Fecal Bacterial Populations Research Articles

Dietary galactooligosaccharides affect ileal and total-tract nutrient digestibility, ileal and fecal bacterial concentrations, and ileal fermentative characteristics of growing pigs.

The objective of this study was to evaluate dietary galactooligosaccharide (Gal OS) addition on swine nutrient digestibility, ileal and fecal bacterial populations, and ileal short-chain fatty acid (SCFA) production, and to determine their impact on ileal fermentative characteristics in vitro. Twelve T-cannulated pigs (BW = 25 kg) were fed a diet free of Gal OS for 21 d. On d 22, ileal digesta samples were collected for an in vitro fermentation experiment (Exp. 1). Substrates included: raffinose/stachyose combination (R + S), soy solubles (SS), and transgalactooligosaccharides (TOS). Also included were the non-OS components of SS and TOS. Nine pigs (three donors per treatment) served as ileal effluent donors. Each substrate was fermented in vitro for 6 h, and pH and SCFA and gas production were determined. Pigs then were allotted to three treatments: a Gal OS-free control diet and the control diet with either 3.5% added Gal OS from SS or TOS. Diets, feces, and digesta samples collected weekly for 6 wk on d 6 (feces) and 7 (digesta) were analyzed for DM, OM, CP, and chromic oxide concentrations. Feces and ileal digesta were analyzed for bifidobacteria and lactobacilli populations. Ileal digesta samples were analyzed for SCFA. On d 64, a second in vitro fermentation experiment (Exp. 2) was conducted using ileal effluent from three pigs per treatment and the same substrates used in Exp. 1. In vivo results showed that ileal and total tract DM and OM digestion were decreased (P < 0.05) by addition of both SS and TOS to the diet. Ileal and total-tract N digestibilities were decreased (P < 0.05) by dietary addition of SS. Fecal bifidobacteria and lactobacilli were increased (P < 0.05) by addition of SS and TOS to the diet. Ileal propionate and butyrate concentrations were greater (P < 0.05) for pigs fed diets containing both sources of Gal OS. In vitro results showed that fermentation data were not affected by donor animal adaptation to treatment. For both in vitro experiments, gas and SCFA production were higher (P < 0.05) for R + S than for SS or TOS. Fermentation of R + S resulted in a higher pH (P < 0.05) than did SS or TOS. Fermentation of non-OS components of SS and TOS resulted in more (P < 0.05) gas and SCFA production, and pH values that did not differ (P > 0.05) compared to SS and TOS. The Gal OS used in this study were prebiotics, increasing beneficial bacteria in vivo and SCFA concentrations both in vivo and in vitro.

Effects of dietary oligosaccharides on microbial diversity and fructo-oligosaccharide degrading bacteria in faeces of piglets post-weaning

Newly weaned 4-week-old piglets were fed a semi-synthetic diet supplemented with 40 g fructo-oligosaccharides (FOS) or 40 g transgalacto-oligosaccharides (TOS) per kg replacing 20 g maize starch and 20 g cellulose per kg in the control diet. Faecal samples were collected on days 0, 3, 7, 14 and 28 after weaning and analysed for dry matter, pH, organic acid concentration and distribution, microbial populations and identity of dominant bacterial species able to ferment FOS. The oligosaccharide addition to diets had no effect on growth performance of the piglets. Dietary oligosaccharides and time after weaning did not affect faecal dry matter content. There was no effect of diet on faecal pH and total organic acid concentration, but the pH decreased ( P<0.001) and total organic acid concentration increased ( P<0.001) during the period after weaning. Faecal bacterial populations (total culturable anaerobic bacteria, lactic acid bacteria, lactobacilli, bifidobacteria, coliforms and lactose-negative enterobacteria) were not affected by diet. The faecal population of yeast was significantly higher in piglets fed the TOS-diet as compared to piglets fed the FOS-diet ( P<0.05) and control diet ( P<0.001). The population of yeast was also higher in piglets fed the FOS-diet as compared to those fed the control diet ( P<0.05). In the faeces of piglets fed the control diet, the level of FOS-degrading bacteria was significantly lower than the level of saccharolytic bacteria ( P<0.01). This difference was not observed in the faeces of piglets fed the FOS-diet, indicating that the microbial population changed in response to the dietary FOS. A 16S rDNA-based identification of the FOS-degrading bacteria indicated that several different species of bacteria present in faeces of piglets are able to ferment FOS. It was concluded that oligosaccharide supplementation in diets to piglets after weaning had no effect on growth performance, and no or little effect on physico-chemical characteristics and bacterial populations of faeces. A strong stimulating effect on the faecal population of yeast was observed and the impact of this on pig performance is discussed.

