms by standard "MPN" and metabolic tests, but this was shown to be due to their thermotolerance, not their origin. Mill coliforms were shown not to be just simple transients from feedwater or furnish (wood), but to be continuously growing, especially in some of the primary clarifiers. Isolated mill coliforms grew very well on a sterilized raw combined mill effluent. The fecal streptococci (enterococci), alternative indicators of fecal health hazards, were common in all mills in the absence of sewage. Ten strains of E. coli isolated from four mills were all shown to be non-toxigenic strains of harmless serotypes. No salmonellas were found. Therefore, the use of total coliform, fecal coliform, enterococci, or E. coli counts as indicators of fecal contamination, and thus of health hazard in pulp and paper mill effluents or biosolids (sludges) known to be free of fecal input is invalid.