scholarly journals The LEE1 Promoters from both Enteropathogenic and Enterohemorrhagic Escherichia coli Can Be Activated by PerC-Like Proteins from Either Organism

2005 ◽  
Vol 187 (2) ◽  
pp. 458-472 ◽  
Author(s):  
Megan E. Porter ◽  
Paul Mitchell ◽  
Andrew Free ◽  
David G. E. Smith ◽  
David L. Gally
Keyword(s):  
Escherichia Coli ◽  
Transcriptional Start Site ◽  
Enterohemorrhagic Escherichia Coli ◽  
Site Directed Mutagenesis ◽  
E Coli ◽  
Vitro Binding ◽  
Multiple Copies ◽  
Enterocyte Effacement ◽  
Genome Analyses

ABSTRACT The PerC protein of enteropathogenic Escherichia coli (EPEC), encoded by the pEAF plasmid, is an activator of the locus of enterocyte effacement (LEE) pathogenicity island via the LEE1 promoter. It has been assumed that the related LEE-containing pathogen enterohemorrhagic E. coli (EHEC) lacks PerC-dependent activation due to utilization of an alternative LEE1 promoter and lack of a perC gene. However, we show here that EPEC PerC can activate both the EPEC and EHEC LEE1 promoters and that the major transcriptional start site is similarly located in both organisms. Moreover, a PerC-like protein family identified from EHEC genome analyses, PerC1 (also termed PchABC), can also activate both promoters in a manner similar to that of EPEC PerC. The perC1 genes are carried by lambdoid prophages, which exist in multiple copies in different EHEC strains, and have a variable flanking region which may affect their expression. Although individual perC1 copies appear to be poorly expressed, the total perC1 expression level from a strain encoding multiple copies approaches that of perC in EPEC and may therefore contribute significantly to LEE1 activation. Alignment of the protein sequences of these PerC homologues allows core regions of the PerC protein to be identified, and we show by site-directed mutagenesis that these core regions are important for function. However, purified PerC protein shows no in vitro binding affinity for the LEE1 promoter, suggesting that other core E. coli proteins may be involved in its mechanism of activation. Our data indicate that the nucleoid-associated protein IHF is one such protein.

scholarly journals Translationskopplung via Termination-Reinitiation in Archaea und Bacteria

2021 ◽  
Author(s):  
◽  
Madeleine Huber
Keyword(s):  
Escherichia Coli ◽  
De Novo ◽  
Stop Codon ◽  
Haloferax Volcanii ◽  
Site Directed Mutagenesis ◽  
Start Codon ◽  
Directed Mutagenesis ◽  
Ribosome Display ◽  
E Coli

