L-374,087, an efficacious, orally bioavailable, pyridinone acetamide thrombin inhibitor

The potential for transfer of Escherichia coli O157:H7 from contaminated ground beef to grinding equipment and the inactivation of attached cells during cleaning and sanitizing was examined. Chub-packed ground beef with lean:fat ratios of 75:25, 80:20 or 90:10 was inoculated with 6 log CFU/g or 2 log CFU/g E. coli O157:H7 strain FRIK 910. Samples were consecutively ground in a Hobart meat grinder with stainless steel (SS) chips (1 cm2) glued to the auger housing. Chips were harvested after grinding, detergent washing with or without manual scrubbing and rinsing, sanitizing in a chlorine or peroxyacetic acid sanitizer, and overnight storage. Survival of E. coli O157:H7 was evaluated both by plate count and enrichment in trypticase soy broth. Approximately 3 to 4 log CFU/cm2 were attached to the SS after grinding with all three fat contents. After washing and sanitizing in a chlorine or peroxyacetic acid sanitizer, viable bacteria were infrequently recovered by plate count. Enrichment of chips resulted in a higher survival rate with both sanitizing treatments, indicating that cell numbers below the limit of detection (5 CFU/cm2) or potentially injured organisms remained on the surface. Manual scrubbing during the washing step reduced the recovery rate. The scrubbing step also increased the number of passing scores assigned using an ATP bioluminescence assay of total residual soil on the chips sanitized in chlorine. The overall results indicate that plate counts alone may not be a reliable indicator of sanitation efficacy and may be validated by enrichment assay.     

Bacterial cross-contamination of meat during liquid nitrogen immersion freezing

Prerigor beef carcass surface tissue (BCT) was used to simulate lamb carcasses on a processing line with a 15-min liquid nitrogen (LN) immersion freezing step, and the potential for the dissemination of bacteria during freezing was examined. Streptomycin-resistant strains of Listeria innocua and Escherichia coli O157:H7 spiked into a fecal slurry were inoculated onto BCT pieces that were introduced into the freezing process to represent contaminated carcasses. Following this introduction, subsequently frozen uninoculated BCT, LN, and LN containers were examined for the inoculated organisms. In the first study, BCT samples were inoculated with ca. 7 log CFU/cm2 of both L. innocua and E. coli O157:H7, spray washed with water and frozen, distributed among uninoculated BCT, in LN for 15 min. In two separate trials, L. innocua was recovered by enrichment from all uninoculated BCT and LN samples. E. coli O157:H7 was also recovered from uninoculated BCT and LN, but this cross-contamination was more sporadic. Both species were recovered from the LN container following freezing. Attempts to enumerate cross-contaminating bacteria in the second trial indicated that contaminating levels were low (< 1.0 CFU/cm2 BCT). In a second study, a 2.0% lactic acid spray wash was used to reduce further the numbers of L. innocua introduced into the freezing system and resulted in fewer positive samples, although this organism was still recovered from many uninoculated BCT samples. When either bacterium was inoculated at lower initial levels (1.35 to 1.77 log CFU/cm2) and BCT was water or 2.0% lactic acid spray washed prior to freezing, neither L. innocua nor E. coli O157:H7 was recoverable by enrichment from uninoculated BCT, LN, or from the freezing container. Results demonstrate that bacterial cross-contamination of meat during LN immersion freezing can occur but indicate that the use of good sanitation practices and product with low microbial numbers can limit this occurrence.     

