Detection of false-positives among total and fecal coliform counts by factorial analysis of correspondence.

Application of an analysis of correspondence to the biochemical characteristics of total and fecal coliforms isolated in the Ivory Coast permitted us to separate two small clusters of isolates different from the main clusters, which included isolates from human and animal feces. The isolates grouped in the small clusters were from water samples. An analysis of the biochemical characteristics which permitted the segregation of the "water-specific" isolates from the main clusters indicates that water-specific total coliforms were citrate positive, indole negative, and amygdaline positive. Water-specific fecal coliforms were either citrate positive, indole negative, amygdaline positive, and inositol negative or indole negative, amygdaline positive, and inositol positive. Any isolates not fitting the above patterns could be considered of fecal origin. If this observation is confirmed under temperate climates and for a greater number of isolates, these simple tests could be used to confirm the fecal origin of coliforms.     

Incidence of coliphage in potable water supplies

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

Incidence of coliphage in potable water supplies.

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

Characterization of the coliform and enteric bacilli in the environment of calves with colibacillosis.

In the first part of the present study the coliform and enteric bacilli in the environment of calves with colibacillosis were examined. The occurrence, number, and pathogenic properties of Escherichia coli in barnyard soils were obtained from six cattle ranches. The O and K serogroups of E. coli isolates obtained from the feces of calves with colibacillosis born at these cattle ranches were determined, and their serotypes were compared with the E. coli O and K serotypes found in soils. The results showed a reservoir of potentially pathogenic E. coli in barnyard soils contaminated with bovine feces. For the second part of this study, 6 healthy calves and 51 calves with colibacillosis were studied. The numbers of total aerobic heterotrophic bacteria, total streptococci, fecal streptococci, total coliforms, and fecal coliforms in the feces of calves were determined. In addition, coliform and enteric bacilli from the feces of both healthy and diseased calves were identified, and their indole, methyl red, Voges-Proskauer, citrate (IMViC) types were described. In parallel, the IMViC types of coliform and enteric bacilli isolated from barnyard soils previously contaminated with bovine feces were compared with those isolated from uncontaminated soils. All fecal specimens were also examined for the presence of rotavirus. No significant effect on the numbers of the bacterial types was found. The results suggest that the predominant IMViC types found in the feces of calves with colibacillosis originate from the soil. From this study it is apparent that the occurrence, number, and survival of E. coli in barnyard soils is related to ranch husbandry and sanitary practices.     

Characterization of the coliform and enteric bacilli in the environment of calves with colibacillosis

In the first part of the present study the coliform and enteric bacilli in the environment of calves with colibacillosis were examined. The occurrence, number, and pathogenic properties of Escherichia coli in barnyard soils were obtained from six cattle ranches. The O and K serogroups of E. coli isolates obtained from the feces of calves with colibacillosis born at these cattle ranches were determined, and their serotypes were compared with the E. coli O and K serotypes found in soils. The results showed a reservoir of potentially pathogenic E. coli in barnyard soils contaminated with bovine feces. For the second part of this study, 6 healthy calves and 51 calves with colibacillosis were studied. The numbers of total aerobic heterotrophic bacteria, total streptococci, fecal streptococci, total coliforms, and fecal coliforms in the feces of calves were determined. In addition, coliform and enteric bacilli from the feces of both healthy and diseased calves were identified, and their indole, methyl red, Voges-Proskauer, citrate (IMViC) types were described. In parallel, the IMViC types of coliform and enteric bacilli isolated from barnyard soils previously contaminated with bovine feces were compared with those isolated from uncontaminated soils. All fecal specimens were also examined for the presence of rotavirus. No significant effect on the numbers of the bacterial types was found. The results suggest that the predominant IMViC types found in the feces of calves with colibacillosis originate from the soil. From this study it is apparent that the occurrence, number, and survival of E. coli in barnyard soils is related to ranch husbandry and sanitary practices.     

Clonal populations of thermotolerant Enterobacteriaceae in recreational water and their potential interference with fecal Escherichia coli counts

Bacterial strains were isolated from beach water samples using the original Environmental Protection Agency method for Escherichia coli enumeration and analyzed by pulsed-field gel electrophoresis (PFGE). Identical PFGE patterns were found for numerous isolates from 4 of the 9 days sampled, suggesting environmental replication. 16S rRNA gene sequencing, API 20E biochemical testing, and the absence of beta-glucuronidase activity revealed that these clonal isolates were Klebsiella, Citrobacter, and Enterobacter spp. In contrast, 82% of the nonclonal isolates from water samples were confirmed to be E. coli, and 16% were identified as other fecal coliforms. These nonclonal isolates produced a diverse range of PFGE patterns similar to those of isolates obtained directly from untreated sewage and gull droppings. beta-Glucuronidase activity was critical in distinguishing E. coli from other fecal coliforms, particularly for the clonal isolates. These findings demonstrate that E. coli is a better indicator of fecal pollution than fecal coliforms, which may replicate in the environment and falsely elevate indicator organism levels.     

