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PubMed2006

Male-specific coliphages as indicators of thermal inactivation of pathogens in biosolids.

Male-specific (F+) coliphages have been proposed as a candidate indicator of fecal contamination and of virus reduction in waste treatment. However, in this and earlier work with a laboratory thermophilic anaerobic digester, a heat-resistant fraction of F+ coliphage populations indigenous to municipal wastewater and sludge was evident. We therefore isolated coliphages from municipal wastewater sludge and from biosolid samples after thermophilic anaerobic digestion to evaluate the susceptibility of specific groups to thermal inactivation. Similar numbers of F+ DNA and F+ RNA coliphages were found in untreated sludge, but the majority of isolates in digested biosolids were group I F+ RNA phages. Separate experiments on individual isolates at 53 degrees C confirmed the apparent heat resistance of group I F+ RNA coliphages as well as the susceptibility of group III F+ RNA coliphages. Although few F+ DNA coliphages were recovered from the treated biosolid samples, thermal inactivation experiments indicated heat resistance similar to that of group I F+ RNA phages. Hence, F+ DNA coliphage reductions during thermophilic anaerobic digestion are probably related to mechanisms other than thermal inactivation. Further studies should focus on the group III F+ RNA coliphages as potential indicators of reductions of heat-resistant pathogens in thermal processes for sludge treatment.

Anaerobiosis↗

PubMed2006

Transport of coliphage in the presence and absence of manure suspension.

Mechanisms of coliphage transport and fate in the presence and absence of manure suspension were studied in saturated column experiments. In the presence of manure suspension, little inactivation of indigenous somatic coliphage occurred and the transport was controlled by deposition. The deposition followed a power law distribution with depth, and the magnitude increased with decreasing sand size. Comparison of the cumulative size distribution of manure components in the suspension initially and after passage through sand, suggested that particles retained by mechanical filtration and/or straining decreased the effective pore size and potentially induced straining of the somatic coliphage. A 2-site kinetic deposition model was used to estimate the magnitudes of attachment and straining in the presence of manure suspension, and provided a good description of the data. Modeling results indicated that straining accounted for 16 to 42% of the deposited somatic coliphage, and that both straining and attachment increased with decreasing sand size due to smaller pores and higher surface area, respectively. In the absence of manure suspension, phiX174 (a representative somatic coliphage) and MS2 (a male-specific RNA coliphage) transport was controlled by inactivation induced by the solid phase. This conclusion was based on comparison of coliphage transport behavior at 5 and 20 degrees C, mass balance information, and numerical modeling. Comparison of somatic coliphage transport data in the presence and absence of manure suspension revealed much higher effluent concentrations in the presence of manure. This difference was attributed to lower inactivation and higher detachment rates. The observed coliphage transport behavior suggests that survival of viruses may be extended in the presence of manure suspensions, and that transport studies conducted in the absence of manure suspension may not accurately characterize the transport potential of viruses in manure-contaminated environments.

Animals↗

PubMed1986

Methodology for enumeration of coliphages in foods.

The effects of eluent composition, pH, and chaotropic agents on the recovery of T2, MS2, and indigenous coliphages from various foods were investigated. Additionally, methods of sample suspension and clarification were evaluated for coliphage recovery and application to various foods. Clarified sample suspensions were assayed for coliphages with a modified agar layer technique and appropriate Escherichia coli hosts. Centrifugation and polypropylene mesh filtration were more rapid and effective than glass wool filtration for clarification of sample suspensions and subsequent recovery of coliphages. Blending, stomaching, and shaking procedures were generally comparable for sample liquefaction and release of coliphages from foods. Complex basal eluents, EC medium and 1% casein, were generally more effective than a less complex eluent, phosphate buffer, for elution of coliphages from foods. For most foods, incorporation of sodium chloride or chaotropic agents, i.e., sodium trichloroacetate, urea, Tween 80, Triton X-100, and sodium nitrate, into basal eluents did not enhance recovery of coliphages. Indigenous coliphage recovery was not affected by sample suspension pH over a range of 6.0 to 9.0. With an optimal procedure, i.e., EC medium eluent, blending, and centrifugation, the recovery of T2 and MS2 ranged from 48 to 81% and from 58 to 100%, respectively, depending on the food type.