Operons wurden zuerst im Jahre 1961 beschrieben. Bis heute ist bekannt, dass die prokaryotischen Domänen Bacteria und Archaea Gene sowohl in monocistronischen als auch in bi- oder polycistronischen Transkripten exprimieren können. Häufig überlappen Gene sogar in ihren Sequenzen. Diese überlappenden Genpaare stehen nicht in Korrelation mit der Kompaktheit ihres Genoms. Das führt zu der Annahme, dass eine Art der Regulation vorliegt, welche weitere Proteine oder Gene nicht benötigt. Diese könnte eine gekoppelte Translation sein. Das bedeutet die Translation des stromabwärts-liegenden Gens ist abhängig von der Translation eines stromaufwärts-liegenden Gens. Diese Abhängigkeit kann zum Beispiel durch lang reichende Sekundärstrukturen entstehen, bei welchen Ribosomenbindestellen (RBS) des stromabwärts-liegenden Gens blockiert sind. Die de novo-Initiation am stromabwärts-liegenden Gen kann nur stattfinden, wenn das erste Gen translatiert wird und dabei die Sekundärstruktur an der RBS aufgeschmolzen wird. Für Genpaare in E. coli ist dieser Mechanismus gut untersucht. Ein anderes Beispiel für die Translationskopplung ist die Termination-Reinitiation, bei welcher ein Ribosom das erste Gen translatiert bis zum Stop-Codon, dort terminiert und direkt am stromabwärts-liegenden Start-Codon reinitiiert. Der Mechanismus via Termination-Reinitiation ist bis jetzt nur für eukaryontische Viren beschrieben worden. Im Gegensatz zu einer Kopplung über Sekundärstrukturen kommt es bei der Termination-Reinitiation am stromabwärts-liegenden Gen nicht zu einer de novo-Initiation sondern eine Reinitiation des Ribosoms findet statt. Diese Arbeit analysiert jene Art der Translationskopplung an Genen polycistronischer mRNAs in jeweils einem Modellorganismus als Vertreter der Archaea (Haloferax volcanii) und Bacteria (Escherichia coli). Hierfür wurden Reportergenvektoren erstellt, welche die überlappenden Genpaare an Reportergene fusionierten. Für diese Reportergene ist es möglich die Transkriptmenge zu quantifizieren sowie für die exprimierten Proteine Enzymassays durchgeführt werden können. Aus beiden Werten können Translationseffizienzen berechnet werden indem jeweils die Enzymaktivität pro Transkriptmenge ermittelt wird. Durch ein prämatures Stop-Codon in diesen Konstrukten ist es möglich zu unterscheiden ob es für die Translation des zweiten Gens essentiell ist, dass das Ribosom den Überlapp erreicht. Hiermit konnte für neun Genpaare in H. volcanii und vier Genpaare in E. coli gezeigt werden, dass eine Art der Kopplung stattfindet bei der es sich um eine Termination-Reinitiation handelt. Des Weiteren wurde analysiert, welche Auswirkungen intragene Shine-Dalgarno Sequenzen bei dem Event der Translationskopplung besitzen. Durch die Mutation solcher Motive und dem Vergleich der Translationseffizienzen der Konstrukte, mit und ohne einer SD Sequenz, wird für alle analysierten Genpaare beider Modellorganismen gezeigt, dass die SD Sequenz einen Einfluss auf diese Art der Kopplung hat. Zwischen den Genpaaren ist dieser Einfluss jedoch stark variabel. Weiterhin wurde der maximale Abstand zwischen zwei bicistronischen Genen untersucht, für welchen Translationskopplung via Termination-Reinitiation noch stattfinden kann. Hierfür wird durch site-directed mutagenesis jeweils ein prämatures Stop-Codon im stromaufwärts-liegenden Gen eingebracht, welches den intergenen Abstand zwischen den Genen in den jeweiligen Konstrukten vergrößert. Der Vergleich aller Konstrukte eines Genpaars zeigt in beiden Modellorganismen, dass die Termination-Reinitiation vom intergenen Abstand abhängig ist und die Translationseffizienz des stromabwärts-liegenden Reporters bereits ab 15 Nukleotiden Abstand abnimmt. Eine weitere Fragestellung dieser Arbeit war es, den genauen Mechanismus der Termination-Reinitiation zu analysieren. Für Ribosomen gibt es an der mRNA nach der Termination der Translation zwei Möglichkeiten: Entweder als 70S Ribosom bestehen zu bleiben und ein weiteres Start-Codon auf der mRNA zu suchen oder in seine beiden Untereinheiten zu dissoziieren, während die 50S Untereinheit die mRNA verlässt und die 30S Untereinheit über Wechselwirkungen an der mRNA verbleiben kann. Um diesen Mechanismus auf molekularer Ebene zu untersuchen, wird ein Versuchsablauf vorgestellt. Dieser ermöglicht das Event bei der Termination-Reinitiation in vitro zu analysieren. Eine Unterscheidung von 30S oder 70S Ribosomen bei der Reinitiation der Translation des stromabwärts-liegenden Gens wird ermöglicht. Die Idee dabei basiert auf einem ribosome display, bei welchem Translationskomplexe am Ende der Translation nicht in ihre Bestandteile zerfallen können, da die eingesetzte mRNA kein Stop-Codon enthält Der genaue Versuchsablauf, die benötigten Bestandteile sowie proof-of-principal Versuche sind in der Arbeit dargestellt und mögliche Optimierungen werden diskutiert.