Evaluation of beta-glucuronidase assay for the detection of Escherichia coli from environmental waters.

The new United States Drinking Water Regulations state that water systems must analyze for Escherichia coli or fecal coliforms on any routine or repeat sample that is positive for total coliforms. The proposed methods for the detection of E. coli are based on beta-glucuronidase activity, using the fluorogenic substrate 4-methylumbelliferyl beta-D-glucuronide (MUG). This study was conducted to determine whether beta-glucuronidase negative E. coli were present in significant numbers in environmental waters. Two hundred and forty E. coli cultures were isolated from 12 water samples collected from different environmental sources. beta-glucuronidase activity was determined using lauryl tryptose broth with MUG, EC broth with MUG, and the Autoanalysis Colilert (AC) procedure. The isolates were also evaluated by the standard EC broth gas fermentation method for fecal coliforms. The results confirm that assaying for the enzyme beta-glucuronidase utilizing the MUG substrate is an accurate method for the detection of E. coli in environmental waters.     

Clonal Populations of Thermotolerant Enterobacteriaceae in Recreational Water and Their Potential Interference with Fecal Escherichia coli Counts

Bacterial strains were isolated from beach water samples using the original Environmental Protection Agency method for Escherichia coli enumeration and analyzed by pulsed-field gel electrophoresis (PFGE). Identical PFGE patterns were found for numerous isolates from 4 of the 9 days sampled, suggesting environmental replication. 16S rRNA gene sequencing, API 20E biochemical testing, and the absence of β-glucuronidase activity revealed that these clonal isolates were Klebsiella, Citrobacter, and Enterobacter spp. In contrast, 82% of the nonclonal isolates from water samples were confirmed to be E. coli, and 16% were identified as other fecal coliforms. These nonclonal isolates produced a diverse range of PFGE patterns similar to those of isolates obtained directly from untreated sewage and gull droppings. β-Glucuronidase activity was critical in distinguishing E. coli from other fecal coliforms, particularly for the clonal isolates. These findings demonstrate that E. coli is a better indicator of fecal pollution than fecal coliforms, which may replicate in the environment and falsely elevate indicator organism levels.     

Unusual Organism Which Gives a Positive Elevated Temperature Test for Fecal Coliforms

Organisms apparently not of fecal origin isolated from a sulfur hot spring gave a positive elevated temperature test for fecal coliforms.     

Occurrence of Coliforms, Fecal Coliforms, and Streptococci on Vegetation and Insects

This study considers the sanitary significance of coliforms, fecal coliforms, and streptococci isolated from 152 species of plants and 40 samples of insects. These specimens were collected from various ecological environments and grouped into several categories. Results indicate that typical coliforms of the warm-blooded animal gut contribute a relatively small percentage of the organisms associated with vegetation (14.1%) and insects, (14.9%). A total of 1,203 coliform strains from vegetation and 1,084 coliform strains from insects were classified as to IMViC type and fecal coliform. No type was predominant in either the vegetation or insect groupings. The biochemical results for 646 streptococci from vegetation and 226 cultures from insects were reported. The predominant group, Streptococcus fecalis, as defined by Sherman criteria, constituted a majority of all strains from vegetation and insects. The “Completed Coliform Test” is recommended for the examination of plant and insect specimens to eliminate the many anaerobic and aerobic sporeforming bacteria that frequently produce false positive reactions by the “Confirmed Test” procedure. These findings support the current interpretation of the significance of the fecal coliform test for stream investigations or for surface water quality evaluations.     

Bacteriology of Dehydrated Space Foods

The initial bacteriological requirement established in 1964 for space foods by the U.S. Army Natick Laboratories are: a total aerobic plate count ( 300,000), chocolate ice cream cubes (20,000), and each of four samples of chocolate candy (12,000 to 61,000); (ii) coliforms: two out of three vanilla milk drinks (16 and 127) and one beef hash bar (14); (iii) fecal coliforms: one sample of chicken soup and gravy base positive; (iv) fecal streptococci: two samples of peanut cubes (40 and 108), coconut cubes (75), chicken soup and gravy base (2,650), beef soup and gravy base (33), and five out of six flavored milk drinks (23 to 300); (v) salmonellae: one each of chicken and beef soup and gravy base were positive.     

