Biomedical subjects

C L Gyles

Publications and source records attributed to C L Gyles.

At least 19 recordsLinked to original sources

PubMed2006

Shiga toxin-producing Escherichia coli: an overview.

The objective of this review is to highlight the importance of cattle in human disease due to Shiga toxin-producing Escherichia coli (STEC) and to discuss features of STEC that are important in human disease. Healthy dairy and beef cattle are a major reservoir of a diverse group of STEC that infects humans through contamination of food and water, as well as through direct contact. Infection of humans by STEC may result in combinations of watery diarrhea, bloody diarrhea, and hemolytic uremic syndrome. Systems of serotyping, subtyping, and virulence typing of STEC are used to aid in epidemiology, diagnosis, and pathogenesis studies. Severe disease and outbreaks of disease are most commonly due to serotype O157:H7, which, like most other highly pathogenic STEC, colonize the large intestine by means of a characteristic attaching and effacing lesion. This lesion is induced by a bacterial type III secretion system that injects effector proteins into the intestinal epithelial cell, resulting in profound changes in the architecture and metabolism of the host cell and intimate adherence of the bacteria. Severe disease in the form of bloody diarrhea and the hemolytic uremic syndrome is attributable to Shiga toxin (Stx), which exists as 2 major types, Stx1 and Stx2. The stx genes are encoded on temperate bacteriophages in the chromosome of the bacteria, and production and release of the toxin are highly dependent on induction of the phages. Regulation of the genes involved in induction of the attaching and effacing lesion, and production of Stx is complex. In addition to these genes that are clearly implicated in virulence, there are several putative virulence factors. A major public health goal is to prevent STEC-induced disease in humans. Studies aimed at understanding factors that affect carriage and shedding of STEC by cattle and factors that contribute to development of disease in humans are considered to be important in achieving this objective.

Animals↗

PubMed2005

Acquisition of resistance to extended-spectrum cephalosporins by Salmonella enterica subsp. enterica serovar Newport and Escherichia coli in the turkey poult intestinal tract.

Salmonella enterica subsp. enterica serovar Newport resistant to the extended-spectrum cephalosporins (ESCs) and other antimicrobials causes septicemic salmonellosis in humans and animals and is increasingly isolated from humans, animals, foods, and environmental sources. Mechanisms whereby serovar Newport bacteria become resistant to ESCs and other classes of antimicrobials while inhabiting the intestinal tract are not well understood. The present study shows that 25.3% of serovar Newport strains isolated from the turkey poult intestinal tract after the animals were dosed with Escherichia coli harboring a large conjugative plasmid encoding the CMY-2 beta-lactamase and other drug resistance determinants acquired the plasmid and its associated drug resistance genes. The conjugative plasmid containing the cmy-2 gene was transferred not only from the donor E. coli to Salmonella serovar Newport but also to another E. coli serotype present in the intestinal tract. Laboratory studies showed that the plasmid could be readily transferred between serovar Newport and E. coli intestinal isolates. Administration of a single dose of ceftiofur, used to prevent septicemic colibacillosis, to 1-day-old turkeys did not result in the isolation of ceftiofur-resistant E. coli or Salmonella serovar Newport. There was a remarkable association between serotype, drug resistance, and plasmid profile among the E. coli strains isolated from the poults. This study shows that Salmonella serovar Newport can become resistant to ESCs and other antibiotics by acquiring a conjugative drug resistance plasmid from E. coli in the intestines.

Animals↗

PubMed2004

Resistance of broiler chickens to Escherichia coli respiratory tract infection induced by passively transferred egg-yolk antibodies.

Egg-yolk antibodies induced by immunizing hens with selected Escherichia coli antigens were evaluated for their ability to protect broiler chickens against respiratory/septicemic disease caused by avian pathogenic E. coli (APEC). Seven groups of broiler breeder hens were vaccinated three times, 1 week apart with live E. coli, killed E. coli, E. coli antigens [lipopolysaccharide (LPS), type 1 pilus adhesin (FimH), P pilus adhesin (PapG), aerobactin outer membrane receptor (IutA)] or phosphate buffered saline (PBS). An O78 APEC strain was used for preparation of all the antigens. Egg yolk immunoglobulins (IgY) were purified from eggs of each group and antibody activity in serum and purified IgY was determined by enzyme-linked immunosorbent assay (ELISA). IgY (100mg) was injected intramuscularly into 11-day-old broiler chickens, which were challenged 3 days later with homologous (O78) or heterologous (O1 or O2) E. coli by the intra-air sac route. Mortality was recorded and surviving chickens were euthanized 1 week after the challenge and examined for macroscopic lesions. Passive antibodies against all antigens except FimH were protective (90-100%) against the homologous challenge, but only anti-PapG and anti-IutA were effective against heterologous challenge. Anti-PapG IgY provided the greatest protection against the three serogroups of E. coli used for challenge. Hence vaccination of broiler breeders to induce anti-PapG and anti-IutA antibodies may provide passive protection of progeny chicks against respiratory/septicemic disease caused by APEC.

