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Biomedical subjects

M P Doyle

Publications and source records attributed to M P Doyle.

At least 73 records · Page 4Linked to original sources
PubMed1997

Emerging issues in microbiological food safety.

Many microorganisms previously unrecognized as food-borne or harmful are emerging as human pathogens transmitted by food. This is a result of recent acquisition of key virulence factors, detection by newly developed isolation procedures, or astute detective-like laboratory skills of microbiologists. Six microbial pathogens, including Shiga toxin-producing Escherichia coli, Listeria monocytogenes, Arcobacter butzleri, Helicobacter pylori, Cryptosporidium parvum, and Cyclospora, have become recognized as significant causes of human illness. Although the ecology and epidemiology of illness caused by some of these pathogens have not been fully elucidated, food has the potential of being an important vehicle in their dissemination. Existing technologies and new approaches such as irradiation and hazard analysis critical control point (HACCP) programs are useful tools in the control of food-borne hazards. However, because of ever-changing products, processes, food-handling practices, societal habits, and pathogens, emerging food-borne diseases will continue to be an important public health concern.

Campylobacter Infections↗
PubMed1997

Acetylcholine induces conducted vasodilation by nitric oxide-dependent and -independent mechanisms.

Conducted vasodilation has been proposed as an important component of local vascular control. Because conducted vasomotor responses have previously been studied only in response to short pulses (<500 ms) of agonist, this study examined conducted vasodilation in response to sustained stimuli. In addition, we examined the contribution of nitric oxide (NO) to initiation and maintenance of conducted responses induced by acetylcholine (ACh). Responses to 2-min applications of ACh, sodium nitroprusside, and 8-bromoguanosine 3',5'-cyclic monophosphate were obtained in cannulated, perfused hamster cheek pouch arterioles (approximately 60 microm in diameter). Changes of luminal diameter in response to pressure ejection of agonists from a micropipette placed close to the downstream end of the vessel were observed at the site of stimulation ("local") as well as 570 and 1,140 microm upstream. At the local site, ACh stimuli produced large changes in diameter (approximately 70% of the maximum response) that peaked within 45 s before declining slowly to levels of approximately 50% of the maximum response. A similar response pattern was observed at both upstream sites, with the conducted responses being maintained for the duration of the stimulus. Local responses of similar magnitude were found with sodium nitroprusside and 8-bromoguanosine 3',5'-cyclic monophosphate, but only minimal responses were observed at the conducted sites. In a separate set of arterioles, ACh responses were obtained before and during perfusion with 10 microM N(omega)-nitro-L-arginine. Inhibition of NO synthesis diminished the local response to ACh, but the initial phase of the conducted response was unaffected. Furthermore, the conducted responses faded more rapidly in the presence of N(omega)-nitro-L-arginine. We conclude from these results that local NO synthesis alone is insufficient to initiate conducted responses but that NO synthesis contributes to maintenance of sustained conducted responses.

Acetylcholine↗
PubMed1997

Hamburger-associated Escherichia coli O157:H7 infection in Las Vegas: a hidden epidemic.

OBJECTIVES: This study sought to determine whether a multistate fast food hamburger-associated outbreak of Escherichia coli O157:H7 infection involved Las Vegas residents as well and, if so, why public health officials had not detected it. METHODS: A matched case-control study was conducted among persons with bloody diarrhea and their healthy meal companions. Hamburger production, distribution, and cooking methods were reviewed. Unused hamburger patties were cultured, and E. coli O157:H7 isolates were characterized. Local laboratory stool culture practices were reviewed. RESULTS: Fifty-eight cases of bloody diarrhea were identified. Illness was associated with eating regular hamburgers (matched odds ratio [OR] = 9.0, 95% confidence interval [CI] = 1.02,433.4), but 25% of ill persons reported eating only jumbo hamburgers. Regular and jumbo hamburger patties yielded E. coli O157:H7 indistinguishable from the lone clinical isolate. No local laboratory cultured routinely for E. coli O157:H7 until after the outbreak. CONCLUSIONS: A large outbreak of E. coli O157:H7 infections escaped timely notice in Las Vegas because local laboratories did not culture for this pathogen. Health officials should encourage laboratories to screen at least all bloody stools on sorbitol-MacConkey medium.

Adolescent↗
PubMed1996

Polymerase chain reaction for detecting Escherichia coli O157: H7.