Animals↗

PubMed1988

Animal viruses, coliphages, and bacteria in aerosols and wastewater at a spray irrigation site.

Aerosol samples collected at the Muskegon County Wastewater Management System Number 1 spray irrigation site in Michigan by using the Army prototype XM2 Biological Sampler/Collector were examined for the presence of animal viruses, coliphages, and bacteria. Air samples, collected in Earle lactalbumen hydrolysate, and wastewater samples were filtered through a 0.45- and 1.2-micron membrane filter sandwich, pretreated with 10% beef extract (pH 7.0), and assayed for animal viruses by the plaque method on Buffalo green monkey kidney cells. Untreated air and wastewater samples were assayed for coliphages by the soft agar overlay method with three Escherichia coli hosts (ATCC 13706, 15597, and 11303) and for bacteria by the heterotrophic plate count method. Filtered air samples were assayed for coliphages by the most-probable-number method with the same three hosts. Although no animal viruses were detected in the aerosol samples, coliphages and bacteria were recovered. E. coli ATCC 13706 coliphage were recovered more often and in greater numbers than either of the other two types of coliphages. Concentrations of animal viruses, coliphages, and bacteria detected in the raw influent decreased as the wastewater was aerated and stored in the lagoons. No animal viruses were detected in the wastewater at the pump station just before distribution to the spray irrigation rigs. The most-probable-number method was more sensitive and consistent than the overlay procedure in detecting low levels of coliphages in air samples.

Aerosols↗

PubMed2005

Barriers to coliphage infection of commensal intestinal flora of laboratory mice.

BACKGROUND: Growth characteristics of coliphage viruses indicate that they are adapted to live with their Eschericia coli hosts in the intestinal tract. However, coliphage experimentally introduced by ingestion persist only transiently if at all in the gut of humans and other animals. This study attempted to identify the barriers to long term establishment of exogenous coliphage in the gastrointestinal (GI) tracts of laboratory mice. Intestinal contents were screened for the presence of coliphage and host bacteria, and strains of E. coli bacteria from different segments of the GI tract were tested for susceptibility to six common laboratory coliphages. RESULTS: Contrary to expectations, coliphage were not evident in the GI tracts of laboratory mice, although they were occasionally detected in feces. Commensal flora showed extreme variability within groups of mice despite identical handling and diet. Less than 20% of 48 mice tested carried E. coli in their gut, and of 22 commensal E. coli strains isolated and tested, 59% were completely resistant to infection by lambda, M13, P1, T4, T7, and PhiX174 coliphage. Lysogeny could not be demonstrated in the commensal strains as mitomycin C failed to induce detectable phage. Pre-existing immunity to phages was not evident as sera and fecal washes did not contain significant antibody titers to six laboratory phage types. CONCLUSION: Lack of sufficient susceptible host bacteria seems to be the most likely barrier to establishment of new coliphage infections in the mouse gut.

Animals↗

PubMed2004

Development and evaluation of methods to detect coliphages in large volumes of water.

New and improved methods have been developed to detect somatic and male-specific coliphages in large volumes of water by single agar layer (SAL), enrichment and membrane filter methods. Somatic coliphages were detected efficiently on E. coli hosts C and CN13, male-specific coliphages were detected more efficiently on E. coli Famp than on Salmonella typhimurium WG49 and both types of coliphages were detected simultaneously on E. coli C3000. For water volumes of up to 100 ml, the SAL method was efficient and reliable. For water volumes of <1 L and as many as 10 multiple 1 L volumes, the enrichment method was efficient in detecting very low numbers of coliphages. Membrane filter methods, in which coliphages were adsorbed to and eluted from filters, also were relatively efficient, but they were less efficient than SAL and enrichment methods and were considered to be more cumbersome. For filter adsorption-elution methods, coliphage recoveries were most efficient for cellulose ester filters, less efficient for electropositive 1 MDS filters and least efficient for a direct membrane filter method. Overall, the enrichment method was preferred because of its ability to easily and rapidly detect low levels of coliphages in large sample volumes by either presence-absence or most probable number quantification.