scholarly journals Fitness of Enterohemorrhagic Escherichia coli (EHEC)/Enteroaggregative E. coli O104:H4 in Comparison to That of EHEC O157: Survival Studies in Food andIn Vitro

2016 ◽  
Vol 82 (21) ◽  
pp. 6326-6334 ◽  
Author(s):  
Christina Böhnlein ◽  
Jan Kabisch ◽  
Diana Meske ◽  
Charles M. A. P. Franz ◽  
Rohtraud Pichner
Keyword(s):  
Escherichia Coli ◽  
Meat Product ◽  
Enterohemorrhagic Escherichia Coli ◽  
Content Type ◽  
E Coli ◽  
Adverse Conditions ◽  
Log Reduction ◽  
Fermented Sausages ◽  
Survival Studies

ABSTRACTIn 2011, one of the world's largest outbreaks of hemolytic-uremic syndrome (HUS) occurred, caused by a rareEscherichia coliserotype, O104:H4, that shared the virulence profiles of Shiga toxin-producingE. coli(STEC)/enterohemorrhagicE. coli(EHEC) and enteroaggregativeE. coli(EAEC). The persistence and fitness factors of the highly virulent EHEC/EAEC O104:H4 strain, grown either in food orin vitro, were compared with those ofE. coliO157 outbreak-associated strains. The log reduction rates of the different EHEC strains during the maturation of fermented sausages were not significantly different. Both the O157:NM and O104:H4 serotypes could be shown by qualitative enrichment to be present after 60 days of sausage storage. Moreover, the EHEC/EAEC O104:H4 strain appeared to be more viable thanE. coliO157:H7 under conditions of decreased pH and in the presence of sodium nitrite. Analysis of specific EHEC strains in experiments with an EHEC inoculation cocktail showed a dominance of EHEC/EAEC O104:H4, which could be isolated from fermented sausages for 60 days. Inhibitory activities of EHEC/EAEC O104:H4 toward severalE. colistrains, including serotype O157 strains, could be determined. Our study suggests that EHEC/EAEC O104:H4 is well adapted to the multiple adverse conditions occurring in fermented raw sausages. Therefore, it is strongly recommended that STEC strain cocktails composed of several serotypes, instead ofE. coliO157:H7 alone, be used in food risk assessments. The enhanced persistence of EHEC/EAEC O104:H4 as a result of its robustness, as well as the production of bacteriocins, may account for its extraordinary virulence potential.IMPORTANCEIn 2011, a severe outbreak caused by an EHEC/EAEC serovar O104:H4 strain led to many HUS sequelae. In this study, the persistence of the O104:H4 strain was compared with those of other outbreak-relevant STEC strains under conditions of fermented raw sausage production. Both O157:NM and O104:H4 strains could survive longer during the production of fermented sausages thanE. coliO157:H7 strains.E. coliO104:H4 was also shown to be well adapted to the multiple adverse conditions encountered in fermented sausages, and the secretion of a bacteriocin may explain the competitive advantage of this strain in an EHEC strain cocktail. Consequently, this study strongly suggests that enhanced survival and persistence, and the presumptive production of a bacteriocin, may explain the increased virulence of the O104:H4 outbreak strain. Furthermore, this strain appears to be capable of surviving in a meat product, suggesting that meat should not be excluded as a source of potentialE. coliO104:H4 infection.

scholarly journals Shiga Toxin Gene Loss and Transfer In Vitro and In Vivo during Enterohemorrhagic Escherichia coli O26 Infection in Humans

