The ecology of Escherichia coli in market calves fed a milk-substitute diet

Dynamic changes in the Escherichia coli population in the calf gut were studied over 21 days in a group of 18 intensively-reared market calves. Isolates were identified by O-serogrouping, biotyping and resistogram patterns. Seventy O-serogroups were identified among nearly 3000 E. coli isolates examined and these were subdivided into 416 strains by means of their biotype and resistogram. Seventy-five per cent of these strains were detected only once or twice, which points to the continual replacement of the E. coli flora with strains that showed low persistence in the gut. The rise in the frequency of antibiotic resistance observed during the study was not due to a change in the proportion of resistant to sensitive strains in the gut flora. It was a consequence of the displacement of the original flora by multiply-resistant strains, which presumably originated from the calves' environment.     

Bacterial conjugation in the digestive tracts of gnotoxenic chickens.

Escherichia coli K-12 Hfr and F- strains were successively implanted in axenic chicks. Conjugation with exchanges of chromosomal genes occurred with high frequencies in the gut of the chicks and could continue as long as fertile strains coexisted in this environment. Almost all of the expected recombinant types were recovered in the feces under these experimental conditions. Furthermore, these recombinants were analogous to those obtained after conjugations in vitro. Recombinants formed in the gut were more numerous when the F- strain was seeded before the Hfr strain. The recombinants showed no apparent selective advantage over the parental strains in the intestinal medium. They were maintained throughout the experimental period and represented more than 10% of the total intestinal flora. The chick gut is usually rapidly colonized by other bacterial types under natural conditions. The possible effects of other components of the bacterial flora on conjugation of E. coli in holoxenic animals will require subsequent work with more complex microbiological conditions.     

A comparison of the ecology of Escherichia coli in the intestine of healthy unweaned pigs and pigs after weaning

A total of 51 clinically healthy pigs (14 unweaned and 37 weaned) from five litters, and aged 21 to 35 d, were studied. Escherichia coli isolates from the duodenum, jejunum, ileum, caecum and colon were differentiated on the basis of O-serogroup, biotype and resistance pattern. The complexity of the flora was influenced considerably by the presence or absence of the enterotoxigenic serotype 0149: K91, K88a, o (Abbotstown strain). When it was absent the E. coli flora of both weaned and unweaned pigs was complex with up to 25 strains being identified. The majority of these E. coli strains identified in each pig were isolated from only one of the five intestinal sites sampled. On the other hand, when the enterotoxigenic strain was present (14 pigs) it tended to dominate the E. coli flora at all levels of the intestine and this dominance was reflected in a corresponding fall in the total number of E. coli strains isolated per pig.     

A comparison of the ecology of Escherichia coli in the intestine of healthy unweaned pigs and pigs after weaning

A total of 51 clinically healthy pigs (14 unweaned and 37 weaned) from five litters, and aged 21 to 35 d, were studied. Escherichia coli isolates from the duodenum, jejunum, ileum, caecum and colon were differentiated on the basis of O-serogroup, biotype and resistance pattern. The complexity of the flora was influenced considerably by the presence or absence of the enterotoxigenic serotype 0149: K91, K88a,c (Abbotstown strain). When it was absent the E. coli flora of both weaned and unweaned pigs was complex with up to 25 strains being identified. The majority of these E. coli strains identified in each pig were isolated from only one of the five intestinal sites sampled. On the other hand, when the enterotoxigenic strain was present (14 pigs) it tended to dominate the E. coli flora at all levels of the intestine and this dominance was reflected in a corresponding fall in the total number of E. coli strains isolated per pig.     