Cellulose↗

PubMed1983

Relationship between the occurrence of coliphages and E. coli in Danish marine bathing areas.

During 2 bathing seasons, 1978 and 1979, water samples were collected with intervals of 2 weeks from 17 marine bathing areas situated on the coast of the island of Fyn. Quantitative examinations for coliphages and fecal coliforms were carried out. Coliphages were present in approximately 50% of the samples. 75% of the positive samples contained less than or equal to 10 coliphages per 100 ml. 6 samples were found with greater than 100 coliphages per 100 ml. A positive correlation between coliphage and fecal coliform concentrations was demonstrated. The ratios of fecal coliforms to coliphages varied only little between individual sampling stations. The mean ratio was 1.9 and 2.0 for the 2 sampling periods. The mean concentration of fecal coliforms was gradually increasing from 2.4/100 ml in May to 7.2/100 ml in September. Conversely the coliphage concentrations were quite uniform during the bathing seasons. As a result of those 2 trends, the fecal coliform/coliphages ratio increased during the seasons from 0.86 to 3.13. Finally, the potential value of coliphages as an indicator of pathogenic microorganisms in marine bathing areas is discussed.

Bathing Beaches↗

PubMed1990

Tissue distribution of a coliphage and Escherichia coli in mussels after contamination and depuration.

Experiments were undertaken to determine the tissue distribution of Escherichia coli and a coliphage after contamination of the common mussel (Mytilus edulis). Mussels were contaminated with high levels of feces-associated E. coli and a 22-nm icosahedral coliphage over a 2-day period in a flowing-seawater facility. After contamination, individual tissues were carefully dissected and assayed for E. coli and the coliphage. Contaminated mussels were also analyzed to determine the tissue distribution of the contaminants after 24- and 48-h depuration periods. The majority of each contaminant was located in the digestive tract (94 and 89% of E. coli and coliphage, respectively). Decreasing concentrations were found in the gills and labial palps, foot and muscles, mantle lobes, and hemolymph. Our results indicate that contamination above levels in water occurred only in the digestive tract. Contaminated mussels were depurated in a commercial-scale recirculating UV depuration system over a 48-h period. The percent reductions of E. coli occurred in the following order: digestive tract, hemolymph, foot and muscles, mantle lobes, and gills and labial palps. The percent reductions of the coliphage were different, occurring in the following order: hemolymph, foot and muscles, gills and labial palps, mantle lobes, and digestive tract. Our results clearly demonstrate that E. coli and the coliphage are differentially eliminated from the digestive tract. The two microorganisms are eliminated at similar rates from the remaining tissues. Our results also clearly show that the most significant coliphage retention after depuration for 48 h is in the digestive tract. Thus, conventional depuration practices are inappropriate for efficient virus elimination from mussels.

Animals↗

PubMed1988

[Quantitative studies of the elimination of coliphages and other fecal indicators during wastewater treatment].