2007 ◽  
Vol 73 (10) ◽  
pp. 3144-3150 ◽  
Author(s):  
Martina Bielaszewska ◽  
Rita Prager ◽  
Robin Köck ◽  
Alexander Mellmann ◽  
Wenlan Zhang ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Shiga Toxin ◽  
Gene Loss ◽  
Pulsed Field Gel Electrophoresis ◽  
Enterohemorrhagic Escherichia Coli ◽  
Toxin Gene ◽  
Shiga Toxins ◽  
E Coli

ABSTRACT Escherichia coli serogroup O26 consists of enterohemorrhagic E. coli (EHEC) and atypical enteropathogenic E. coli (aEPEC). The former produces Shiga toxins (Stx), major determinants of EHEC pathogenicity, encoded by bacteriophages; the latter is Stx negative. We have isolated EHEC O26 from patient stools early in illness and aEPEC O26 from stools later in illness, and vice versa. Intrapatient EHEC and aEPEC isolates had quite similar pulsed-field gel electrophoresis (PFGE) patterns, suggesting that they might have arisen by conversion between the EHEC and aEPEC pathotypes during infection. To test this hypothesis, we asked whether EHEC O26 can lose stx genes and whether aEPEC O26 can be lysogenized with Stx-encoding phages from EHEC O26 in vitro. The stx 2 loss associated with the loss of Stx2-encoding phages occurred in 10% to 14% of colonies tested. Conversely, Stx2- and, to a lesser extent, Stx1-encoding bacteriophages from EHEC O26 lysogenized aEPEC O26 isolates, converting them to EHEC strains. In the lysogens and EHEC O26 donors, Stx2-converting bacteriophages integrated in yecE or wrbA. The loss and gain of Stx-converting bacteriophages diversifies PFGE patterns; this parallels findings of similar but not identical PFGE patterns in the intrapatient EHEC and aEPEC O26 isolates. EHEC O26 and aEPEC O26 thus exist as a dynamic system whose members undergo ephemeral interconversions via loss and gain of Stx-encoding phages to yield different pathotypes. The suggested occurrence of this process in the human intestine has diagnostic, clinical, epidemiological, and evolutionary implications.

scholarly journals Cross-Reactive Protection against Enterohemorrhagic Escherichia coli Infection by Enteropathogenic E. coli in a Mouse Model

2011 ◽  
Vol 79 (6) ◽  
pp. 2224-2233 ◽  
Author(s):  
Carla Calderon Toledo ◽  
Ida Arvidsson ◽  
Diana Karpman
Keyword(s):  
Escherichia Coli ◽  
Weight Loss ◽  
Mouse Model ◽  
Enterohemorrhagic Escherichia Coli ◽  
Epec Strain ◽  
Content Type ◽  
E Coli ◽  
Renal Histology ◽  
Escherichia Coli Infection ◽  
Enterocyte Effacement

ABSTRACTEnteropathogenicEscherichia coli(EPEC) and enterohemorrhagicE. coli(EHEC) are related attaching and effacing (A/E) pathogens. The genes responsible for the A/E pathology are carried on a chromosomal pathogenicity island termed the locus of enterocyte effacement (LEE). Both pathogens share a high degree of homology in the LEE and additional O islands. EHEC prevalence is much lower in areas where EPEC is endemic. This may be due to the development of antibodies against common EPEC and EHEC antigens. This study investigated the hypothesis that EPEC infections may protect against EHEC infections. We used a mouse model to inoculate BALB/c mice intragastrically, first with EPEC and then with EHEC (E. coliO157:H7). Four control groups received either a nonpathogenicE. coli(NPEC) strain followed by EHEC (NPEC/EHEC), phosphate-buffered saline (PBS) followed by EHEC (PBS/EHEC), EPEC/PBS, or PBS/PBS. Mice were monitored for weight loss and symptoms. EPEC colonized the intestine after challenge, and mice developed serum antibodies to intimin andE. colisecreted protein B (encoded in the LEE). Prechallenge with an EPEC strain had a protective effect after EHEC infection, as only a few mice developed mild symptoms, from which they recovered. These mice had an increase in body weight similar to that in control animals, and tissue morphology exhibited mild intestinal changes and normal renal histology. All mice that were not prechallenged with the EPEC strain developed mild to severe symptoms after EHEC infection, with weight loss as well as intestinal and renal histopathological changes. These data suggest that EPEC may protect against EHEC infection in this mouse model.