A Halophilic Vibrio Isolated from a Case of Chronic Cholecystitis

We found a halophilic vibrio in B bile from a 75-year-old female patient with chronic cholecystitis, and examined its biochemical characteristics. The organisms are gram-negative short rods or comma shaped, with some ring forms. They have a single polar flagellum, but no capsule. The strains can grow in peptone water with 1.0 to 4.0% NaCl, but not with no NaCl or 6.0% NaCl. The characteristics of the organisms are positive dextrose fermentation, catalase, oxidase, and ornithine decarboxylase, and negative lysine decarboxylase, arginine dihydrolase, and absence of gas from glucose. They are sensitive to 2,4-diamine-6,7-diisopropyl pteridine (0/129). These characteristics indicate that the isolated strain should be a halophilic vibrio. However, no growth on Salmonella-Shigella (SS), SS with added sucrose and bromcresol purple (SS-SB), MacConkey's or thiosulfate citrate bile salts (TCBS) agar plates was demonstrated. Nitrate reduction, Simmons' citrate agar, indole, o-nitrophenol-β-d-galactopyranoside (ONPG), motility and esculin hydrolysis were positive. Urease, gelatinase, Voges-Proskauer, phenylalanine deaminase and malonate reactions were negative. Acid was produced from amygdalin, arabinose, cellobiose, fructose, galactose, glucose, lactose, maltose, mannitol, salicin, starch, sucrose, trehalose, and xylose, but not from adonitol, dulcitol, inositol, mannose, melibiose, raffinose, rhamnose and sorbitol. From these characteristics the isolate is considered to be not identical with V. parahaemolyticus, V. cholerae, V. vulnificus or other vibrios. It can be presumed that this isolate represents another species of halophilic vibrio.     

Differentiation of Aerobacter-Klebsiella isolated from sugarcane

Three hundred and eighty-four isolates were obtained in the completed test portion of the most probable number determinations of coliforms in sugarcane sources. Of these isolates, 88% were of the (- - + +) indole, methyl red, Voges-Proskauer, citrate (IMViC) type and were identified as Aerobacter aerogenes according to the protocol of the American Public Health Association (1). Employing 359 of these cultures, a comparative biochemical, serological, and pathogenicity study was carried out with Klebsiella pneumoniae CDC no. 2211-66 type 9. More than 86% of the organisms tested gave biochemical reactions typical of K. pneumoniae. Of the other isolates, 2% were Enterobacter aerogenes, and the remaining 12% were identified as atypical, nonmotile IMViC types. Comparable agglutination titers were also observed between A. aerogenes and the CDC strain of K. pneumoniae when several randomly selected sugarcane strains were reacted with prepared K. pneumoniae whole cell antiserum. Neither the K. pneumoniae reference organism nor selected sugarcane isolates displayed pathogenicity for mice. On the basis of all the analyses performed, it was suggested that such organisms be classified as K. pneumoniae.     

Identification of fecal Escherichia coli from humans and animals by ribotyping

Fecal pollution of water resources is an environmental problem of increasing importance. Identification of individual host sources of fecal Escherichia coli, such as humans, pets, production animals, and wild animals, is prerequisite to formulation of remediation plans. Ribotyping has been used to distinguish fecal E. coli of human origin from pooled fecal E. coli isolates of nonhuman origin. We have extended application of this technique to distinguishing fecal E. coli ribotype patterns from human and seven individual nonhuman hosts. Classification accuracy was best when the analysis was limited to three host sources. Application of this technique to identification of host sources of fecal coliforms in water could assist in formulation of pollution reduction plans.     

Detection of Acinetobacter spp. in rural drinking water supplies.

A bacteriological survey was conducted of untreated, individual groundwater supplies in Preston County, W.Va. Nearly 60% of the water supplies contained total coliforms in excess of the U.S. Environmental Protection Agency maximum contaminant level of 1 CFU/100 ml. Approximately one-third of the water systems contained fecal coliforms and/or fecal streptococci. Acinetobacter spp. were detected in 38% of the groundwater supplies at an arithmetic mean density of 8 CFU/100 ml and were present in 16% of the water supplies in the absence of total coliforms, posing some concern about the usefulness of total coliforms as indicators of the presence of this opportunistic pathogen. Slime production, a virulence factor for A. calcoaceticus, was not significantly different between well water isolates and clinical strains, suggesting some degree of pathogenic potential for strains isolated from groundwater. In addition, several Acinetobacter isolates were able to interfere with sheen production by some coliform bacteria on M-Endo medium, adding further to the possible significance of Acinetobacter spp. in groundwater supplies.     