Gram-negative bacteria aggravate murine small intestinal Th1-type immunopathology following oral infection with Toxoplasma gondii

Oral infection of susceptible mice with Toxoplasma gondii results in Th1-type immunopathology in the ileum. We investigated gut flora changes during ileitis and determined contributions of gut bacteria to intestinal inflammation. Analysis of the intestinal microflora revealed that ileitis was accompanied by increasing bacterial load, decreasing species diversity, and bacterial translocation. Gram-negative bacteria identified as Escherichia coli and Bacteroides/Prevotella spp. accumulated in inflamed ileum at high concentrations. Prophylactic or therapeutic administration of ciprofloxacin and/or metronidazole ameliorated ileal immunopathology and reduced intestinal NO and IFN-gamma levels. Most strikingly, gnotobiotic mice in which cultivable gut bacteria were removed by quintuple antibiotic treatment did not develop ileitis after Toxoplasma gondii infection. A reduction in total numbers of lymphocytes was observed in the lamina propria of specific pathogen-free (SPF), but not gnotobiotic, mice upon development of ileitis. Relative numbers of CD4(+) T cells did not differ in naive vs infected gnotobiotic or SPF mice, but infected SPF mice showed a significant increase in the frequencies of activated CD4(+) T cells compared with gnotobiotic mice. Furthermore, recolonization with total gut flora, E. coli, or Bacteroides/Prevotella spp., but not Lactobacillus johnsonii, induced immunopathology in gnotobiotic mice. Animals recolonized with E. coli and/or total gut flora, but not L. johnsonii, showed elevated ileal NO and/or IFN-gamma levels. In conclusion, Gram-negative bacteria, i.e., E. coli, aggravate pathogen-induced intestinal Th1-type immunopathology. Thus, pathogen-induced acute ileitis may prove useful to study bacteria-host interactions in small intestinal inflammation and to test novel therapies based on modulation of gut flora.     

The Effect of Tetracycline on the Coliform Gut Flora of Broiler Chickens with Special Reference to Antibiotic Resistance and O-Serotypes of Escherichia coli

The effect of terramycin, administered prophylactically in drinking water, on the gut flora of broiler birds was investigated. Exposure to the antibiotic for only 24 h profoundly affected the counts of tetracycline-resistant strains and selected O-serotypes carrying resistance determinants. Large numbers of Escherichia coli resistant to sulphonamides were found in treated and control birds and this is discussed in relation to the use of sulphaquinoxaline as a coccidiostat. Evidence of carcass contamination by antibiotic resistant E. coli found in the gut is presented.     

A case of shigellosis caused by Shigella dysenteriae type 1 and Shigella flexneri type 5B

Shigella dysenteriae type 1 and Shigella flexneri type 5b strains were isolated as causative agents of bacterial dysentery in a patient having visited South-East Asia. Both strains are a rare finding for Bulgaria. S. dysenteriae 1 strains have not been isolated since 1962, and there were only single isolates of S. flexneri 5b. The strains were of the same antibiotic resistance patterns. Conjugation experiments showed that resistance is determined by transferrable R-plasmids having identical characteristics. It is assumed that in the patient's gut transfer of an R-plasmid occurs from E. coli of the normal flora to the pathogenic shigellae.     

Drug Resistance in Strains of Escherichia Coli Isolated from the Intestinal Tract of Pigs in Norway

Faecal samples from 95 healthy pigs and samples of jejunal content from 85 piglets suffering from colienterotoxaemia were tested for the presence of drug resistant E. coli strains. Practically all pigs in both groups harboured E. coli strains resistant to one or more of the 6 antibiotics/chemotherapeutic agents tested (Oxytetracycline, streptomycin, sulphaisodimidin, neomycin, ampicillin, chloramphenicol). Almost 100% of healthy and approx. 90% of diseased pigs harboured strains resistant to Oxytetracycline, streptomycin and sulphaisodimidin. Pigs with strains resistant to neomycin, ampicillin and chloramphenicol were less frequently found. The predominant coliform flora consisted of E. coli strains” resistant to Oxytetracycline, streptomycin and sulphaisodimidin in 71% to 81% of diseased pigs and in 47% to 69% of the healthy pigs. In diseased pigs ¾ of the animals had a coliform flora dominated by neomycinresistant E. coli strains. Of the 721 resistant E. coli strains isolated from healthy pigs, 11% were single resistant while the corresponding figure for the 518 resistant strains isolated from diseased pigs was 6%. Thus 89% and 94% of strains showed simultaneous resistance to 2 or more antibiotics. E. coli strains resistant to 3 or more drugs were found in approx. 60% and 70% of the isolates from healthy and diseased animals, respectively. Oxytetracycline/streptomycin/sulphaisodimidin resistance was most commonly found, approx. 22% and 38% of the strains from healthy and diseased pigs, respectively, showing this resistance pattern. Transmission of drug resistance which was examined in E. coli strains originating from the diseased pigs was demonstrated in approx. 76% of the isolates. The incidence of drug resistance transfer in single, double, triple and quadruple resistant strains was 11%, 68%, 97% and 98%, respectively.     