Escherichia coli O157:H7 is known as an important cause of hemorrhagic colitis and hemolytic uremic syndrome. Real-time procedures that are sensitive for detecting small populations of this bacterium in food are lacking and needed. An expression library was constructed by ligation of BamHI-EcoRI DNA fragments of E. coli O157:H7 to plasmid vector pUC19 and transformation of recombinant plasmids to E. coli JM109. A clone that contained a specific DNA fragment of E. coli O157:H7 was identified by colony immunoblot assay using monoclonal antibody MAb 4E8C12 that uniquely links to E. coli O157:H7 and a few other serotypes of verotoxin-producing E. coli. The DNA sequence of the clone consisted of 110 bp of 5' region of enterohemorrhagic E. coli (EHEC) eae gene and a 688 bp DNA fragment adjacent to 5' end of the eae gene, including an unknown function gene encoding 156 amino acids. A pair of oligonucleotide primers was synthesized based on the sequence of the 688 bp fragment. The primers were used in a polymerase chain reaction (PCR) to amplify a target DNA of 633 bp. The primers amplified 1 ng of DNA from 67 strains of E. coli O157:H7, two strains of E. coli O157:NM, and 7 of 11 E. coli O55:H7 and O55:NM strains, but not 50 ng of DNA from 34 strains of 29 other E. coli serotypes and 25 strains of 8 other bacterial species. Annealing temperatures from 60 to 63 degrees C could be used for the PCR without loss of specificity. The minimum amount of target DNA detected by the PCR was 5 pg. When a boiling method and GeneReleaser were used, the PCR was able to detect as few as 25 and 38 CFU of E. coli O157:H7, respectively, in 3 h.

Antibodies, Monoclonal↗
PubMed1996

A low molecular weight outer-membrane protein of Escherichia coli O157:H7 associated with adherence to INT407 cells and chicken caeca.

Enterohaemorrhagic Escherichia coli (EHEC) O157:H7 and O26:H11 have been shown to produce a low mol.wt outer-membrane protein (OMP) that is unique to a few serotypes of E. coli. A mutant (A10) of E. coli O157:H7 strain HA1 deficient in the OMP was constructed by TnphoA mutagenesis and assayed for its adherent ability. Adherence of A10 to intestinal epithelial cells (INT407) was significantly less than that of its parent strain (HA1). Adherence of HA1 to INT407 cells was significantly decreased by treatment with a monoclonal antibody (4E8C12) that specifically binds to the OMP. When chickens were infected experimentally with E. coli O157:H7 strains, the average number of cfu of strain A10 recovered from chicken caeca was significantly less than those of strain HA1 and wild-type strain 932 at 14 and 21 days after peroral inoculation. These data suggest that the OMP of EHEC is associated with adherence of E. coli O157:H7 to epithelial cells in vitro and chicken caeca in vivo.

Animals↗
PubMed1996

Fate of enterohemorrhagic Escherichia coli O157:H7 in bovine feces.

Dairy cattle have been identified as a principal reservoir of Escherichia coli O157:H7. The fate of this pathogen in bovine feces at 5, 22, and 37 degrees C was determined. Two levels of inocula (10(3) and 10(5) CFU/g) of a mixture of five nalidixic acid-resistant E. coli O157:H7 strains were used. E. coli O157:H7 survived at 37 degrees C for 42 and 49 days with low and high inocula, respectively, and at 22 degrees C for 49 and 56 days with low and high inocula, respectively. Fecal samples at both temperatures had low moisture contents (about 10%) and water activities ( < 0.5) near the end of the study. E. coli O157:H7 at 5 degrees C survived for 63 to 70 days, with the moisture content (74%) of feces remaining high through the study. Chromosomal DNA fingerprinting of E. coli O157:H7 isolates surviving near the completion of the study revealed that the human isolate strain 932 was the only surviving strain at 22 or 37 degrees C. All five strains were isolated near the end of incubation from feces held at 5 degrees C. Isolates at each temperature were still capable of producing both verotoxin 1 and verotoxin 2. Results indicate that E. coli O157:H7 can survive in feces for a long period of time and retain its ability to produce verotoxins. Hence, bovine feces are a potential vehicle for transmitting E. coli O157:H7 to cattle, food, and the environment. Appropriate handling of bovine feces is important to control the spread of this pathogen.