Characterization of fecal bacterial populations in canines: effects of age, breed and dietary fiber.

The effects of age, breed, and diet on fecal chemistry, enzyme activity, and bacterial populations of dogs were studied. Eighteen dogs from two age groups (young: 2.5 +/- 0.5 years, old: 10.9 +/-0.7 years) and three different breeds (German shepherds, miniature schnauzers, and English setters) were rotated through a Latin Square design such that every dog was fed each of the diets. The test diets included a low-fiber (control) diet and a 10% fiber diet which contained 5% soybean hulls and 5% beet pulp. Inclusion of 10% fiber in the diet decreased the fecal concentration of ammonia, sulfide, and indole. Fiber inclusion significantly increased acetic, propionic, and butyric acid concentrations, while fecal pH decreased by 0.4 units. Fresh fecal samples were plated on selected aerobic and anaerobic culture media and DNA extracted for denaturing gradient gel electrophoresis (DGGE) analysis of PCR-amplified 16S ribosomal DNA fragments. Plate counts showed significant effects of breed (p < or = 0.05) and age (p < or = 0.01) on selected aerobic and anaerobic bacterial counts, while no significant effect of diet was found. Analysis of PCR-DGGE banding patterns showed there was a tendency for individual dogs to cluster together according to age (young or old dogs) and also for size (large or small dogs). However, the outstanding conclusion obtained from the DGGE analysis of fecal bacterial profiles was that individual dogs had their own characteristic banding pattern which was unique and stable. The relative stability and individuality of the patterns indicates that each individual harbored a characteristic fecal bacterial community which was independent of diet.

Cellulolytic Bacteria in Human Gut and Irritable Bowel Syndrome

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Administration of Bifidobacterium bifidum to Chicken Broilers Reduces the Number of Carcass Condemnations for Cellulitis at the Abattoir

A trial was conducted to evaluate the effect of Bifidobacterium bifidum on fecal bacterial populations, incidence of cellulitis, and growth performance in broiler chickens. Two groups of 9,385 broiler chickens each were administered 0 or 107B. bifidum cells/mL of drinking water for 38 d. Ten chickens from each group were randomly selected and euthanized every week; cecal samples were taken and analyzed for enumeration of total aerobic and anaerobic bacteria, coliforms, clostridia, and bifidobacteria. Mortality was recorded as it occurred. On Day 38, the chickens were weighed and shipped to the abattoir where carcass condemnations due to cellulitis and other causes were enumerated. The administration of B. bifidum had no significant effect on growth performance; however, the chickens that received the treatment had significantly increased numbers of fecal bifidobacteria on Days 28 and 35 of the experiment. Throughout the experiment, there was a trend of reduction of total aerobic bacteria, coliforms, and clostridia in the chickens that received B. bifidum. The number of condemnations for cellulitis and the total number of condemnations at slaughter were significantly reduced in the chickens that received B. bifidum. Results indicate that the administration of B. bifidum may provide a method of reducing cellulitis in broiler chickens.

Molecular Ecological Analysis of Dietary and Antibiotic-Induced Alterations of the Mouse Intestinal Microbiota

A cultivation-independent approach, polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE), was used to characterize changes in fecal bacterial populations resulting from consumption of a low residue diet or oral administration of a broad-spectrum antibiotic. C57BL/6NHsd mice were weaned to either a standard nonpurified diet (LC-diet) or a low residue diet (LR-diet) and at 17 wk of age were randomly assigned to receive drinking water with or without 25 ppm cefoxitin for 14 d. On d 1, 2, 7 and 14, microbial DNA was extracted from feces, and the V3 region of the 16S rDNA gene was amplified by PCR and analyzed by DGGE. The diversity of fecal microbial populations, assessed using Shannon's index (H'), which incorporates species richness (number of species, or in this case, PCR-DGGE bands) and evenness (the relative distribution of species), was not affected by cefoxitin. However, use of Sorenson's pairwise similarity coefficient (C(s)), an index that measures the species in common between different habitats, indicated that the species composition of fecal bacterial communities was altered by cefoxitin in mice fed either diet. Dietary effects on fecal microbial communities were more pronounced, with greater H' values (P < 0.05) in mice fed the LR-diet (1.9 +/- 0.1) compared with the LC-diet (1.6 +/- 0.1). The C(s) values were also greater (P < 0.05) in fecal bacterial populations from mice fed the LR-diet (C(s) = 69.8 +/- 2.0%) compared with mice fed the LC-diet (C(s) = 50.1 +/- 3.8%), indicating greater homogeneity of fecal bacterial communities in mice fed the LR-diet. These results demonstrate the utility of cultivation-independent PCR-DGGE analysis combined with measurements of ecological diversity for monitoring diet- and antibiotic-induced alterations of the complex intestinal microbial ecosystem.

