Survival of coliform bacteria in sewage sludge applied to a forest clearcut and potential movement into groundwater.

Anaerobically digested dewatered sludge (10 to 15 cm thick) was applied to a forest clearcut as a fertilizer source in northwest Washington on gravelly glacial outwash soil. This sludge is not microbiologically sterile and may contain pathogenic organisms. Fecal coliform bacterial counts in sludge applied in summer (July) fell from 1.08 X 10(5) to 358/g in 204 days and to 0/g in 267 days. Dieoff appeared more rapid in winter (January)-applied sludge, when colnts fell from 1.2 X 10(5) to 20/g in 162 days. Initial death rates were related to sludge temperature, moisture, pH, physical composition, and microbial competition. Aftergrowth of fecal coliforms occurred in warm summer and fall months, but counts were of similar magnitude to background levels in forest soils, where a maximum count of 54/g was recorded. Total coliform counts in fresh sludge ranged from 1.4 X 10(4) to 1.9 X 10(6)/g. Numbers stabilized at 10(3) to 10(4)/g in spring, fall, and summer, with lower numbers in winter. Both total and fecal bacteria moved from the sludge to the soil beneath, but few penetrated past the first 5 cm. The soil acts as an effective biological filter. Few fecal coliform bacteria were recorded in the groundwater, generally being less than 5/100 ml and mostly 0/100 ml. A maximum count of 52/100 ml was recorded. Groundwater contamination from vertical movement of potential pathogens appears unlikely, but hazards from surface runoff and direct handling in the first year may arise.     

Seasonal Variations in Survival of Indicator Bacteria in Soil and Their Contribution to Storm-water Pollution

Survival of a fecal coliform (Escherichia coli) and a fecal streptococcus (Streptococcus faecalis var. liquifaciens) was studied through several years at shaded and exposed outdoor soil plots. Death rates for both organisms were calculated for the different seasons at both sites. The 90% reduction times for the fecal coliform ranged from 3.3 days in summer to 13.4 days in autumn. For the fecal streptococcus, 90% reduction times were from 2.7 days in summer to 20.1 days in winter. During summer, the fecal coliform survived slightly longer than the fecal streptococcus; during autumn, survival was the same; and in spring and winter the fecal streptococcus survived much longer than the fecal coliform. Both organisms were isolated from storm-water runoff collected below a sampling site when counts were sufficiently high in soil. Isolation was more frequent during prolonged rains, lasting up to 10 days, than during short rain storms. There was evidence of aftergrowth of nonfecal coliforms in the soil as a result of temperature and rainfall variations. Such aftergrowth may contribute to variations in bacterial count of storm-water runoff which have no relation to the sanitary history of the drainage area.     

Bacteriological quality of crops irrigated with wastewater in the Xochimilco plots, Mexico City, Mexico

Xochimilco county plots (Mexico City), one of the most fertile agricultural areas in the Valley of Mexico, produce a large portion of the fresh vegetables consumed in the city. These plots are generally irrigated with domestic wastewater, and for this reason, it was deemed important to examine and evaluate the bacteriological quality of the water, soil, and vegetables from these plots that are harvested and marketed. The soils were also examined for the classical parameters such as nitrates, ammonia, etc., and organic matter and texture. The crops selected for this study were radishes, spinach, lettuce, parsley, and celery because they are usually consumed raw. The highest bacterial counts were encountered in leafy vegetables, i.e., spinach (8,700 for total coliform and 2,400 for fecal coliform) and lettuce (37,000 for total coliform and 3,600 for fecal coliform). Statistically significant differences in bacterial counts between rinsed and unrinsed edible portions of the crops were observed even in rinsed vegetables, and high densities of fecal coliform were detected, indicating that their consumption represents a potential health hazard. The total coliform values found in irrigation water ranged from 4 X 10(4) to 29 X 10(4), and for fecal coliform the values ranged from 5 X 10(2) to 30 X 10(2).     

R factors in coliform-fecal coliform sewage flora of the prairies and Northwest Territories of Canada.