Detection of Acinetobacter spp. in rural drinking water supplies

A bacteriological survey was conducted of untreated, individual groundwater supplies in Preston County, W.Va. Nearly 60% of the water supplies contained total coliforms in excess of the U.S. Environmental Protection Agency maximum contaminant level of 1 CFU/100 ml. Approximately one-third of the water systems contained fecal coliforms and/or fecal streptococci. Acinetobacter spp. were detected in 38% of the groundwater supplies at an arithmetic mean density of 8 CFU/100 ml and were present in 16% of the water supplies in the absence of total coliforms, posing some concern about the usefulness of total coliforms as indicators of the presence of this opportunistic pathogen. Slime production, a virulence factor for A. calcoaceticus, was not significantly different between well water isolates and clinical strains, suggesting some degree of pathogenic potential for strains isolated from groundwater. In addition, several Acinetobacter isolates were able to interfere with sheen production by some coliform bacteria on M-Endo medium, adding further to the possible significance of Acinetobacter spp. in groundwater supplies.     

Morphological and Biochemical Differentiation of Three Types of Small Oral Spirochetes

THIRTY STRAINS OF HUMAN ORAL ANAEROBIC SPIROCHETES WERE ISOLATED IN THREE DIFFERENT MEDIA: veal heart infusion-ascitic fluid, Spirolate-Brain Heart Infusion-rabbit serum, and supplemented PPLO broth. The morphological and biochemical characteristics of the isolates permitted their differentiation into three distinct species: Treponema denticola, T. macrodentium, and T. oralis (proposed new species). These species could be differentiated as follows. Organisms of the T. denticola type had a "2-4-2" axial fibril relationship as determined by electron microscopy, required serum for growth, did not utilize glucose or lactate, and produced indole, ammonia, acetate, and lactate as end products. T. macrodentium had a "1-2-1" axial fibril relationship, did not require serum, utilized glucose but not lactate, did not produce indole or ammonia, and produced formate, acetate, lactate, and succinate as acid end products. T. oralis had a "1-2-1" axial fibril relationship, required serum for growth, utilized lactate but not glucose, produced indole but not ammonia, and produced propionate and acetate as acid end products.     

Validity of fecal coliforms, total coliforms, and fecal streptococci as indicators of viruses in chlorinated primary sewage effluents.

Quantities of combined chlorine that usually destroyed more than 99.999% of the indigenous fecal coliforms, total coliforms, and fecal streptococci in primary sewage effluents destroyed only 85 to 99% of the indigenous viruses present. Viruses were recovered from five of eight chlorinated primary effluents from which fecal coliforms were not recovered by standard most-probable-number procedures. The limited volumes of such chlorinated effluents that can be tested for indicator bacteria with currently available multiple-tube and membrane filter techniques restrict the value of fecal coliforms, fecal streptococci, and even total coliforms as indicators of viruses in these effluents. Although fecal coliforms and fecal streptococci are useful indicators of viruses in effluents from which these bacteria are recovered, the absence of these bacteria and even total coliforms from disinfected effluents (in standard tests) does not assure that viruses are also absent.     

Validity of fecal coliforms, total coliforms, and fecal streptococci as indicators of viruses in chlorinated primary sewage effluents.

Quantities of combined chlorine that usually destroyed more than 99.999% of the indigenous fecal coliforms, total coliforms, and fecal streptococci in primary sewage effluents destroyed only 85 to 99% of the indigenous viruses present. Viruses were recovered from five of eight chlorinated primary effluents from which fecal coliforms were not recovered by standard most-probable-number procedures. The limited volumes of such chlorinated effluents that can be tested for indicator bacteria with currently available multiple-tube and membrane filter techniques restrict the value of fecal coliforms, fecal streptococci, and even total coliforms as indicators of viruses in these effluents. Although fecal coliforms and fecal streptococci are useful indicators of viruses in effluents from which these bacteria are recovered, the absence of these bacteria and even total coliforms from disinfected effluents (in standard tests) does not assure that viruses are also absent.     

Identification of Fecal Escherichia coli from Humans and Animals by Ribotyping

Fecal pollution of water resources is an environmental problem of increasing importance. Identification of individual host sources of fecal Escherichia coli, such as humans, pets, production animals, and wild animals, is prerequisite to formulation of remediation plans. Ribotyping has been used to distinguish fecal E. coli of human origin from pooled fecal E. coli isolates of nonhuman origin. We have extended application of this technique to distinguishing fecal E. coli ribotype patterns from human and seven individual nonhuman hosts. Classification accuracy was best when the analysis was limited to three host sources. Application of this technique to identification of host sources of fecal coliforms in water could assist in formulation of pollution reduction plans.