Reduction of microorganisms on citrus fruit surfaces during packinghouse processing

Citrus fruit surface microbial populations were evaluated following various packingline processes of seven Florida commercial packinghouses. At each packinghouse, six fruits (oranges or tangerines) were collected at each of four sampling points. The sampling was conducted in duplicate; thus, 336 fruit were evaluated during this survey. Average aerobic plate counts and yeast and mold counts on fruit surfaces before washing were about 4.0 log CFU/cm2 and 3.3 log CFU/cm2, respectively, and were reduced to 2.1 log CFU/cm2 and 1.3 log CFU/cm2, respectively, by packinghouse processing. Waxing alone reduced the average fruit surface aerobic plate counts and coliform counts from 3.7 log CFU/cm2 and 35.2 most probable number (MPN)/cm2, respectively, to 2.6 log CFU/cm2 and 1.4 MPN/cm2. No Escherichia coli was recovered from fruit at the end of packinghouse processing, and no salmonellae were found on fruit during the entire processing. In an inoculation study to test the effect of packinghouse processes, test organism E. coli was applied to fruit to achieve a high level (4.8 log CFU/cm2) of contamination. The average E. coli count was reduced about 2.4 log cycles by washing and rinsing with potable water (40 psi, 25 degrees C) for about 30 s. The combination of washing and waxing significantly reduced the inoculated level of E. coli from 4.8 to 1.4 log CFU/cm2.     

PCR-based DNA amplification and presumptive detection of Escherichia coli O157:H7 with an internal fluorogenic probe and the 5' nuclease (TaqMan) assay

Presumptive identification of Escherichia coli O157:H7 is possible in an individual, nonmultiplexed PCR if the reaction targets the enterohemorrhagic E. coli (EHEC) eaeA gene. In this report, we describe the development and evaluation of the sensitivity and specificity of a PCR-based 5' nuclease assay for presumptively detecting E. coli O157:H7 DNA. The specificity of the eaeA-based 5' nuclease assay system was sufficient to correctly identify all E. coli O157:H7 strains evaluated, mirroring the previously described specificity of the PCR primers. The SZ-primed, eaeA-targeted 5' nuclease detection assay was capable of rapid, semiautomated, presumptive detection of E. coli O157:H7 when >/=10(3) CFU/ml was present in modified tryptic soy broth (mTSB) or modified E. coli broth and when >/=10(4) CFU/ml was present in ground beef-mTSB mixtures. Incorporating an immunomagnetic separation (IMS) step, followed by a secondary enrichment culturing step and DNA recovery with a QIAamp tissue kit (Qiagen), improved the detection threshold to >/=10(2) CFU/ml. Surprisingly, immediately after IMS, the sensitivity of culturing on sorbitol MacConkey agar containing cefeximine and tellurite (CT-SMAC) was such that identifiable colonies were demonstrated only when >/=10(4) CFU/ml was present in the sample. Several factors that might be involved in creating these false-negative CT-SMAC culture results are discussed. The SZ-primed, eaeA-targeted 5' nuclease detection system demonstrated that it can be integrated readily into standard culturing procedures and that the assay can be useful as a rapid, automatable process for the presumptive identification of E. coli O157:H7 in ground beef and potentially in other food and environmental samples.     

Efficacy of spray application of chlorinated water in killing pathogenic bacteria on raw apples, tomatoes, and lettuce

Washing whole and cut produce by dipping or submerging in chlorinated water has a sanitizing effect, although reduction in microbial populations is minimal and is usually less than 100-fold. A study was undertaken to evaluate the efficacy of a spray application of chlorine in killing Salmonella, Escherichia coli O157:H7, Listeria monocytogenes, yeasts and molds, and total aerobic mesophilic microorganisms on whole apples, tomatoes, and lettuce leaves. Inoculated produce was treated (sprayed and then soaked) with water (control) or solutions containing 200 or 2,000 ppm of chlorine for 0, 1, 3, 5, or 10 min, rinsed with sterile water, and analyzed for populations (CFU/cm2) of target microorganisms. Compared to the control treatment, further reductions in numbers of pathogens of 0.35 to 2.30 log CFU/cm2 were achieved by treatment with chlorine. Chlorine was generally more effective at 2,000 ppm than at 200 ppm. Inactivation of microorganisms occurred essentially within 1 min after application of chlorine. These reductions are significant relative to populations of pathogenic microorganisms that may be present on produce. Spray application of chlorine to raw produce at food service or household levels may be a suitable, and more convenient, alternative to treatment by dipping or submersion.     