Concentrations of coliphages, coliforms, enterococci and fluorescent Pseudomonas were monitored in several wastewater purification steps of the treatment plant Wolfenbüttel during one year. Their number varied widely during the investigation period, but was independent of seasons. In the course of sewage treatment, including primary settling, activated sludge purification, simultaneous precipitation, trickling filters and oxidation pond, the concentration of indicators decreased gradually. The coliphages were most resistant, exhibiting only a decimal elimination value of 1.7 log10 units as compared to the bacterial indicators with elimination values ranging between 2.4 and 2.8 log10 units in the whole process. The most efficient purification step revealed to be the activated sludge procedure including simultaneous phosphate precipitation with iron hydroxides and sedimentation. On an average 1.7% of the coliphages present in raw sewage or 9.8.10.11 phages were discharged into the river Oker everyday, 0.64% remained in the sludge. Numbers of indicators in the water of the oxidation pond and those seeded into river water were continuously reduced during 3 days. Also in these laboratory experiments, the coliphages were more resistant than the bacteria, but no evidence was found to support the view that coliphages play a role in the reduction of the number of coliform bacteria. Even after addition of peptone which stimulated growth of E. coli the coliphages were inactivated more rapidly. The behaviour of coliphages during the purification process is compared with literature data about enteroviruses.

Bacteria↗

PubMed2003

Evaluation of Bacillus subtilis and coliphage MS2 as indicators of advanced water treatment efficiency.

The assessment of water treatment facilities for their efficiency using alternate indicators is of paramount importance. Current methods for assessing efficiency are limited by the specific characteristics of the microorganisms, such as their different sensitivities to disinfectants. A pilot study was carried out to compare different treatment scenarios for the future upgrade of the Sergio Cuevas Water Treatment plant (the largest in the Caribbean) in San Juan, Puerto Rico. The treatment units under investigation included a coagulation-flocculation-sedimentation unit, dual-media filters, micro-filtration units, intermediate ozone injection and contact columns as well as a biological filtration unit. The plant was challenged at different stages of treatment with Bacillus subtilis spores and MS2 coliphages in an attempt to test them as possible alternate indicators of treatment plant performance. These organisms were chosen because of their resistance to disinfection and desiccation, their low analysis costs and ease of detection. The removal of spores and coliphages by each treatment unit tested was calculated by seeding a known concentration (5-7 log10) of spores and coliphages and following the removal or disinfection rates. The seeded indicators were detected using traditional culture techniques. Ballasted clarification was shown to be highly efficient at removing 99.1% (approximately 3 log10) of the spores and 85.1% (approximately 0.86 log10) of MS2. Ozone treatment inactivated 80.37% (approximately 1.4 log10) spores and 99.95% (approximately 3.07 log10) coliphages. The coliphage inactivation rate obtained confirmed data obtained by previous studies indicating that MS2 was less resistant to ozonation than B subtilis spores. The membrane technology had the best efficiency in terms of physical removal of spores achieving over 99.9% (> 3 log10) removal. Coliphage removal mechanisms remain to be determined and will be a future focus of the study. Preliminary results indicate that aerobic spores and coliphages may be useful as indicators to determine the efficiency of different drinking water treatment technologies.

Bacillus subtilis↗

PubMed2003

Occurrence of coliphages in urban stormwater and their fate in stormwater management systems.

AIMS: To investigate the occurrence of coliphages in, and their removal from, urban stormwater. METHODS AND RESULTS: Inflow and outflow concentrations of somatic and f-specific RNA coliphages to two stormwater treatment systems were determined on 21 occasions over a period of 5 months. Somatic coliphages were detected in 19 (90%) of the constructed wetland inlet samples, 13 (62%) of the pond inlet samples, and less frequently at the outlets of the two systems. F-specific RNA coliphages were detected at the inlets but only occasionally at the pond outlet. Somatic coliphages were found to attach preferentially to particles <5 microm in size and persisted in the sediments of the two systems. CONCLUSIONS: Treatment systems providing conditions that are conducive to the settlement of fine particles may effectively remove sediment-bound coliphages and, therefore, possibly enteric viruses from stormwater. SIGNIFICANCE AND IMPACT OF THE STUDY: The results will aid the design of systems for effective removal of viral contaminants from urban stormwater.

Coliphages↗

PubMed1982

Evaluation of coliphage detection as a rapid indicator of water quality.