Coliform and fecal coliform populations found in the raw sewages and final sewage effluents of the prairie provinces and the Northwest Territories were examined for antibiotic resistance and the possession of R factors. It was determined that 8.91% of the total coliform and 10.80% of the fecal coliform populations carried R factors. The following numbers of combinations of R determinants were found: 39 in the Escherichia coli population, 6 in the Citrobacter population, 20 in the Enterobacter populations, 10 in the Klebsiella populations, and 11 in the Aeromonas populations. The maximum number of R determinants transferable simultaneously was seven; organisms with R factors containing determinants for chloramphenicol usually contained determinants for ampicillin. Of the coliform and fecal coliform populations, 2 to 4% were resistant to chloramphenicol in some provinces, and from 17 to 30% of the populations were resistant to three or more antibiotics. It was calculated that coliforms containing R factors in the raw sewage reached population levels of 1.5 X 10(7)/100 ml, and fecal coliforms containing R factors reached population levels of 8.6 X 10(5) ml. Final effluent discharges to the receiving environment contained R factor-containing coliform and fecal coliform populations of 3.1 X 10(4)/100 ml and 5.8 X 10(2)/100 ml, respectively. The incidence of bacteria containing R factors in sewage appears to be increasing with time, and their removal from sewage before discharge to the receiving environment is desirable. Consideration of data on bacteria with R factors should be made in future water quality deliberations and in discharge regulations.     

Persistence and distribution of pollution indicator bacteria on land used for disposal of piggery effluent.

Numbers of pollution indicator bacteria (fecal coliforms and fecal streptococci) were assessed on land to which effluent from intensively housed pigs had been applied. Topsoil (to a 30-mm depth) was found to provide a more favorable environment for fecal coliform persistence than was pasture or subsoil. Times required for a 90% reduction in number (T90) in topsoil (calculated by linear regression of log counts obtained in a 6-week period after effluent application) ranged from 7 to 20 days (mean T90, 11 days). T90 values for fecal coliforms fell within this range irrespective of the season of application and for a number of soil types and climatic conditions. The range in die-off times was encountered irrespective of the fecal coliform count in the applied effluent or the application regimen (125 to 1,000 kg of elemental nitrogen in the form of effluent per ha; return periods, 3 to 12 months). Autumn and winter conditions were conducive to the persistence of a survivor tail of these bacteria at 10(1) to 10(3) cells per g of topsoil. Fecal streptococci survived similarly on soil and pasture (T90, ca. 14 days) and appeared slightly more suited to survival in the environment than did fecal coliforms. Contamination of subsoils after effluent applications occurred at a rate well in excess of the infiltration capacity of the soil, presumably by percolation of the effluent through soil cracks. Contamination levels of subsoils in the experimental area generally remained low.     

Effect of waterfowl (Anas platyrhynchos) on indicator bacteria populations in a recreational lake Madison, Wisconsin.

A public swimming beach in Madison, wis., experienced intermittent high fecal coliform counts during the late summer and early fall of 1978. Public health officials closed the beach on a number of occasions. A public health survey identified a combination of waterfowl wastes and meteorological events as the explanation for the high bacteria counts. Fecal coliform bacteria were deposited by mallard ducks and multiplied in the beach sands. The bacteria were subsequently transported into the lake and resulted in high fecal coliform counts in the swimming area.     

Effect of waterfowl (Anas platyrhynchos) on indicator bacteria populations in a recreational lake Madison, Wisconsin.

A public swimming beach in Madison, wis., experienced intermittent high fecal coliform counts during the late summer and early fall of 1978. Public health officials closed the beach on a number of occasions. A public health survey identified a combination of waterfowl wastes and meteorological events as the explanation for the high bacteria counts. Fecal coliform bacteria were deposited by mallard ducks and multiplied in the beach sands. The bacteria were subsequently transported into the lake and resulted in high fecal coliform counts in the swimming area.     

Occurrence of Potential Pathogens in Water Containing Ornamental Fishes

The bacterial population of the water supplied with ornamental fish purchased from retail outlets was examined qualitatively and quantitatively. As many as 10(9) viable aerobic organisms per 100 ml were present, with fecal coliform counts as high as 10(5) per 100 ml. Citrobacter, Escherichia, Pseudomonas, and Vibrio were isolated from 75% or more of the samples, whereas Aeromonas, Alcaligenes, Enterobacter, Flavobacterium, and Streptococcus were isolated from 45 to 65% of the samples. Pseudomonas aeruginosa, Edwardsiella tarda, and Klebsiella pneumoniae were also isolated.     