Prevention of drug-resistant Escherichia coli colonization in chickens by treatment with a faecal fluid

This study was performed to prove that intestinal colonization in chickens by resistant Escherichia coli strains present in the environment might be prevented when faeces in which sensitive E. coli strains were dominant was administered to newly hatched chicks. The appearance of resistant E. coli strains was markedly reduced. Escherichia coli O49:H12 was the sensitive E. coli strain which formed the major colonizer in the intestinal tract. In young chickens, this strain persisted as a major component, and even when it was a minor colonizer in the faecal fluid administered, it appeared as a major component soon afterwards. This strain is considered to be a good colonizer in the gut of young chickens.     

Prevention of drug-resistant Escherichia coli colonization in chickens by treatment with a faecal fluid

This study was performed to prove that intestinal colonization in chickens by resistant Escherichia coli strains present in the environment might be prevented when faeces in which sensitive E. coli strains were dominant was administered to newly hatched chicks. The appearance of resistant E. coli strains was markedly reduced. Escherichia coli O49:H12 was the sensitive E. coli strain which formed the major colonizer in the intestinal tract. In young chickens, this strain persisted as a major component, and even when it was a minor colonizer in the faecal fluid administered, it appeared as a major component soon afterwards. This strain is considered to be a good colonizer in the gut of young chickens.     

Attempts to Displace the Indigenous Antibiotic Resistant Gut Flora of Chicken by Feeding Sensitive Strains of Escherichia coli Prior to Slaughter

Attempts to limit the use of antibiotics have not, in general, resulted in the gut flora in farm animals becoming predominantly sensitive. Partial success has been demonstrated, however, by feeding chickens with antibiotic sensitive Escherichia coli known to be good colonizers of the chicken gut. Where feeding was done prior to slaughter a corresponding reduction in carcass contamination by resistant E. coli was observed.     

Phenotypic and genotypic characterization of encapsulated Escherichia coli isolated from blooms in two Australian lakes

Escherichia coli has long been used as an indicator organism for water quality assessment. Recently there has been an accumulation of evidence that suggests some strains of this organism are able to proliferate in the environment, a characteristic that would detract from its utility as an indicator of faecal pollution. Phenotypic and genotypic characterization of E. coli isolated from blooms in two Australian lakes, separated by a distance of approximately 200 km, identified that the blooms were dominated by three E. coli strains. A major phenotypic similarity among the three bloom strains was the presence of a group 1 capsule. Genetic characterization of a conserved region of the cps gene cluster, which encodes group 1 capsules, identified a high degree of genetic variation within the bloom isolates. This differs from previously described encapsulated E. coli strains which are highly conserved at the cps locus. The phenotypic or genotypic profiles of the bloom strains were not identified in 435 E. coli strains isolated from vertebrates. The occurrence of these encapsulated strains suggests that some E. coli have evolved a free-living lifestyle and do not require a host in order to proliferate. The presence of the same three strains in bloom events in different geographical regions of a temperate climate, and at different times, indicates that free-living E. coli strains are able to persist in these water reservoirs. This study provides further evidence of circumstances where caution is required in using E. coli as an indicator organism for water quality.     