Animals↗
PubMed1995

Cloning and nucleotide sequence of a gene upstream of the eaeA gene of enterohemorrhagic Escherichia coli O157:H7.

A DNA segment located immediately upstream of the eaeA gene of enterohemorrhagic Escherichia coli O157:H7 strain HA1 was cloned and sequenced. This segment contained an open reading frame encoding a predicted protein of 156 amino acids. A database search identified similar open reading frames upstream of the eaeA gene in two other bacterial pathogens, i.e. enteropathogenic E. coli and Citrobacter freundii. The predicted amino acid sequence of the enterohemorrhagic E. coli protein shared 96.8% and 94.2% identity with the enteropathogenic E. coli and C. freundii sequences, respectively. Because the open reading frame is located within the locus of enterocyte effacement region of the E. coli chromosome, a 'hot spot' for insertion of virulence factor genes, and shares high sequence homology with attaching and effacing EPEC and C. freundii, this protein may be associated with pathogenicity of E. coli O157:H7.

Amino Acid Sequence↗
PubMed1995

Risk factors for fecal shedding of Escherichia coli O157:H7 in dairy calves.

A case-control study was conducted to determine risk factors for fecal shedding of Escherichia coli O157:H7 (ECO) in dairy calves. Three herds previously found to lack calves that shed ECO in their feces were selected for each herd previously found to have calves that shed ECO. Fecal samples from 965 calves on 64 farms were tested for ECO by microbial culture. Sample prevalence of ECO in calves less than 8 weeks old was 1.4% and in calves 8 weeks or older was 4.8%. Calves were 3 times more likely to shed ECO after weaning than before weaning. Shedding of ECO was associated with grouping calves before weaning; feeding whole cottonseed was negatively associated with ECO shedding. The change in results between testing periods illustrated that an individual herd's status cannot be defined by a single testing of a small sample of cattle.

Age Factors↗
PubMed1995

Molecular characterisation of Escherichia coli O157:H7 isolates by pulsed-field gel electrophoresis and plasmid DNA analysis.

Foods of bovine origin have been identified as sources of Escherichia coli O157:H7. Genomic DNA of E. coli O157:H7 isolates from patients (six isolates), food samples (18 isolates from ground beef and six isolates from raw milk) and calf faecal samples (31 isolates) were characterised by pulsed-field gel electrophoresis (PFGE) and plasmid DNA analysis. These isolates originated from different locations in the USA during 1992 and 1993. Twenty-one distinct genomic profiles were generated from the 61 isolates by PFGE after digestion with the endonuclease XbaI. Four genomic profiles were identified among five patient isolates and the remaining patient isolate was not typable. Five different profiles were detected amongst the isolates from ground beef, one of which was associated with 13 ground beef isolates from an outbreak in the Pacific Northwest of the USA in 1993. The PFGE profile of five calf isolates from Washington and Wisconsin was identical to the profile of the ground beef isolates from the outbreak, suggesting that these isolates were related. Similarly, one PFGE profile accounted for three isolates from calf faeces and one from ground beef. Six raw milk isolates from Georgia were indistinguishable both from each other and from one isolate from calf faeces. Fourteen genomic profiles were identified among 31 calf faecal isolates from 18 different herds in 11 states. Only five plasmid profiles were identified among the 61 isolates. PFGE was shown to be a useful typing technique for E. coli O157:H7.

Animals↗
PubMed1995

Death of enterohemorrhagic Escherichia coli O157:H7 in real mayonnaise and reduced-calorie mayonnaise dressing as influenced by initial population and storage temperature.

This study was undertaken to determine the survivability of low-density populations (10(0) and 10(2) CFU/g) of enterohemorrhagic Escherichia coli O157:H7 inoculated into real mayonnaise and reduced-calorie mayonnaise dressing and stored at 20 and 30 degrees C, temperatures within the range used for normal commercial mayonnaise distribution and storage. Inactivation patterns at 5 degrees C and inactivation of high-inoculum populations (10(6) CFU/g) were also determined. The pathogen did not grow in either mayonnaise formulation, regardless of the inoculum level or storage temperature. Increases in storage temperature from 5 to 20 degrees C and from 20 to 30 degrees C resulted in dramatic increases in the rate of inactivation. Populations of E. coli O157:H7 in the reduced-calorie and real formulations inoculated with a population of 0.23 to 0.29 log10 CFU/g and held at 30 degrees C were reduced to undetectable levels within 1 and 2 days, respectively; viable cells were not detected after 1 day at 20 degrees C. In mayonnaise containing an initial population of 2.23 log10 CFU/g, viable cells were not detected after 4 days at 30 degrees C or 7 days at 20 degrees C; tolerance was greater in real mayonnaise than in reduced-calorie mayonnaise dressing stored at 5 degrees C. The tolerance of E. coli O157:H7 inoculated at the highest population density (6.23 log 10 CFU/g) was less in reduced-calorie mayonnaise dressing than in real mayonnaise at all storage temperatures. In reduced-calorie mayonnaise dressing and real mayonnaise initially containing 2.23 log10 CFU/g, levels were undetectable after 28 and 58 days at 5 degrees C, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Colitis↗
PubMed1995