Coliforms removal in two UASB + ASP based systems

Two full scale up-flow anaerobic sludge blanket (UASB) reactor with activated sludge process based STPs (43 Ml d⁻¹ and 100 Ml d⁻¹ capacities) purely for municipal wastewater having two different aeration systems (surface and diffused) were continuously evaluated for one year. Total coliforms (TC) and fecal coliforms (FC) were measured along with other performance monitoring parameters in raw sewage, UASB effluents, and in final effluents. The concentration of total and fecal coliforms ranged from 9.3 × 10⁶ to 1.2 × 10¹⁴ MPN/100 ml and 9.3 × 10⁶ to 2.4 × 10¹³ MPN/100 ml respectively in raw sewage. At both the plants, it was observed that finally treated effluent still contained significant number of total coliforms (5.71 × 10⁵ and 6.7 × 10⁵) and fecal coliforms (3.67 × 10⁵ and 2.2 × 10⁵) after overall removal of 99.9%, which is still greater than the permissible limit of 1000 MPN/100 ml and needs further treatment. Seasonal variation indicated the effective removal of coliforms in raw sewage was in summer. Fecal coliforms are highly correlated with phosphorous and total coliform concentrations.     

Evaluation of porcelain cup soil water samplers for bacteriological sampling

The validity of obtaining soil water for fecal coliform analyses by porcelain cup soil water samplers was examined. Numbers from samples of manure slurry drawn through porcelain cups were reduced 100- to 10,000,000-fold compared to numbers obtained from the external manure slurry, and 65% of the cups yielded coliform-free samples. Fecal coliforms adsorbed to cups apparently were released, thus influencing the counts of subsequent samples. Fecal coliforms persisted in soil water samplers buried in soil and thus could significantly influence the coliform counts of water samples obtained a month later. These studies indicate that porcelain cup soil water samplers do not yield valid water samples for fecal coliform analyses.     

Cultural and environmental factors affecting the longevity of Escherichia coli in Histosols.

The survival of Escherichia coli in organic soils (Histosols) was examined. The death rate of this organism in Pahokee muck was less than that observed in Pompano fine sand. The number of viable E. coli cells found in the muck was approximately threefold greater than that found in the sand following 8 days of incubation. The initial population of the coliform affected the death rate. The rate of loss of viability varied 100-fold when the population size decreased from 2.5 x 10(7) to 3.4 x 10(4). Other factors affecting the viability of E. coli in muck were aerobic versus anaerobic growth of the organism and moist versus flooded conditions in the soil. The greatest survival of the coliform was noted with anaerobically grown cells amended to flooded soil. That the observed decrease in E. coli viability in soil was the result of biotic factors was demonstrated with amendment of sterile soil with E. coli. When 1.1 x 10(5) bacteria per g of soil were added to sterile muck, a population of 3.0 x 10(7) organisms per g of soil developed over a 10-day period. The role of the protozoa in eradication of the coliform from the muck was indicated by a sixfold increase in the protozoan population in natural soil amended with E. coli. Higher organic matter content in a Histosol compared with a mineral soil resulted in an increased survival of the fecal coliforms. Biotic factors are instrumental in the decline in coliform populations, but the potential for growth of the coliform in the organic soil could extend the survival of the organism.     

Impact of Mount St. Helens Eruption on Bacteriology of Lakes in the Blast Zone

Lakes lying within the blast zone of Mount St. Helens showed dramatic increases in heterotrophic bacterial numbers after the eruption of 18 May 1980. The total microscopic counts of bacteria in some of the most severely affected lakes were more than 10⁷ cells per ml, an order of magnitude above the counts in outlying control lakes. Likewise, the numbers of viable bacteria reached levels of more than 10⁶ cells per ml, compated with fewer than 10⁴ cells per ml in control lakes. The CPS medium used for enumeration provided growth of up to 81.5% of the bacteria during sampling of one of the blast zone lakes. The high numbers of bacteria and the efficacy of the viable enumeration procedure are evidence that the lakes have been transformed rapidly from oligotrophy to eutrophy due to the eruption and its aftermath. Organic material leached from the devastated forest vegetation is thought to be responsible for the enrichment of heterotrophs. Total coliform bacteria were found in all of the blast zone lakes, and some lakes contained fecal coliform bacteria. Klebsiella pneumoniae was the predominant total coliform and was also identified as one of the fecal coliform bacteria, although Escherichia coli was the predominant species in that category. Our data indicate that bacterial populations peaked in the outer blast zone lakes in the summer of 1980 and in most of the inner lakes during the summer of 1981.     