Effect of the dietary supplementation of fructooligosaccharides and Bifidobacterium longum to early-weaned pigs on performance and fecal bacterial populations

Two experiments were conducted to evaluate the effect of administration of fructooligosaccharides (FOS) and Bifidobacterium longumon growth performance and fecal bacterial populations in pigs weaned at 18 d of age. In exp. 1, two groups of 20 pigs each were fed diets containing 0 or 0.5% FOS for 21 d. On days 1 and 3, pigs receiving FOS were administered an oral dose of 1010 B. longumcells. During week 1, average daily gain was higher and feed efficiency was improved (P &lt; 0.05) in pigs fed FOS and bifidobacteria relative to the control. On day 7, supplemented pigs had reduced numbers of total anaerobes and clostridia, and increased numbers of bifidobacteria in faeces (P &lt; 0.05). In exp. 2, 80 pigs were divided into four groups of 20 animals each in a 2 x 2 factorial design. Main effects included FOS supplementation (0 or 0.5% of diet) and B. longum supplementation (0 or 107 cells per gram feed). FOS supplementation reduced growth while B. longum supplementation increased growth (P&lt; 0.05). On days 12 and 19, serum insulin-like growth factor I (IGF-I) was reduced with FOS supplementation and increased with B. longum supplementation (P &lt; 0.05). It was concluded that the continuous administration of bifidobacteria provided a beneficial effect on growth parameters studied in this experiment. Key words: Pigs, fructooligosaccharides, bifidobacteria, Bifidobacterium longum, performance

Denaturing Gradient Gel Electrophoresis Analysis of 16S Ribosomal DNA Amplicons To Monitor Changes in Fecal Bacterial Populations of Weaning Pigs after Introduction of Lactobacillus reuteri Strain MM53

The diversity and stability of the fecal bacterial microbiota in weaning pigs was studied after introduction of an exogenous Lactobacillus reuteri strain, MM53, using a combination of cultivation and techniques based on genes encoding 16S rRNA (16S rDNA). Piglets (n = 9) were assigned to three treatment groups (control, daily dosed, and 4th-day dosed), and fresh fecal samples were collected daily. Dosed animals received 2.5 x 10(10) CFU of antibiotic-resistant L. reuteri MM53 daily or every 4th day. Mean Lactobacillus counts for the three groups ranged from 1 x 10(9) to 4 x 10(9) CFU/g of feces. Enumeration of strain L. reuteri MM53 on MRS agar (Difco) plates containing streptomycin and rifampin showed that the introduced strain fluctuated between 8 x 10(3) and 5 x 10(6) CFU/g of feces in the two dosed groups. Denaturing gradient gel electrophoresis (DGGE) of PCR-amplified 16S rDNA fragments, with primers specific for variable regions 1 and 3 (V1 and V3), was used to profile complexity of fecal bacterial populations. Analysis of DGGE banding profiles indicated that each individual maintained a unique fecal bacterial population that was stable over time, suggesting a strong host influence. In addition, individual DGGE patterns could be separated into distinct time-dependent clusters. Primers designed specifically to restrict DGGE analysis to a select group of lactobacilli allowed examination of interspecies relationships and abundance. Based on relative band migration distance and sequence determination, L. reuteri was distinguishable within the V1 region 16S rDNA gene patterns. Daily fluctuations in specific bands within these profiles were observed, which revealed an antagonistic relationship between L. reuteri MM53 (band V1-3) and another indigenous Lactobacillus assemblage (band V1-6).