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.

Animal Husbandry↗
PubMed1994

Inactivation of Escherichia coli O157:H7, salmonellae, and Campylobacter jejuni in raw ground beef by gamma irradiation.

Raw ground beef patties inoculated with stationary-phase cells of Escherichia coli O157:H7, salmonellae, or Campylobacter jejuni were subjected to gamma irradiation (60Co) treatment, with doses ranging from 0 to 2.52 kGy. The influence of two levels of fat (8 to 14% [low fat] and 27 to 28% [high fat]) and temperature (frozen [-17 to -15 degrees C] and refrigerated [3 to 5 degrees C]) on the inactivation of each pathogen by irradiation was investigated. In ascending order of irradiation resistance, the D10 values ranged from 0.175 to 0.235 kGy (C. jejuni), from 0.241 to 0.307 kGy (E. coli O157:H7), and from 0.618 to 0.800 kGy (salmonellae). Statistical analysis revealed that E. coli O157:H7 had a significantly (P < 0.05) higher D10 value when irradiated at -17 to -15 degrees C than when irradiated at 3 to 5 degrees C. Regardless of the temperature during irradiation, the level of fat did not have a significant effect on the D10 value. Salmonellae behaved like E. coli O157:H7 in low-fat beef, but temperature did not have a significant effect when the pathogen was irradiated in high-fat ground beef. Significantly higher D10 values were calculated for C. jejuni irradiated in frozen than in refrigerated low-fat beef. C. jejuni was more resistant to irradiation in low-fat beef than in high-fat beef when treatment was at -17 to -15 degrees C. Regardless of the fat level and temperature during inactivation, these pathogens were highly sensitive to gamma irradiation.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Variable colonization of chickens perorally inoculated with Escherichia coli O157:H7 and subsequent contamination of eggs.

Challenging 1-day-old White Leghorn chicks perorally with 2.6 x 10(1) to 2.6 x 10(5) Escherichia coli O157:H7 bacteria per chick resulted in cecal colonization at all levels. Two of six chicks inoculated with only 2.6 x 10(1) E. coli O157:H7 bacteria carried 10(3) to 10(4) E. coli O157:H7 bacteria per g of cecal tissue when sacrificed 3 months postinoculation. E. coli O157:H7 colonization persisted at least 10 to 11 months when chicks were administered 10(8) E. coli O157:H7 bacteria. Eggs from five hens that were fecal shedders of E. coli O157:H7 until the termination of the study (10 to 11 months) were assayed for E. coli O157:H7. The organism was isolated from the shells of 14 of 101 (13.9%) eggs but not from the yolks and whites. Considering that chicks can be readily colonized by small populations of E. coli O157:H7 and continue to be long-term shedders, it is possible that chickens and hen eggs can serve as vehicles of this human pathogen.

Animals↗
PubMed1994

Nucleoside-induced arteriolar constriction: a mast cell-dependent response.