Effect of dietary stress on fecal shedding of Escherichia coli O157:H7 in calves

Two groups of calves were subjected to dietary stress by withholding of food beginning 1 or 14 days after inoculation with 10(10) CFU of Escherichia coli O157:H7. Following treatment, neither group had a significant increase in fecal shedding of E. coli O157:H7. A third group of calves had food withheld for 48 h prior to inoculation with 10(7) CFU of E. coli O157:H7. These calves were more susceptible to infection and shed significantly more E. coli O157:H7 organisms than calves maintained on a normal diet.     

New antiepileptic drugs

Survival of Escherichia coli O157:H7 strains QA 326, and ATCC 43889, 43894, and 43895 after freezing (-20 degrees C, 24 h) and thawing (4 degrees C for 12 h, 23 degrees C for 3 h, or microwave heating of 700 W for 120 s) in ground beef patties was determined by reference most probable number (MPN), hydrophobic grid membrane filter SD-39 agar, and sorbitol MacConkey agar (SMA) spread-plating methods. Populations decreased from 0.62 to 2.52 log10 CFU/g, with the extent varying significantly by strain. Strain QA 326 populations almost always decreased the most, up to 1.87 log10 CFU/g more than the least sensitive strain. Microwave heating was the most lethal thawing treatment for strain QA 326, and 4 degrees C thawing was the most lethal treatment for strain ATCC 43894. Thawing treatments varied in relative lethality for the other two strains. For strain QA 326 (4 degrees C and microwave thaw treatments) and strain ATCC 43889 (4 and 23 degrees C thawing), the enumeration method significantly affected a population decrease. The SD-39 agar method best recovered strain QA 326 while the SD-39 agar method and the reference MPN method best recovered strain ATCC 43889 after 4 and 23 degrees C thawing, respectively. The greatest difference in population decrease measured by any two methods was 0.58 log10 CFU/g. Results showed (i) a wide range in freeze-thaw sensitivity among E. coli O157:H7 strains, (ii) no thawing method had consistently and significantly greater lethality, and (iii) the reference MPN, SD-39 agar, and SMA methods differed little in ability to enumerate E. coli O157:H7.     

Prevalence of enterohemorrhagic Escherichia coli O157:H7 in a survey of dairy herds

The prevalence of Escherichia coli O157:H7 in dairy herds is poorly understood, even though young dairy animals have been reported to be a host. From February to May 1993, 662 fecal samples from 50 control herds in 14 states, and from June to August 1993, 303 fecal samples from 14 case herds in 11 states were collected for isolation of E. coli O157:H7. Case herds were those in which E. coli O157:H7 was isolated from preweaned calves in a previous U.S. Department of Agriculture study, whereas control herds from which E. coli O157:H7 had not been isolated previously were randomly selected from the same states as case herds. Among the control herds, E. coli O157:H7 was isolated from 6 of 399 calves (1.5%) that were between 24 h old and the age of weaning and from 13 of 263 calves (4.9%) that were between the ages of weaning and 4 months. Eleven of 50 control herds (22%) were positive. Among the case herds, E. coli O157:H7 was isolated from 5 of 171 calves (2.9%) that were between 24 h old and the age of weaning and from 7 of 132 calves (5.3%) that were between the ages of weaning and 4 months. Seven of 14 case herds (50%) were positive. Sixteen of 31 isolates were obtained by direct plating, with populations ranging from 10(3) to 10(5) CFU/g. Fifteen of 31 isolates were isolated by enrichment only. Nineteen of the isolates produced both verocytotoxin 1 (VT-1) and VT-2, whereas 12 produced VT-2 only.     