Effect of environmental factors on the relationship between concentrations of coprostanol and fecal indicator bacteria in tropical (Mekong Delta) and temperate (Tokyo) freshwaters

A reliable assessment of microbial indicators of fecal pollution (total coliform, Escherichia coli, and fecal streptococcus) is critical in tropical environments. Therefore, we investigated the relationship between concentrations of indicator bacteria and a chemical indicator, coprostanol (5beta-cholestan-3beta-ol), in tropical and temperate regions. Water samples were collected from the Mekong Delta, Vietnam, during wet and dry seasons, and from Tokyo, Japan, during summer, the aftermath of a typhoon, and winter. During the wet season in the Mekong Delta, higher bacterial densities were observed in rivers, probably due to the higher bacterial inputs from soil particles with runoff. In Tokyo, higher bacterial densities were usually observed during summer, followed by those in the typhoon aftermath and winter. A strong logarithmic correlation between the concentrations of E. coli and coprostanol was demonstrated in all surveys. Distinctive seasonal fluctuations were observed, as concentrations of coprostanol corresponding to 1,000 CFU of E. coli/100 ml were at their lowest during the wet season in the Mekong Delta and the typhoon aftermath in Tokyo (30 ng/liter), followed by the dry season in the Mekong Delta and the summer in Tokyo (100 ng/liter), and they were much higher during the winter in Tokyo (400 ng/liter). These results suggested that E. coli is a specific indicator of fecal contamination in both tropical and temperate regions but that the densities are affected by elevated water temperature and input from runoff of soil particles. The concurrent determination of E. coli and coprostanol concentrations could provide a possible approach to assessing the reliability of fecal pollution monitoring data.     

Isolation of Klebsielleae from within living wood.

Previous studies from this laboratory have documented the presence of coliform bacteria emanating from wooden reservoirs containing finished drinking water. Coliforms were identified as Klebsiella pneumoniae and Enterobacter spp. In the present report, evidence is presented which suggests that the origin of these coliforms is from the wood used to construct the reservoirs. In liquid expressed from freshly cut redwood, total bacterial counts in the range of 10(5) to 10(6)/ml were commonly observed. When present, coliform counts were over 10(3)/ml of expressed liquid. E. agglomerans was the most prevalent coliform present, but Klebsiella was isolated from freshly cut logs. Citrobacter freundii was also occasionally isolated. No fecal coliform-positive Klebsiella were obtained from any of the samples. Highest total bacteria and coliform counts were observed in sapwood specimens. Coliforms were present throughout sapwood as evidenced by contact plating serial sections of freshly cut wood. Scanning electron micrographs illustrate the presence of bacterial colonies within sapwood tracheids. Other wood species also contained coliform bacteria but in numbers lower than found in redwood.     

Isolation of Klebsielleae from within living wood.

Previous studies from this laboratory have documented the presence of coliform bacteria emanating from wooden reservoirs containing finished drinking water. Coliforms were identified as Klebsiella pneumoniae and Enterobacter spp. In the present report, evidence is presented which suggests that the origin of these coliforms is from the wood used to construct the reservoirs. In liquid expressed from freshly cut redwood, total bacterial counts in the range of 10(5) to 10(6)/ml were commonly observed. When present, coliform counts were over 10(3)/ml of expressed liquid. E. agglomerans was the most prevalent coliform present, but Klebsiella was isolated from freshly cut logs. Citrobacter freundii was also occasionally isolated. No fecal coliform-positive Klebsiella were obtained from any of the samples. Highest total bacteria and coliform counts were observed in sapwood specimens. Coliforms were present throughout sapwood as evidenced by contact plating serial sections of freshly cut wood. Scanning electron micrographs illustrate the presence of bacterial colonies within sapwood tracheids. Other wood species also contained coliform bacteria but in numbers lower than found in redwood.     

Coliform concentrations in Lake Erie — 1966 to 1970

Total coliform concentrations obtained from cruises of Lake Erie, conducted from the Canada Centre for Inland Waters between 1966 and 1970, have been analysed to determine the spatial and temporal distribution of total coliforms in the lake. Although year to year comparisons were made on a qualitative level due to the limitations in the data, some consistent seasonal features of the spatial distribution are evident. The location of zones of higher concentration have been attributed to proximity to large urban areas and lake currents. The contribution of fecal coliforms to the total coliform concentration has been examined with the conclusion that, at the low total coliform concentrations ( 30 per 100 ml) observed in much of the lake, there is no relationship between the two concentrations but, over a broader range, fecal coliforms increase with total coliform concentrations and, at very high total coliform concentrations (1000 per 100 ml), fecal coliforms are always present. The analysis demonstrates 1) the value of examining observed and fitted distributions to compare subsets of a large collection of data and 2) a clustering procedure which is generally applicable to data consisting of counts.     