Short-tailed stx phages exploit the conserved YaeT protein to disseminate Shiga toxin genes among enterobacteria

Infection of Escherichia coli by Shiga toxin-encoding bacteriophages (Stx phages) was the pivotal event in the evolution of the deadly Shiga toxin-encoding E. coli (STEC), of which serotype O157:H7 is the most notorious. The number of different bacterial species and strains reported to produce Shiga toxin is now more than 500, since the first reported STEC infection outbreak in 1982. Clearly, Stx phages are spreading rapidly, but the underlying mechanism for this dissemination has not been explained. Here we show that an essential and highly conserved gene product, YaeT, which has an essential role in the insertion of proteins in the gram-negative bacterial outer membrane, is the surface molecule recognized by the majority (ca. 70%) of Stx phages via conserved tail spike proteins associated with a short-tailed morphology. The yaeT gene was initially identified through complementation, and its role was confirmed in phage binding assays with and without anti-YaeT antiserum. Heterologous cloning of E. coli yaeT to enable Stx phage adsorption to Erwinia carotovora and the phage adsorption patterns of bacterial species possessing natural yaeT variants further supported this conclusion. The use of an essential and highly conserved protein by the majority of Stx phages is a strategy that has enabled and promoted the rapid spread of shigatoxigenic potential throughout multiple E. coli serogroups and related bacterial species. Infection of commensal bacteria in the mammalian gut has been shown to amplify Shiga toxin production in vivo, and the data from this study provide a platform for the development of a therapeutic strategy to limit this YaeT-mediated infection of the commensal flora.     

Establishment of specific pathogen-free (SPF) rat colonies using gnotobiotic techniques.

Gnotobiotic Wistar rats were produced using gnotobiotic techniques, which were established in the production of a SPF mouse colony, in order to establish a barrier-sustained colony. One strain of Escherichia coli, 28 strains of Bacteriodaceae (B-strains), three strains of Lactobacillus (L-strains) and a chloroform-treated fecal suspension (CHF, Clostridium mixture) were prepared from conventional Wistar rats as the microflora source. Two groups of limited-flora rats, E. coli plus B-strains and E. coli plus CHF, were produced. After confirmation that Clostridium difficile was not detected in the CHF-inoculated rats, two groups of limited-flora rats were transferred to an isolator and housed together in a cage. These rats were then orally inoculated with L-strains. The gnotobiotic rats showed colonization resistance to Pseudomonas aeruginosa, and the number of E. coli in the feces was 10(5) to 10(6)/g. The gnotobiotic rats were transferred to a barrier room as a source of intestinal flora for SPF colonies. In the SPF rats, basic cecal flora was mainly composed of Bacteroidaceae, clostridia, fusiform-shaped bacteria and lactobacilli, and did not change over a long period. Their flora became similar to that of conventional rats.     

The ecology of Escherichia coli in calves reared as dairy-cow replacements

A continual turn-over in the strains forming the majority of faecal Escherichia coli flora was demonstrated in 16 calves reared as dairy cow replacements. The incidence of antibiotic resistance among isolates, as measured by an Antibiotic Resistance Index (ARI), changed markedly with the age of the calf. The value was low initially, when the calves were 1–2 days old and housed with adult animals. It then rose rapidly during the first week after the animals had been weaned and moved into nursery pens. This change in ARI was associated with the isolation of strains resistant to four or five of the six drugs included in the sensitivity test. The ARI then fell from the third week to low levels by the time that the calves were five months of age. This fall was due to the isolation of an increasing proportion of sensitive E. coli strains. These differed from the sensitive strains which had colonized the calves in the early days of life so demonstrating that the change was not due to the reemergence of strains identified several weeks previously. The source of E. coli strains was presumed to be the calfs' environment but further investigations are required to prove this conclusively.     

Molecular Characterization of Commensal Escherichia coli Adapted to Different Compartments of the Porcine Gastrointestinal Tract

The role of Escherichia coli as a pathogen has been the focus of considerable study, while much less is known about it as a commensal and how it adapts to and colonizes different environmental niches within the mammalian gut. In this study, we characterize Escherichia coli organisms (n = 146) isolated from different regions of the intestinal tracts of eight pigs (dueodenum, ileum, colon, and feces). The isolates were typed using the method of random amplified polymorphic DNA (RAPD) and screened for the presence of bacteriocin genes and plasmid replicon types. Molecular analysis of variance using the RAPD data showed that E. coli isolates are nonrandomly distributed among different gut regions, and that gut region accounted for 25% (P < 0.001) of the observed variation among strains. Bacteriocin screening revealed that a bacteriocin gene was detected in 45% of the isolates, with 43% carrying colicin genes and 3% carrying microcin genes. Of the bacteriocins observed (H47, E3, E1, E2, E7, Ia/Ib, and B/M), the frequency with which they were detected varied with respect to gut region for the colicins E2, E7, Ia/Ib, and B/M. The plasmid replicon typing gave rise to 25 profiles from the 13 Inc types detected. Inc F types were detected most frequently, followed by Inc HI1 and N types. Of the Inc types detected, 7 were nonrandomly distributed among isolates from the different regions of the gut. The results of this study indicate that not only may the different regions of the gastrointestinal tract harbor different strains of E. coli but also that strains from different regions have different characteristics.     