Adhesins, Escherichia coli↗

PubMed2003

A novel pathogenicity island integrated adjacent to the thrW tRNA gene of avian pathogenic Escherichia coli encodes a vacuolating autotransporter toxin.

We report the complete nucleotide sequence and genetic organization of the Vat-encoding pathogenicity island (PAI) of avian pathogenic Escherichia coli strain Ec222. The 22,139-bp PAI is situated adjacent to the 3' terminus of the thrW tRNA gene, has a G+C content of 41.2%, and includes a bacteriophage SfII integrase gene, mobile genetic elements, two open reading frames with products exhibiting sequence similarity to known proteins, and several other open reading frames of unknown function. The PAI encodes an autotransporter protein, Vat (vacuolating autotransporter toxin), which induces the formation of intracellular vacuoles resulting in cytotoxic effects similar to those caused by the VacA toxin from Helicobacter pylori. The predicted 148.3-kDa protein product possesses the three domains that are typical of serine protease autotransporters of Enterobacteriaceae: an N-terminal signal sequence of 55 amino acids, a 111.8-kDa passenger domain containing a modified serine protease site (ATSGSG), and a C-terminal outer membrane translocator of 30.5 kDa. Vat has 75% protein homology with the hemagglutinin Tsh, an autotransporter of avian pathogenic E. coli. A vat deletion mutant of Ec222 showed no virulence in respiratory and cellulitis infection models of disease in broiler chickens. We conclude that the newly described PAI and Vat may be involved in the pathogenicity of avian septicemic E. coli strain Ec222 and other avian pathogenic E. coli strains.

Amino Acid Sequence↗

PubMed2002

Shiga toxin genes in avian Escherichia coli.

The purpose of this study was to determine the presence of stx genes in avian pathogenic Escherichia coli (APEC). We examined 97 APEC isolates: 34 from lesions of avian cellulitis, 31 from avian septicemia, 13 from swollen head syndrome (SHS) in chickens, and 19 from diseased turkeys. We also examined five isolates from the feces of healthy chickens. All 102 E. coli isolates were tested for the presence of stx genes by PCR amplification and by colony blots using probes specific for stx1 and stx2. Fifty-three percent (52) of the 97 APEC carried stx gene sequences: one isolate carried stx2 sequences, two carried both stx1 and stx2 sequences, and the remaining 49 isolates carried only stx1 sequences. Twenty-six isolates were positive by both hybridization and PCR amplification, 10 were positive by PCR only, and 16 were positive by hybridization only. All the stx-positive isolates were negative by PCR for the eae and E-hlyA genes. The five isolates from healthy chickens were all negative for stx. All 13 SHS isolates were positive for the stx1 gene and had low titres for cytotoxicity in the Vero cell assay (VCA). Other stx-positive isolates were negative in the VCA. The stx1 gene from one SHS E. coli isolate was cloned and sequenced and shown to be identical to that of the stx gene of Shigella dysenteriae. The observations indicate that stx1 gene sequences are widespread among APEC but that cytotoxicity on Vero cells is uncommon.

Animals↗

PubMed2002

Pathogenic Shiga toxin-producing Escherichia coli in the intestine of calves.