Recovery of thermally-stressed Escherichia coli O157:H7 by media supplemented with pyruvate

Unheated and heat-stressed (57 degrees C, 50 min and 60 min) cells of Escherichia coli O157:H7, were enumerated using three media supplemented with 1% sodium pyruvate (NaPyr): plate count agar (PCA), tryptic soy agar (TSA) and phenol red sorbitol agar (PhRSA) using the spread plate method. The medium recovering the greatest numbers of severely heated E. coli O157:H7 was PCA with 1% NaPyr. Recovery of heat stressed E. coli O157:H7 on this medium was significantly higher (P < 0.05) than the two other media with pyruvate: 16.3% (50 min heating) and 0.55% (60 min heating) of the total population was recovered with TSA + 1% NaPyr when compared to those numbers found on PCA + 1% NaPyr. The ability of PhRSA + 1% NaPyr to recover heat-stressed E. coli O157:H7 was similar to that of TSA + 1% NaPyr. Using PhRSA + 1% NaPyr media. 12.9% (50 min heating) and 0.61% (60 min heating) of the total population were recovered when compared with the cells enumerated on PCA + 1% NaPyr. Recovery of the heat-stressed cells using the spread plate method was greater than using pour plate method. Recovery was significantly higher (P < 0.05) on the spread plates for highly stressed E. coli O157:H7(1.2 log) heated for 60 min than on the pour plates. Overall, the populations on the TSA spread and pour plates were low compared with the same heat-stressed cells recovered on media containing pyruvate. The     

Immunomagnetic separation and flow cytometry for rapid detection of Escherichia coli O157:H7

A rapid method for detecting Escherichia coli O157:H7 combining immunomagnetic beads (IMB) and flow cytometry was developed. Labeling antigens separated by IMB with fluorescent antibody enabled the detection of < 10(3) CFU bacteria per ml in pure culture. The optimum concentration of magnetic beads for flow cytometry was lower (ca. 10(5) particles per ml) than that reported for conventional IMB assay (more than 6 x 10(6) to 8 x 10(6) particles per ml). Immunomagnetic separation and flow cytometry (IMFC) were evaluated for detecting E. coli O157:H7 in the presence of a competing microorganism and for detecting antibodies in potassium phosphate buffer. The total assay time from separating antigens with IMB to analyzing with flow cytometry was about 1 h. IMFC detected 10(3) to 10(4) CFU of E. coli O157:H7 per ml in ground beef enrichment broth and could effectively discriminate between E. coli O157:H7 and competing natural flora. The new assay system provides another approach to separation and detection of low populations of pathogens and shows potential for detecting low concentrations of toxins and other soluble antigens directly from food in a short time.     

Molecular lipophilicity potential by CLIP, a reliable tool for the description of the 3D distribution of lipophilicity: application to 3-phenyloxazolidin-2-one, a prototype series of reversible MAOA inhibitors

Six methods were compared for detection of three strains of Escherichia coli O157:H7 in enrichments of inoculated apple juice. Juice was inoculated at levels varying from 0.1 to 100 CFU/ml and centrifuged after overnight storage at 4 degrees C, and pellets were incubated at 37 degrees C in nonselective enrichment broth. At hourly intervals between 5 and 10 h and at 24 h, the enrichments were tested for E. coli O157:H7 by direct fluorescent antibody (DFA), antibody-direct epifluorescent filter technique (Ab-DEFT), direct selective plating on sorbitol MacConkey agar (SMA), immunomagnetic separation coupled to either selective plating (IMS-SMA) or the polymerase chain reaction (IMS-PCR), and flow cytometry (FC). The most consistent detection of 0.1 CFU/ml of the slowest growing strain of the pathogen was provided by the IMS-SMA and IMS-PCR after 8 h of enrichment. The time required for detection at the level of 0.1 CFU/ml for each assay was Ab-DEFT, 11 h; IMS-PCR, 16 h; FC, 24 h; IMS-SMA, 32 h; and SMA, 48 h. Absolute detection limits (without enrichment) were: IMS-PCR, 10(3) CFU/ml; Ab-DEFT and IMS-SMA, 10(4) CFU/ml; SMA, 10(5) CFU/ml; and DFA, 10(6) CFU/ml. Recovery of the pathogen (10 CFU/ml) in apple juice after 28 days of 4 degrees C storage was possible by means of an 8-h enrichment and Ab-DEFT, IMS-PCR, or IMS-SMA.     