Growth of Escherichia coli in a pulp and cardboard mill

The coliform flora of a pulp and cardboard mill that uses birch as the raw material and ammonium sulphate as the process chemical was studied. Escherichia coli was observed to multiply in the mill. It persisted as the dominant thermotolerant coliform in the effluent. Klebsiellae were encountered among total coliforms only. The E. coli strains isolated had the biochemical characteristics and maximum growth temperatures typical to the species. However, serotyping and hemolysin test differentiated these strains from pathogenic and fecal E. coli.     

Bacterial metabolism of natural and synthetic sex hormones undergoing enterohepatic circulation

Steroids undergoing enterohepatic circulation are exposed to bacterial metabolism particularly by obligate anaerobes which account for 99.99% of the fecal flora. The most common transformation is hydrolysis of conjugated steroids. The glucuronidases are synthesized by Escherichia coli and Bacteroides species. The bacterial catabolism of unconjugated steroids may be considered under several headings: 1. Reduction of ring-A due to clostridia species synthesizing specific enzymes; C. paraputrificum, 3 alpha,5 beta-reductase; C. innocuum, 3 beta,5 beta-reductase; and a new species C.J-1, 3 beta,5 alpha-reductase. 2. Reduction of the delta 5 bond by human fecal flora. The specific strain(s) synthesizing the enzyme have not yet been identified. 3. Reduction of 17-keto estrogens by the above mentioned ring-A reducing clostridia and by Eubacterium lentum. 4. Reduction of 17-keto androstenes by Bacteroides fragilis. 5. Desmolase mediated side chain cleavage at C17-C20 position of 17 alpha-hydroxysteroids by two new species Clostridium scindens and Eubacterium desmolans isolated from human and cat fecal flora respectively and by Clostridium cadavaris isolated from New York City sewage. 6. And 16 alpha- and 21-dehydroxylase by E. lentum a normal inhabitant of the human gut; it is the only organism known to synthesize 16 alpha- or 21-dehydroxylases. Due to the high specificity of the enzymes and the simplicity of extracting the metabolites, biosynthesis of reference compounds and radioimmunoassay reagents is practical and inexpensive. The enzymes can also be used for titration of specific bacterial strains in fecal flora and as markers for bacterial identification in particular for the strains difficult to be defined by regular biochemical reactions.     

Target genes for virulence assessment of Escherichia coli isolates from water, food and the environment

The widespread species Escherichia coli includes a broad variety of different types, ranging from highly pathogenic strains causing worldwide outbreaks of severe disease to avirulent isolates which are part of the normal intestinal flora or which are well characterized and safe laboratory strains. The pathogenicity of a given E. coli strain is mainly determined by specific virulence factors which include adhesins, invasins, toxins and capsule. They are often organized in large genetic blocks either on the chromosome ('pathogenicity islands'), on large plasmids or on phages and can be transmitted horizontally between strains. In this review we summarize the current knowledge of the virulence attributes which determine the pathogenic potential of E. coli strains and the methodology available to assess the virulence of E. coli isolates. We also focus on a recently developed procedure based on a broad-range detection system for E. coli-specific virulence genes that makes it possible to determine the potential pathogenicity and its nature in E. coli strains from various sources. This makes it possible to determine the pathotype of E. coli strains in medical diagnostics, to assess the virulence and health risks of E. coli contaminating water, food and the environment and to study potential reservoirs of virulence genes which might contribute to the emergence of new forms of pathogenic E. coli.