Adenosine (Ado) is a potent vasodilator that has occasionally been shown to cause vasoconstriction. Constrictor responses are generally attributed to A1-receptor stimulation or interactions with the renin-angiotensin system. We describe a previously unreported vasoconstrictor action of Ado and inosine (Ino) in hamster cheek pouch arterioles and examine the mechanism by which these nucleosides induce constriction. Arterioles were dissected from male Golden hamster cheek pouches, transferred to a 37 degrees C tissue chamber, and cannulated at both ends. Changes of luminal diameter in response to Ado were measured to generate cumulative concentration-response curves. The concentration-response curves were biphasic: 10(-6) M Ado elicited an intense, transient constriction, and higher concentrations induced dilator responses. Pretreatment with 8(p-sulfophenyl)theophylline, an Ado receptor antagonist, inhibited the dilator responses but did not alter the constriction. Inhibition of Ado uptake with S-(4-nitrobenzyl)-6-thio-inosine eliminated the constrictor response without altering dilator responses. Similar effects were found after pretreatment with an Ado deaminase inhibitor erythro-9-(2-hydroxy-3-nonyl)adenine hydrochloride. Finally, Ino, a metabolite of Ado, induced constrictions of similar magnitude to those seen with Ado, but at higher concentrations. The constrictor response was focal in nature, suggesting discrete sites of action of Ado. Methylene blue staining after Ado application revealed degranulated mast cells closely associated with the vessel wall, indicating a possible role for mast cell degranulation in the constrictor response. Supporting this idea were the observations that inhibition of degranulation by 10 microM cromolyn blocked the constrictor response, and compound 48/80 (a mast cell secretagogue) caused constriction similar to that elicited by Ado.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenine↗
PubMed1994

Cardiovascular responses to hemorrhage during acute and chronic hypoxia.

Previous work from our laboratory had demonstrated attenuation of systemic vasoreactivity to pressor agents in rats after acute or chronic exposure to hypoxia. Therefore we hypothesized that hemorrhage of acutely hypoxic (12% O2) or chronically hypoxic (barometric pressure 380 mmHg for 3 wk) rats would deter the normal increase in total peripheral resistance (TPR) and thus decrease the ability to maintain blood pressure. Progressive hemorrhage (2% of blood volume per min) was performed under conditions of either normoxia or acute hypoxia in conscious rats. In control animals, the increase in TPR observed during normoxic hemorrhage was absent when hemorrhage was performed in acute hypoxia. Furthermore, the amount of blood removal required to achieve hypotension was reduced under conditions of acute hypoxia. In contrast, chronically hypoxic rats exhibited no difference in the threshold for hypotension between conditions of acute normoxia and hypoxia and demonstrated an increased hypotensive threshold under both normoxic and hypoxic conditions compared with control animals. We next hypothesized that the prolonged threshold for hypotension observed in chronically hypoxic rats might be due to hypoxia-induced right ventricular hypertrophy. Such ventricular hypertrophy may minimize stimulation of ventricular volume receptors thought to elicit the reflex fall in heart rate and TPR occurring in extreme underfill conditions. Therefore we compared the cardiovascular responses to hemorrhage in rats with right ventricular hypertrophy resulting from administration of monocrotaline with those from rats treated with vehicle.(ABSTRACT TRUNCATED AT 250 WORDS)

Acute Disease↗
PubMed1993

An outbreak of diarrhea and hemolytic uremic syndrome from Escherichia coli O157:H7 in fresh-pressed apple cider.

OBJECTIVE: Escherichia coli O157:H7 causes hemorrhagic colitis and the hemolytic uremic syndrome. In the fall of 1991, an outbreak of E coli O157:H7 infections in southeastern Massachusetts provided an opportunity to identify transmission by a seemingly unlikely vehicle. DESIGN: Case-control study to determine the vehicle of infection. New England cider producers were surveyed to assess production practices and determined the survival time of E coli O157:H7 organisms in apple cider. RESULTS: Illness was significantly associated with drinking one brand of apple cider. Thirteen (72%) of 18 patients but only 16 (33%) of 49 controls reported drinking apple cider in the week before illness began (odds ratio [OR], 8.3; 95% confidence interval [CI], 1.8 to 39.7). Among those who drank cider, 12 (92%) of 13 patients compared with two (13%) of 16 controls drank cider from cider mill A (lower 95% CI, 2.9; P < .01). This mill pressed cider in a manner similar to that used by other small cider producers: apples were not washed, cider was not pasteurized, and no preservatives were added. In the laboratory, E coli O157:H7 organisms survived for 20 days in unpreserved refrigerated apple cider. Addition of sodium benzoate 0.1% reduced survival to less than 7 days. CONCLUSIONS: Fresh-pressed, unpreserved apple cider can transmit E coli O157:H7 organisms, which cause severe infections. Risk of transmission can be reduced by washing and brushing apples before pressing, and preserving cider with sodium benzoate. Consumers can reduce their risk by only drinking cider made from apples that have been washed and brushed.

Adolescent↗