Effect of frequency and amplitude of vibration on void formation in dies poured from polyvinyl siloxane impressions

To determine the susceptibility of Escherichia coli O157:H7 to freezing and thawing in apple cider, methods that recover injured cells are needed for accurate enumeration. This study compared the ISO-GRID hydrophobic grid membrane filter (HGMF) SD-39 agar method to two other methods: a reference most probable number (MPN) method, and plating on sorbitol MacConkey agar (SMA). To determine numbers of injured cells, SMA spread plating was also compared to Trypticase soy agar (TSA) spread plating. Two strains of E. coli O157:H7 QA 326 and ATCC 43895, were inoculated into presterilized apple cider (10 ml) which was then frozen (-20 degrees C for 24 h). Samples were thawed at 4 degrees C for 4 h, or at 23 degrees C for 1.5 h, or in a microwave oven (700 W for 10 s). Substantial cell death (0.69- to 6.33 log10 CFU/ml decreases) and injury (0.70- to 2.38-log10 CFU/ml decreases) occurred during freezing and thawing. The extent of death and injury varied with strain and thawing method. The TSA spread plating method recovered the most cells while the HGMF method always recovered more viable cells than the reference MPN method and also either recovered significantly more (P < 0.05) cells or a not significantly different number of cells than SMA spread plating. Some injured cells of both strains were not counted by the HGMF method. Significant numbers of cells injured by freezing and thawing at 4 degrees C in apple cider were enumerated in the cider was diluted 1:2 Trypticase soy broth immediately before plating. Two epifluorescent microscopic methods showed that injury was not associated with loss of cell membrane integrity.     

Evaluation of dietary influences on Escherichia coli O157:H7 shedding by sheep

The effect of diet, an abrupt diet change, and fasting on the shedding of Escherichia coli O157:H7 was investigated with experimentally inoculated sheep as a ruminant model. Sheep were fed a grass hay diet (G), which was low in protein and digestible energy and high in fiber, or a mixture of corn and pelleted alfalfa (C), which was high in protein and digestible energy and low in fiber. After a single oral inoculation of E. coli O157:H7, all the animals shed fecal E. coli O157:H7. However, sheep that were fed G shed the bacterium almost twice as long as, and in larger numbers than, did sheep that were fed C. The number of culture-positive animals increased after the diet was abruptly changed from C to G and decreased with the opposite change (G to C). A 24-h fast did not influence E. coli O157:H7 shedding. Horizontal transmission of infection between animals occurred. Recent shedding of E. coli O157:H7 did not affect recolonization with E. coli O157:H7. The findings presented in this study indicate that preharvest control of diet may reduce the risk of E. coli O157:H7-positive animals entering the food chain.     

Symptom relief with amitriptyline in the irritable bowel syndrome

An assay was developed for the specific detection of Escherichia coli O157 using PCR, because O serological cross-reactivities have been reported between E. coli O157 and some E. coli, other bacterial species. PCR amplification of E. coli O157 rfbE (Ec O157:H7) gene that is necessary for the expression of the O157 antigen, was performed for the identification of E. coli O157. All Shiga toxin-producing Escherichia coli (STEC) O157:H7 and O157:H, non-STEC O157 strains were positive, and other non-O157 E. coli strains were negative by PCR. All tested strains of other bacterial species, like Salmonella O30 and Citrobacter freundii which gave positive results with O157 detection kits, were negative by PCR. It is recommended that PCR amplification of O157 rfbE gene is one of the most specific method for E. coli O157 identification.     