The effect of a model melanoidin mixture on faecal bacterial populationsin vitro

The Maillard reaction produces coloured, macromolecular materials (melanoidins) in a variety of foods, on heating. Significant quantities may enter the human gut on a daily basis, but there is little information on their metabolism in the human colon. As the large bowel contains a diverse population of bacteria involved in normal bowel function, it is possible that melanoidins are metabolized therein. Depending on the bacteria involved, there may be disease or health implications. The aim of the present study was to use in vitro models to determine the digestibility of melanoidins and the effect of melanoidins on colonic bacteria in the gastrointestinal tract. Melanoidins were prepared and the effects of simulated upper-gut secretions on their stability determined in a model system. The effects of faecal bacteria were also determined, in batch culture, with a combination of phenotypic and genotypic (probes) criteria being used to identify the microbial diversity involved. Simulation of peptic and pancreatic digestion showed that the melanoidins did not produce detectable amounts of low-molecular-mass degradation products. However, melanoidins affected the growth of gut bacteria during mixed culture growth. The effect was to cause a non-specific increase in the anaerobic bacteria enumerated. This in vitro study indicates that melanoidins can affect the growth of human large-bowel bacteria and serves to demonstrate possible effects that may occur in vivo. Given the large and varied number of food items that contain Maillard reaction products, this may have relevance for lower-gut health.

Fecal coliform-related bacterial and coliphage populations in five lakes of southeastern Spain

Aerobic heterotrophic bacteria, fecal and total coliforms, fecal streptococci and coliphages were isolated from five protected lakes in the Antequera area of Spain over the time from January to March (1994–96). The water samples contained large number of heterotrophic bacteria (mean counts 0.2 to 5.0 × 10 7 cfu per 100 ml). Most of the lakes contained fecal streptococci and a relationship between streptococci and salinity of the water samples was stablished. Coliphages were isolated from lakes containing fecal coliform and these bacteria were taxonomically identified as E. coli. Coliform bacilli do not seem to be an adequate indicator of fecal pollution for these ephemeral small lakes.

Monoclonal immunoglobulin A derived from peritoneal B cells is encoded by both germ line and somatically mutated VH genes and is reactive with commensal bacteria.

We transferred peritoneal cells from BALB/c mice into C.B17 scid/scid mice. Six to eight months after injection, only cells with the B1 phenotype were retained in the spleens and peritoneal cavities of these mice. The lamina propria of the intestine contained many peritoneal, donor-derived, immunoglobulin A (IgA)-producing cells. The mesenteric lymph nodes of these mice were found to be a major site of proliferation and generation of IgA plasmablasts. We established eight IgA-producing hybridomas from the mesenteric lymph nodes of such mice, and all the hybridomas reacted with different but partially overlapping fecal bacterial populations. Cloning and sequencing of the VH genes of these hybridomas showed that two hybridomas utilized germ line-encoded VH genes while the VH genes of the six hybridomas showed somatic mutations, some of which are indicative of an antigen-driven selection process.

Occurrence of glycopeptide resistance among Enterococcus faecium isolates from conventional and ecological poultry farms.

Occurrence of glycopeptide resistance among Enterococcus faecium isolates from conventional and ecological poultry farms.

Effect of Administration ofLactobacillus caseiStrain GG on the Gastrointestinal Microbiota of Newborns

The aim of the study was to determine whether Lactobacillus casei strain GG could colonise the intestine of newborns and the influence of its administration on establishment of the microbiota. The faecal bacterial population of 25 under 1 mth old newborns was studied: in addition to breastfeeding, 15 babies (GG group) received for 2 wk immediately after birth Lactobacillus GG supplement as freeze-dried powder diluted in water in a dose of 10 10 -10 11 c.f.u./g; 10 newborns (control group) did not receive any supplement to breastfeeding. The faecal bacterial composition of meconium was similar in both groups studied. Ten newborns (67 per cent) excreted Lactobacillus GG, while in eight cases (53.3 per cent) Lactobacillus GG was found even 2 wk after the administration was stopped. The faecal concentrations and the relative proportion of Lactobacillus GG were individually different. In 3-4 d, 5-7 d and 1 mth old newborns of the GG group the faecal concentrations of lactobacilli exceeded those of the control group. The faecal microorganisms predominance pattern did not differ in the case of 1 wk and 1 mth old newborns of the GG group. The study shows that 2 wk administration of Lactobacillus GG, which starts right after birth, increases intestinal lactobacilli concentrations and does not impair the establishment of a normal faecal bacterial microbiota. Keywords - Lactobacillus casei GG, Neonatal faecal bacteria.