A rapid coliphage analysis technique for enumerating coliphages in natural waters has been evaluated by water quality laboratories located throughout the United States. Correlations were established between coliphages and coliforms in natural water systems. These correlations were highly significant. This relationship can thus be used to determine the number of fecal or total coliforms present in natural water samples based on an enumeration of coliphages. With this method, coliphages in natural water systems (containing greater than or equal to six coliphages per 100 ml) can be enumerated within 6 h.

Coliphages↗

PubMed1984

Coliphages as indicators of enteroviruses.

Coliphages were monitored in conjunction with indicator bacteria and enteroviruses in a drinking-water plant modified to reduce trihalomethane production. Coliphages could be detected in the source water by direct inoculation, and sufficient coliphages were detected in enterovirus concentrates to permit following the coliphage levels through different water treatment processes. The recovery efficiency by different filter types ranged from 1 to 53%. Statistical analysis of the data indicated that enterovirus isolates were better correlated with coliphages than with total coliforms, fecal coliforms, fecal streptococci, or standard plate count organisms. Coliphages were not detected in finished water.

Coliphages↗

PubMed1985

Comparison of selective media for assay of coliphages in sewage effluent and lake water.

Selective media, including EC medium, gram-negative broth, nutrient broth (with 0.05% sodium deoxycholate), and lactose broth (with 0.05% sodium deoxycholate), as well as nonselective nutrient and lactose broths, were compared for the enumeration of coliphages by the agar layer method from activated-sludge effluent and eutrophic-lake water from a lake receiving treated sewage effluent. Samples were plated directly or after chloroform treatment with Escherichia coli B, E. coli C, or a mixed host of both E. coli B and C. With the exception of gram-negative broth, direct assays of all samples with the selective media generally resulted in significantly higher (P less than 0.05) recoveries of coliphages than did assays of chloroform-treated samples with nutrient broth medium regardless of the host used. In addition, chloroform pretreatment resulted in decreased recovery of coliphages with each selective medium in most analyses. The highest recoveries of coliphages from all samples with each host, except lake water with E. coli C, were obtained by direct assay on EC medium. The selectivity of the EC and gram-negative media resulted in suppression of bacterial interference on direct assay plates comparable to that observed in nutrient agar medium with chloroform-treated samples. The use of certain selective media for the direct assay of environmental materials for coliphage may enhance the recovery of coliphages and obviate bacterial decontamination procedures.

Chloroform↗

PubMed1989

Differential depuration of poliovirus, Escherichia coli, and a coliphage by the common mussel, Mytilus edulis.

The elimination of sewage effluent-associated poliovirus, Escherichia coli, and a 22-nm icosahedral coliphage by the common mussel, Mytilus edulis, was studied. Both laboratory-and commercial-scale recirculating, UV depuration systems were used in this study. In the laboratory system, the logarithms of the poliovirus, E. coli, and coliphage levels were reduced by 1.86, 2.9, and 2.16, respectively, within 52 h of depuration. The relative patterns and rates of elimination of the three organisms suggest that they are eliminated from mussels by different mechanisms during depuration under suitable conditions. Poliovirus was not included in experiments undertaken in the commercial-scale depuration system. The differences in the relative rates and patterns of elimination were maintained for E. coli and coliphage in this system, with the logarithm of the E. coli levels being reduced by 3.18 and the logarithm of the coliphage levels being reduced by 0.87. The results from both depuration systems suggest that E. coli is an inappropriate indicator of the efficiency of virus elimination during depuration. The coliphage used appears to be a more representative indicator. Depuration under stressful conditions appeared to have a negligible affect on poliovirus and coliphage elimination rates from mussels. However, the rate and pattern of E. coli elimination were dramatically affected by these conditions. Therefore, monitoring E. coli counts might prove useful in ensuring that mussels are functioning well during depuration.

Animals↗

PubMed1991

Comparative inactivation of poliovirus type 3 and MS2 coliphage in demand-free phosphate buffer by using ozone.