Effects of starvation on physiological activity and chlorine disinfection resistance in Escherichia coli O157:H7

Escherichia coli O157:H7 can persist for days to weeks in microcosms simulating natural conditions. In this study, we used a suite of fluorescent, in situ stains and probes to assess the influence of starvation on physiological activity based on membrane potential (rhodamine 123 assay), membrane integrity (LIVE/DEAD BacLight kit), respiratory activity (5-cyano-2,3-di-4-tolyl-tetrazolium chloride assay), intracellular esterase activity (ScanRDI assay), and 16S rRNA content. Growth-dependent assays were also used to assess substrate responsiveness (direct viable count [DVC] assay), ATP activity (MicroStar assay), and culturability (R2A agar assay). In addition, resistance to chlorine disinfection was assessed. After 14 days of starvation, the DVC values decreased, while the values in all other assays remained relatively constant and equivalent to each other. Chlorine resistance progressively increased through the starvation period. After 29 days of starvation, there was no significant difference in chlorine resistance between control cultures that had not been exposed to the disinfectant and cultures that had been exposed. This study demonstrates that E. coli O157:H7 adapts to starvation conditions by developing a chlorine resistance phenotype.     

Expression of vascular endothelial growth factor gene and its receptor (flt-1) gene in urinary bladder cancer

Factors affecting the ability of Escherichia coli O157:H7 to survive in foods with a(w) less than required for growth have not been fully defined. This study was undertaken to determine the ability of E. coli O157:H7 to survive in a commercial dry infant rice cereal as affected by a(w) (0.35+/-0.04, 0.52+/-0.03 and 0.73+/-0.03), pH (4.0 and 6.8), and temperature (5, 25, 35 and 45 degrees C), and in nine other reduced-a(w) foods. Death of E. coli O157:H7 in cereal was enhanced with increased temperature and decreased pH during a 16- to 24-week storage period. Survival was enhanced at pH 6.8 compared to pH 4.0 in cereal at a(w) 0.34+/-0.04 during initial storage at 5 and 25 degrees C. The effect of temperature (8, 15, 21 and 30 degrees C) on survival and growth of acid-adapted cells of E. coli O157:H7 inoculated into cereal reconstituted with milk or apple juice at two inoculum levels (8.2-12.3 cfu/ml and 82-123 cfu/ml of slurry) was also studied. Growth occurred in cereal reconstituted with milk at all test temperatures and in cereal reconstituted with apple juice at 15, 21 and 30 degrees C. Populations increased by >1 log10 cfu/ml within 3-6 h at 21 and 30 degrees C. Acid-adapted and unadapted cells had similar growth patterns. The effects of temperature and acid adaptation on survival of E. coli O157:H7 in nine commercial foods and food ingredients with pH 4.07-6.49 and a(w) 0.17-0.82 were determined. The pathogen survived in these foods for various lengths of time, depending the storage temperature, with an order of survival of 5 degrees C >21 degrees C >37 degrees C. Survival appeared to be enhanced in foods with highest pH, and acid-adapted cells retained higher viability than unadapted cells in only two of the nine test foods. Of particular importance is the ability of E. coli O157:H7 to survive well in dry foods with a wide range in a(w) and pH, particularly at refrigeration temperature.     