scholarly journals Recycling of Shiga Toxin 2 Genes in Sorbitol-Fermenting Enterohemorrhagic Escherichia coli O157:NM

2007 ◽  
Vol 74 (1) ◽  
pp. 67-72 ◽  
Author(s):  
Alexander Mellmann ◽  
Shan Lu ◽  
Helge Karch ◽  
Jian-guo Xu ◽  
Dag Harmsen ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Shiga Toxin ◽  
Integration Site ◽  
Hot Spot ◽  
Blot Hybridization ◽  
Enterohemorrhagic Escherichia Coli ◽  
Biologically Active ◽  
E Coli ◽  
Shiga Toxin 2

ABSTRACT Using colony blot hybridization with stx 2 and eae probes and agglutination in anti-O157 lipopolysaccharide serum, we isolated stx 2-positive and eae-positive sorbitol-fermenting (SF) enterohemorrhagic Escherichia coli (EHEC) O157:NM (nonmotile) strains from initial stool specimens and stx-negative and eae-positive SF E. coli O157:NM strains from follow-up specimens (collected 3 to 8 days later) from three children. The stx-negative isolates from each patient shared with the corresponding stx 2-positive isolates fliC H7, non-stx virulence traits, and multilocus sequence types, which indicates that they arose from the stx 2-positive strains by loss of stx 2 during infection. Analysis of the integrity of the yecE gene, a possible stx phage integration site in EHEC O157, in the consecutive stx 2-positive and stx-negative isolates demonstrated that yecE was occupied in stx 2-positive but intact in stx-negative strains. It was possible to infect and lysogenize the stx-negative E. coli O157 strains in vitro using an stx 2-harboring bacteriophage from one of the SF EHEC O157:NM isolates. The acquisition of the stx 2-containing phage resulted in the occupation of yecE and production of biologically active Shiga toxin 2. We conclude that the yecE gene in SF E. coli O157:NM is a hot spot for excision and integration of Shiga toxin 2-encoding bacteriophages. SF EHEC O157:NM strains and their stx-negative derivatives thus represent a highly dynamic system that can convert in both directions by the loss and gain of stx 2-harboring phages. The ability to recycle stx 2, a critical virulence trait, makes SF E. coli O157:NM strains ephemeral EHEC that can exist as stx-negative variants during certain phases of their life cycle.

scholarly journals Enterohemorrhagic Escherichia coli Virulence Regulation by Two Bacterial Adrenergic Kinases, QseC and QseE

2011 ◽  
Vol 80 (2) ◽  
pp. 688-703 ◽  
Author(s):  
Jacqueline Njoroge ◽  
Vanessa Sperandio
Keyword(s):  
Escherichia Coli ◽  
Response Regulator ◽  
Enterohemorrhagic Escherichia Coli ◽  
Dependent Manner ◽  
Content Type ◽  
E Coli ◽  
Virulence Regulation ◽  
Sensor Kinases ◽  
Virulence Factor Gene ◽  
Enterocyte Effacement

ABSTRACTThe human pathogen enterohemorrhagicEscherichia coli(EHEC) O157:H7 has two histidine sensor kinases, QseC and QseE, which respond to the mammalian adrenergic hormones epinephrine and norepinephrine by increasing their autophosphorylation. Although QseC and QseE are present in nonpathogenic strains ofE. coli, EHEC exploits these kinases for virulence regulation. To further investigate the full extent of epinephrine and its sensors' impact on EHEC virulence, we performed transcriptomic and phenotypic analyses of single and double deletions ofqseCandqseEgenes in the absence or presence of epinephrine. We showed that in EHEC, epinephrine sensing seems to occur primarily through QseC and QseE. We also observed that QseC and QseE regulate expression of the locus of enterocyte effacement (LEE) genes positively and negatively, respectively. LEE activation, which is required for the formation of the characteristic attaching and effacing (A/E) lesions by EHEC on epithelial cells, is epinephrine dependent. Regulation of the LEE and the non-LEE-contained virulence factor genenleAby QseE is indirect, through transcription inhibition of the RcsB response regulator. Finally, we show that coincubation of HeLa cells with epinephrine increases EHEC infectivity in a QseC- and QseE-dependent manner. These results genetically and phenotypically map the contributions of the two adrenergic sensors QseC and QseE to EHEC pathogenesis.