MS2 coliphage (ATCC 15597-B1) has been proposed by the U.S. Environmental Protection Agency as a surrogate for enteric viruses to determine the engineering requirements of chemical disinfection systems on the basis of previous experience with chlorine. The objective of this study was to determine whether MS2 coliphage was a suitable indicator for the inactivation of enteric viruses when ozone disinfection systems were used. Bench-scale experiments were conducted in 2-liter-batch shrinking reactors containing ozone demand-free 0.05 M phosphate buffer (pH 6.9) at 22 degrees C. Ozone was added as a side stream from a concentrated stock solution. It was found that an ozone residual of less than 40 micrograms/liter at the end of 20 s inactivated greater than 99.99% of MS2 coliphage in the demand-free buffer. When MS2 was compared directly with poliovirus type 3 in paired experiments, 1.6 log units more inactivation was observed with MS2 coliphage than with poliovirus type 3. It was concluded that the use of MS2 coliphage as a surrogate organism for studies of enteric virus with ozone disinfection systems overestimated the inactivation of enteric viruses. It is recommended that the regulatory agencies evaluate their recommendations for using MS2 coliphage as an indicator of enteric viruses.

Buffers↗

PubMed1995

Genotyping male-specific RNA coliphages by hybridization with oligonucleotide probes.

F-specific (F+) RNA coliphages are prevalent in sewage and other fecal wastes of humans and animals. There are four antigenically distinct serogroups of F+ RNA coliphages, and those predominating in humans (groups II and III) differ from those predominating in animals (groups I and IV). Hence, it may be possible to distinguish between human and animal wastes by serotyping F+ RNA coliphage isolates. Because serotyping is laborious and requires scarce antiserum reagents, we investigated genotyping using synthetic oligonucleotide probes as an alternative approach to distinguishing the four groups of F+ RNA coliphages. Oligoprobes I, II, III, IV, A, and B were selected to detect group I, II, III, IV, I plus II, and III plus IV phages, respectively. Methods for phage transfer from zones of lysis on a host cell lawn to candidate membrane filters and fixation of genomic nucleic acid on the membranes were optimized. The oligoprobes, which were end labeled with digoxigenin, were applied in DNA-RNA hybridization, and hybrids were observed by colorimetric, immunoenzymatic detection. Of 203 isolates of F+ RNA coliphages from environmental samples of water, wastes, and shellfish, 99.5 and 96.6% could be classified into each group by serotyping and genotyping, respectively. Probes A and B correctly identified 100% of the isolates. On the basis of these results, this method for genotyping F+ RNA coliphages appears to be practical and reliable for typing isolates in field samples.

Animals↗

PubMed1991

Fate of human enteric viruses, coliphages, and Clostridium perfringens during drinking-water treatment.

The elimination of human enteric viruses, coliphages, and Clostridium perfringens was studied during a conventional complete drinking-water treatment process. The respective concentrations (geometric mean) of these microorganisms in 100-L samples of river water were, respectively, as follows: viruses, 79 mpniu (most probable number of infectious units) per 100 L, coliphages, 6565 pfu (plaque-forming units) per 100 L. and clostridia, 11,349 cfu (colony-forming units) per 100 L. After predisinfection, flocculation with alum, and settling, human enteric viruses were not detected in any of the 100-L samples (less than 4 mpniu/100 L), but coliphages were detected in 7 of 14 samples and clostridia in 15 of 16 samples. In filtered water samples, human enteric viruses were detected in 2 of 31 samples, coliphages in 10 of 33, and clostridia in 17 of 33. Finished water was free of human enteric viruses (0/162 samples), but coliphages were detected in one sample (1.5 pfu/100 L) and clostridia in three, at 1.0, 4.1, and 7.0 cfu/100 L. It thus appears that coliphages and clostridia, which are present in larger numbers than viruses in river water and which may have similar resistance to drinking-water treatments, may be useful for estimating the level of treatment attained when large volumes of water (1000 L or greater) are sampled.

Clostridium perfringens↗