Effect of Environmental Factors on the Relationship between Concentrations of Coprostanol and Fecal Indicator Bacteria in Tropical (Mekong Delta) and Temperate (Tokyo) Freshwaters

A reliable assessment of microbial indicators of fecal pollution (total coliform, Escherichia coli, and fecal streptococcus) is critical in tropical environments. Therefore, we investigated the relationship between concentrations of indicator bacteria and a chemical indicator, coprostanol (5β-cholestan-3β-ol), in tropical and temperate regions. Water samples were collected from the Mekong Delta, Vietnam, during wet and dry seasons, and from Tokyo, Japan, during summer, the aftermath of a typhoon, and winter. During the wet season in the Mekong Delta, higher bacterial densities were observed in rivers, probably due to the higher bacterial inputs from soil particles with runoff. In Tokyo, higher bacterial densities were usually observed during summer, followed by those in the typhoon aftermath and winter. A strong logarithmic correlation between the concentrations of E. coli and coprostanol was demonstrated in all surveys. Distinctive seasonal fluctuations were observed, as concentrations of coprostanol corresponding to 1,000 CFU of E. coli/100 ml were at their lowest during the wet season in the Mekong Delta and the typhoon aftermath in Tokyo (30 ng/liter), followed by the dry season in the Mekong Delta and the summer in Tokyo (100 ng/liter), and they were much higher during the winter in Tokyo (400 ng/liter). These results suggested that E. coli is a specific indicator of fecal contamination in both tropical and temperate regions but that the densities are affected by elevated water temperature and input from runoff of soil particles. The concurrent determination of E. coli and coprostanol concentrations could provide a possible approach to assessing the reliability of fecal pollution monitoring data.     

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.     

Cadmium tolerance and antibiotic resistance in Escherichia coli isolated from waste stabilization ponds

The incidence pattern of cadmium tolerance and antibiotics resistance by Escherichia coli was examined periodically from the samples of water, sludge and intestine of fish raised in waste stabilization ponds in a sewage treatment plant. Samples of water and sludge were collected from all the selected ponds and were monitored for total counts of fecal coliform (FC), total coliform (TC) and the population of Escherichia coli, which was also obtained from the intestine of fishes. Total counts of both FC and TC as well as counts of E. coli were markedly reduced from the facultative pond to the last maturation pond. Tolerance limit to cadmium by E. coli tended to decline as the distance of the sewage effluent from the source increased; the effective lethal concentration of cadmium ranged from 0.1 mM in split chamber to 0.05 mM in first maturation pond. E. coli isolated from water, sludge and fish gut were sensitive to seven out of ten antibiotics tested. It appears that holistic functions mediated through the mutualistic growth of micro algae and heterotrophic bacteria in the waste stabilization ponds were responsible for the promotion of water quality and significant reduction of coliform along the sewage effluent gradient.     

Evaluation of Standard and Modified M-FC, MacConkey, and Teepol Media for Membrane Filtration Counting of Fecal Coliforms in Water

MacConkey agar, standard M-FC agar, M-FC agar without rosolic acid, M-FC agar with a resuscitation top layer, Teepol agar, and pads saturated with Teepol broth, were evaluated as growth media for membrane filtration counting of fecal coliform bacteria in water. In comparative tests on 312 samples of water from a wide variety of sources, including chlorinated effluents, M-FC agar without rosolic acid proved the medium of choice because it generally yielded the highest counts, was readily obtainable, easy to prepare and handle, and yielded clearly recognizable fecal coliform colonies. Identification of 1,139 fecal coliform isolates showed that fecal coliform tests cannot be used to enumerate Escherichia coli because the incidence of E. coli among fecal coliforms varied from an average of 51% for river water to 93% for an activated sludge effluent after chlorination. The incidence of Klebsiella pneumoniae among fecal coliforms varied from an average of 4% for the activated sludge effluent after chlorination to 32% for the river water. The advantages of a standard membrane filtration procedure for routine counting of fecal coliforms in water using M-FC agar without rosolic acid as growth medium, in the absence of preincubation or resuscitation steps, are outlined.