scholarly journals Enterohemorrhagic Escherichia coli O157:H7 gal Mutants Are Sensitive to Bacteriophage P1 and Defective in Intestinal Colonization

2006 ◽  
Vol 75 (4) ◽  
pp. 1661-1666 ◽  
Author(s):  
Theresa Deland Ho ◽  
Matthew K. Waldor
Keyword(s):  
Escherichia Coli ◽  
Genetic Manipulation ◽  
Enterohemorrhagic Escherichia Coli ◽  
Deletion Strain ◽  
Growth Defect ◽  
Intestinal Colonization ◽  
Wild Type ◽  
E Coli ◽  
O Antigen

ABSTRACT Enterohemorrhagic Escherichia coli (EHEC), especially E. coli O157:H7, is an emerging cause of food-borne illness. Unfortunately, E. coli O157 cannot be genetically manipulated using the generalized transducing phage P1, presumably because its extensive O antigen obscures the P1 receptor, the lipopolysaccharide (LPS) core subunit. The GalE, GalT, GalK, and GalU proteins are necessary for modifying galactose before it can be assembled into the repeating subunit of the O antigen. Here, we constructed E. coli O157:H7 gal mutants which presumably have little or no O antigen. These strains were able to adsorb P1. P1 lysates grown on the gal mutant strains could be used to move chromosomal markers between EHEC strains, thereby facilitating genetic manipulation of E. coli O157:H7. The gal mutants could easily be reverted to a wild-type Gal+ strain using P1 transduction. We found that the O157:H7 galETKM::aad-7 deletion strain was 500-fold less able to colonize the infant rabbit intestine than the isogenic Gal+ parent, although it displayed no growth defect in vitro. Furthermore, in vivo a Gal+ revertant of this mutant outcompeted the galETKM deletion strain to an extent similar to that of the wild type. This suggests that the O157 O antigen is an important intestinal colonization factor. Compared to the wild type, EHEC gal mutants were 100-fold more sensitive to a peptide derived from bactericidal permeability-increasing protein, a bactericidal protein found on the surface of intestinal epithelial cells. Thus, one way in which the O157 O antigen may contribute to EHEC intestinal colonization is to promote resistance to host-derived antimicrobial polypeptides.

scholarly journals Type 2 Secretion Promotes Enterohemorrhagic Escherichia coli Adherence and Intestinal Colonization

2008 ◽  
Vol 76 (5) ◽  
pp. 1858-1865 ◽  
Author(s):  
Theresa D. Ho ◽  
Brigid M. Davis ◽  
Jennifer M. Ritchie ◽  
Matthew K. Waldor
Keyword(s):  
Escherichia Coli ◽  
Rabbit Model ◽  
Enterohemorrhagic Escherichia Coli ◽  
Growth Defect ◽  
Intestinal Colonization ◽  
Colonization Factor ◽  
Enterocyte Effacement ◽  
Type 2 Secretion