Polysaccharide side chains are not required for attaching and effacing adhesion of Escherichia coli O157:H7

Escherichia coli of the serotype O157:H7 is an enterohemorrhagic human pathogen which demonstrates attaching and effacing adhesion to colonocytes in vivo and to epithelial cells grown in tissue culture. Transposon TnphoA mutants of E. coli O157:H7 strain CL-8 were produced. Two of 300 alkaline phosphatase positive mutants, designated JB6 and JB27, did not express O157 side chains as assessed by agglutination with specific polyclonal O157 antiserum, silver staining of lipopolysaccharide extracts separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and Western immunoblots with polyclonal O157-specific antiserum. Both O157-negative mutants and the parent strain demonstrated localized adherence to HEp-2 cells when examined by Giemsa staining and bright-field microscopy. Furthermore, both O157-negative mutants showed enhanced adherence to HEp-2 cells compared with the parent strain when assessed by quantification of adherent bacterial CFUs. The parent strain, CL-8, and both of the mutants produced fluorescent foci when adherent bacteria and HEp-2 cells were stained with fluorescein isothiocyanate-labelled phalloidin. Transmission electron microscopy confirmed attaching and effacing adherence of strain CL-8 and the OO7-negative mutants to HEp-2 cells. These findings indicate that mutants deficient in O157 polysaccharide repeats exhibit adherence to tissue culture cells in vitro and that O157 polysaccharide repeats are not required to produce the attaching and effacing lesion.     

Survival and recovery of Escherichia coli O157:H7 in inoculated bottled water

A methodology used to isolate Escherichia coli O157:H7 from water and survival of this pathogen in inoculated water is described. The methodology used in the isolation of E. coli O157:H7 included the use of selective plating on Sorbitol MacConkey agar (supplemented with potassium tellurite [2.5 mg/liter], cefixime [0.05 mg/liter], and cefsulodin [10 mg/liter], and modified hemorrhagic colitis agar (also supplemented with potassium tellurite [2.5 mg/liter]) and cefsulodin [10 mg/liter]). There were no significant differences (P < 0.05) between the recoveries of E. coli O157:H7 on these two selective media. Direct plating on these selective agars was used to determine the length of time that E. coli O157:H7 was able to grow, remain viable, and be resistant to the selective agents. E. coli O157:H7 survived in inoculated water for up to > 300 days, depending on the type of water. Observation by scanning electron microscopy indicated that E. coli O157:H7 cells attached to, and multiplied on, the container walls.     

Enterohaemorrhagic Escherichia coli O157:H7 infection

Escherichia coli O157:H7 differs from previously described diarrheagenic E. coli classes (enteropathogenic, enteroinvasive, enterotoxigenic) by distinct clinical symptoms, production of verotoxin (VT) and a specific plasmid. Cattle are the primary reservoirs of E. coli O157:H7. The organism may be transmitted through the consumption of contaminated foods (mainly of bovine origin) and by person-to-person contact. The most typical clinical manifestations of E. coli O157:H7 infection are hemorrhagic colitis and hemolytic-uremic syndrome. Since the 1982 many outbreaks of E. coli O157:H7 infections as well as sporadic cases have been documented. Diagnosis of E. coli O157:H7 is based on a positive stool culture, presence of VT and elevated serum antibodies. The best currently available and inexpensive method for diagnosing E. coli O157:H7 is culture of stool on sorbitol-Mac Conkey agar medium.     

Over-expression of acetylcholinesterase stimulates the expression of agrin in NG108-15 cells

A 16-month old female child living on an Ontario dairy farm was taken to hospital suffering from bloody diarrhoea. Escherichia coli O157:H7 was isolated from her stool. Initial tests of well water samples were negative for E. coli by standard methods but culture of selected coliform colonies on sorbitol-MacConkey agar led to isolation of E. coli O157:H7. E. coli O157:H7 was also isolated from 63% of cattle on the farm. The E. coli O157:H7 isolates from the child, the water and the cattle were phage type 14, produced verotoxins 1 and 2, and were highly related on analysis by pulsed field gel electrophoresis. The child did not have known direct contact with the cattle and did not consume unpasteurized milk. Hydrogeological investigation revealed the design and location of the well would allow manure-contaminated surface water to flow into the well. This investigation demonstrates that cattle farm well water is a potential source of E. coli O157:H7 which may not be identified by standard screening for E. coli in water.