Mucin Glycoprotein Degradation by Mucin Oligosaccharide-degrading Strains of Human Faecal Bacteria. Characterisation of Saccharide Cleavage Products and their Potential Role in Nutritional Support of Larger Faecal Bacterial Populations

Mucin Glycoprotein Degradation by Mucin Oligosaccharide-degrading Strains of Human Faecal Bacteria. Characterisation of Saccharide Cleavage Products and their Potential Role in Nutritional Support of Larger Faecal Bacterial Populations

Mucin Glycoprotein Degradation by Mucin Oligosaccharide-degrading Strains of Human Faecal Bacteria. Characterisation of Saccharide Cleavage Products and their Potential Role in Nutritional Support of Larger Faecal Bacterial Populations

A subset of normal human faecal bacteria composed of mucin oligosaccharide-degrading (MOD) strains appears to be the principal source of intraluminal, extracellular glycosidases degrading the oligosaccharides of gut mucin glycoproteins and epithelial membrane glycoconjugates. Here we report studies characterising the saccharides released from hog gastric mucin during incubation with the extracellular enzymes in their cell-free culture supernates. We have also tested the ability of the saccharide products to support growth of larger populations of faecal bacteria.Gel exclusion chromatography of the residual mucin following bacterial growth in mucin-containing medium confirmed that MOD strains extensively degraded the carbohydrate moieties (60–96 per cent) whereas strains of bacteroides and bifidobacteria of larger faecal populations degraded the carbohydrate moieties less extensively (median loss, 29 per cent; range 8–42 per cent). Saccharides released from hog gastric mucin by MOD strain enzymes were identified by biochemical and NMR spectroscopic methods as galactose, fucose. N-acetylglucosaminc, N-acetylgalactosamme and the disaccharides Galβ3GalNAc and Galβ3GlcNAc. Larger oligosaccharides were not recovered. Carbon-13 NMR spectra of the residual mucin were consistent with complete removal of some but not all oligosaccharide chains. The monosaccharides but not the disaccharides were also recovered following incubation of mucin with the sterile cell-free supernate of a heavily inoculated faecal culture. Saccharides released from hog gastric mucin by partially purified culture supernate enzymes from Ruminococcus torques strain IX-70 were 70–98 per cent utilised by larger populations of faecal bacteria during growth in culture. Bacteria growing from 1 × 10−10 g faecal inocula from five subjects showed minimal growth in media containing intact hog gastric mucin but grew well if the mucin-containing medium was preincubated with a sterile enzyme preparation from R. torques IX-70. We conclude that glycosidases produced by MOD strains can serve a nutritionally supportive role for larger populations of enteric bacteria, especially during fasting.

Antibiotic Resistance and Transferable R Plasmids in Coliforms from a Polluted Estuary

AbstractThe aim of this work was to find out how far the Nervion Estuary serves as a reservoir of R plasmids and/or for their dissemination. The fecal pollution bacterial indicator population was estimated along the estuary and 167 coliform isolates were assayed for resistance to 16 antibiotics. About sixty‐nine per cent of the coliform isolates were antibiotic‐resistant and forty‐two per cent of these were simultaneously resistant to three or more of 16 antibiotics tested. Furthermore, 31.5% of enterobacterial strains resistant to any of the antibiotics tested could transfer their resistance determinants in conjugal matings. Most of the transferred plasmids confer resistance to ampicillin or to other β‐lactamic antibiotics and several codify resistance to tetracyclin, chloramphenicol of fosfomycin. Taking account of the coliforms density found and the ratio of these isolates able to transfer R plasmids, an average of 5 × 103 coliform strains able to transfer R plasmids in 100 ml of water was obtained. This amount must indicate a significant potential public health risk because these polluted waters get into an area with intensive fishing and recreational use.

Colonic bacterial proteases to IgA1 and sIgA in patients with ulcerative colitis.

The colonic faecal and mucosal associated bacterial populations of five patients with ulcerative colitis and four control patients were studied in detail to assess their ability to produce IgA1-proteases. A total of 330 bacterial strains were isolated from the patients with ulcerative colitis and IgA1-protease activity was unable to be reliably shown in any. It is therefore unlikely that such enzyme production by colonic bacteria plays a significant role in the pathogenesis of ulcerative colitis.

Morphological aspects of dietary fibers in the intestine.