ABSTRACT Enterohemorrhagic Escherichia coli (EHEC) is a noninvasive food-borne pathogen that colonizes the distal ileum and colon. Proteins encoded in the EHEC locus of enterocyte effacement (LEE) pathogenicity island are known to contribute to this pathogen's adherence to epithelial cells and intestinal colonization. The role of non-LEE-encoded proteins in these processes is not as clear. We found that the Z2053 gene (designated adfO here), a gene located in a cryptic EHEC prophage, exhibits similarity to adherence and/or colonization factor genes found in several other enteric pathogens. An EHEC adfO mutant exhibited marked reductions in adherence to HeLa cells and in the secretion of several proteins into the supernatant. YodA, one of these secreted proteins, was found to be a substrate of the EHEC pO157-encoded type 2 secretion system (T2SS). Both the T2SS and YodA proved to be essential for EHEC adherence to cultured HeLa cell monolayers. Using an infant rabbit model of infection, we found that the adfO mutation did not affect colonization but that the colonization of an etpC (T2SS) mutant was reduced ∼5-fold. A strain deficient in YodA had a more severe colonization defect; however, this strain also exhibited a growth defect in vitro. Overall, our findings indicate that the pO157-encoded T2SS contributes to EHEC adherence and intestinal colonization and thus show that EHEC pathogenicity depends on type 2 secretion as well as type 3 secretion.

scholarly journals Reduction of Carriage of Enterohemorrhagic Escherichia coli O157:H7 in Cattle by Inoculation with Probiotic Bacteria

1998 ◽  
Vol 36 (3) ◽  
pp. 641-647 ◽  
Author(s):  
Tong Zhao ◽  
Michael P. Doyle ◽  
Barry G. Harmon ◽  
Cathy A. Brown ◽  
P. O. Eric Mueller ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Enterohemorrhagic Escherichia Coli ◽  
Probiotic Bacteria ◽  
Escherichia Coli O157 ◽  
Bacterial Isolates ◽  
Intestinal Tissue ◽  
Tissue Samples ◽  
E Coli ◽  
Observation Period

Bacteria inhibitory to Escherichia coli O157:H7 were isolated from cattle and evaluated for their potential for reducing carriage of E. coli O157:H7 in calves. Eighteen of 1,200 bacterial isolates from cattle feces and intestinal tissue samples were screened and determined to inhibit the growth of E. coliO157:H7 in vitro. Seventeen of the isolates were E. coli and one was Proteus mirabilis. None produced Shiga toxin. Genomic DNA fingerprinting by pulsed-field gel electrophoresis revealed 13 distinguishable profiles among the 18 isolates. Two calves inoculated perorally with a mixture of all 18 isolates (1010 CFU) appeared to be normal and did not develop signs of clinical disease throughout a 25- to 27-day observation period. These bacteria colonized segments of the gastrointestinal tract and were in feces at the termination of the experiment (25 and 27 days postinoculation) at levels of 50 to 200 CFU/g. Fifteen cannulated calves were studied to determine the efficiency of the probiotic bacteria in reducing or eliminating the carriage of E. coli O157:H7. Nine calves served as controls, with each animal receiving perorally 1010 CFU ofE. coli O157:H7. E. coliO157:H7 was detected intermittently in the rumen samples from all control animals throughout 3 weeks postinoculation, whereasE. coli O157:H7 was shed at various levels in feces continuously throughout the experiment (mean, 28 days).E. coli O157:H7 was isolated from the rumens and colons of eight of nine and nine of nine calves, respectively, at the termination of the study. Six calves each received perorally 1010 CFU of probiotic bacteria and then 2 days later received 1010 CFU of E. coli O157:H7.E. coli O157:H7 was detected in the rumen for only 9 days postinoculation in two animals, for 16 days in one animal, for 17 days in two animals, and for 29 days in one animal. E. coli O157:H7 was detected in feces for only 11 days postinoculation in one animal, for 15 days in one animal, for 17 days in one animal, for 18 days in one animal, for 19 days in one animal, and for 29 days in one animal. At the end of the experiment (mean, 30 days), E. coli O157:H7 was not recovered from the rumen of any of the six animals treated with probiotic bacteria; however, E. coli O157:H7 was recovered from the feces of one of the animals. This animal was fasted twice postinoculation. These studies indicate that selected probiotic bacteria administered to cattle prior to exposure to E. coli O157:H7 can reduce the level of carriage ofE. coli O157:H7 in most animals.

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