Publisher Summary This chapter discusses the morphological aspects of dietary fibers in the intestine. It presents the results from a comparative assessment of the structural responses to the inclusion of carefully defined semi purified fibers and bile salt-binding resins in paired groups of animals. The morphological and analytical modes deployed include routine light microscopy, transmission electron microscopy, and scanning electron microscopy of architectural surface topography of the mucosa. There is a lack of consistent information in regard to the antiatherogenic potential of individual dietary fibers in different species. The mechanisms proposed to account for their hypolipidemic properties include reduced intestinal transit time; binding of bile acids or phospholipids in the intestinal lumen, thereby reducing absorptive capacity; alterations in hepatic lipid and bile acid metabolism; and effects on the serum and tissue distribution of lipids. The effects of high-fiber feeding on fecal sterol concentrations and bacterial populations in the colon have also received much attention in the field of colon carcinogenesis. It is becoming apparent that the presence of fiber in the gastrointestinal tract may affect the regulation of endocrinological or peptide factors that are critical to gastrointestinal functioning.

Suppression of human fecal 7α-hydroxysteroid dehydrogenase by a short-term low cholesterol diet

Fourteen healthy human volunteers were placed on short-term (1–4 weeks) isocaloric dietary regimes on an outpatient basis. Each diet consisted of approx. 2200 calories and contained 17% protein, 35% fat, 48% carbohydrate and varied only in mg/day cholesterol intake and polyunsaturated/saturated (P/S) fatty acid ratio. Diets A, B, C and D consisted of a daily cholesterol intake of 100 mg cholesterol (chicken and/or fish based P/S = 2.0), 500 mg cholesterol (fish and/or chicken based P/S = 0.17), 500 mg cholesterol (beef based P/S = 0.35) and 1000 mg cholesterol (beef based P/S = 0.10). When both NAD- and NADP-dependent bile salt dehydrogenases in mixed fecal cultures were measured before and after weekly dietary sequences: A alone ( n = 2), B alone ( n = 2), C alone ( n = 2), A-D ( n = 5), A-D-D-D ( n = 2), B-A ( n = 2), and D-A ( n = 4), the NAD-dependent 7α-hydroxysteroid dehydrogenase (HSDH) activity was shown to be significantly depressed or eliminated after one week of diet A, and returned when subjects were taken off diet A or placed on another diet. No clear effect on 7α-HSDH was measurable by diets B, C or D. No dietary effect was seen on 3α-, 12α-HSDH or the NADP-component of 7α-HSDH. On serial dilution of fecal microorganisms and growth of the bacterial populations to stationary phase, NAD-dependent 7α-HSDH consistently increased on dilution, while NADP-dependent 7α-HSDH and NADP-dependent 12α-HSDH decreased and vanished on dilution. When NAD-dependent 7α-HSDH was eliminated by diet A, it could be restored by diluting fecal bacterial populations, simultaneously decreasing and eliminating NADP-7α-HSDH and NADP-12α-HSDH. The results suggest that these three enzymes may be markers for ecologically related fecal sub-populations. The dietary suppression of NAD-dependent 7α-HSDH has been interpreted as a transient suppression of that subpopulation carrying this enzyme.

Relationship between various bacterial populations and coprostanol and cholesterol.

A year-long study was conducted at three different locations in Canada to evaluate the relationship between fecal sterols and bacterial populations. A statistical model was constructed to study the following: (i) seasonal variations in the sterols and bacterial parameters; (ii) the differences between morning and afternoon samples; (iii) the relationship between the two sterols (coprostanol and cholesterol) as well as their relationship to the bacterial parameters; and (iv) the geographical differences between the three sampling sites. Application of the model to the data indicated that (i) the relationship in receiving waters between coprostanol and cholesterol was found to be highly significant; (ii) at two sampling sites the relationship between fecal sterols and fecal coliforms was positively significant at the 5% level; (iii) at one sampling site there was a positive relationship between fecal sterols and the fecal coliform - fecal streptococcus ratio; and (iv) in a.m. vs. p.m. samples significant differences were noted in coliform, fecal coliform, and fecal streptococcus densities while no significant differences were found in heterotroph densities and coprostanol and cholesterol levels. Raw and treated sewage from four sewage-treatment plants were also studied to provide baseline control data. Data support the conclusion that while no consistent relationship exists between bacterial parameters and fecal sterols, fecal sterols could be considered as unequivocal indicators of the presence of fecal material in water.