The purpose of this study was to compare the pathological effects of Shiga toxin-producing Escherichia coli (STEC) that vary in their association with bovine and human disease. Shiga toxin-producing E. coli of serotypes associated with both dysentery in calves and hemolytic uremic syndrome (HUS) in humans (O5:H-, O26:H11, O111:H-,O113:H21) were compared with O157:H7 STEC, which are associated with HUS in humans but not with disease in calves. The STEC were administered orally to 80 day-old chicks and into ligated loops in the ileum and colon of four 2- to 6-day-old calves. Examination of the ceca of the chickens 10 d postchallenge showed no adherence or tissue abnormality for any isolate. The calves were euthanized 8 to 10 h postinoculation, and sections of the intestinal loops were examined by light microscopy, transmission and scanning electron microscopy, and immunohistochemistry. All strains showed consistent focal adherence associated with mild lesions in the colon. Attaching and effacing lesions were observed with the eae-positive strains. Ileal lesions were similar to the colonic ones but were sometimes severe, with marked polymorphonuclear leukocyte proliferation in the lamina propria. It is concluded that chickens were unsuitable for studying interaction of STEC with the intestine and that there was no difference in the interaction of the ligated calf intestine with STEC of serotypes associated with disease in calves compared with O157:H7 STEC.

Animals↗

PubMed2000

Estimation of the under-reporting rate for the surveillance of Escherichia coli O157:H7 cases in Ontario, Canada.

Two models estimating the proportion of Escherichia coli O157:H7 cases not reported in the Ontario notifiable diseases surveillance system are described. The first model is a linear series of adjustments in which the total number of reported cases is corrected by successive underreporting coefficients. The structure of the second model is based on a relative difference in the proportion of E. coli O157:H7 cases which are hospitalized between the surveillance database and the underlying population. Based on this analysis, the rate of under-reporting of symptomatic cases of E. coli O157:H7 infection in Ontario ranges from 78 to 88% corresponding to a ratio of 1 reported case for approximately 4-8 symptomatic cases missed by the surveillance system. This study highlights the need to increase awareness among public health workers of the potential biases that may exist in the interpretation of routine surveillance data.

Escherichia coli Infections↗

PubMed2000

Use of a Shiga toxin (Stx)-enzyme-linked immunosorbent assay and immunoblot for detection and isolation of Stx-producing Escherichia coli from naturally contaminated beef.

The purpose of this study was to evaluate an enzyme-linked immunosorbent assay (ELISA) and an immunoblot procedure for detection and isolation of Shiga toxin-producing Escherichia coli (STEC) from beef, and to correlate the presence of STEC in beef with E. coli and total coliform counts. A total of 120 samples of boneless beef supplied to a meat processor in southern Ontario were tested for the presence of STEC, E. coli, and total coliforms. Following enrichment in modified tryptic soy broth, samples were screened for Shiga toxin (Stx) by a Stx-ELISA and a Vero cell assay (VCA). Samples that were positive in the Stx-ELISA were subjected to the Stx-immunoblot for STEC isolation. Overall, 33.3% of samples were positive in the VCA, and 34.2% were positive in the Stx-ELISA. There was almost complete agreement between the Stx-ELISA and the VCA results (kappa = 0.98). The sensitivity and specificity of the Stx-ELISA with respect to the VCA were 100% and 98.75%, respectively. STEC were isolated by the Stx-immunoblot from 87.8% of the samples that were positive in the Stx-ELISA. The STEC isolates belonged to 19 serotypes, with serotype O113:H21 accounting for 10 of 41 isolates. No STEC of serotype O157:H7 were isolated. There was a significant correlation between E. coli counts and total coliform counts (Spearman correlation coefficient = 0.68, P < 0.01). The E. coli count was positively correlated with detection of STEC by both the Stx-ELISA and the VCA (P < 0.01).

Animals↗

PubMed2000

Prevention of edema disease in pigs by passive immunization.

The effect of treatment with verotoxin 2e (VT2e) specific antiserum was evaluated in 3 Danish pig herds with edema disease (ED). The antiserum was prepared by immunizing horses with a VT2e toxoid. The study was performed as a randomized blind field trial with parallel treatment and control groups. There were approximately 50 piglets in each group in each of the 3 herds and 741 piglets were included in the study (244 from herd A, 249 from herd B, and 247 from herd C). Treatment groups received 2, 4, or 6 mL anti-VT2e serum intramuscularly the day before weaning. Control groups were treated with 6 mL normal horse serum or 6 mL RPMI 1640 medium as placebo. All pigs that died in the trial period (1 d before weaning to 44 d after weaning) were examined pathologically and microbiologically. Mortality due to ED, mortality due to other causes, and adverse effects due to treatment were recorded. As there was no mortality due to ED, herd B was excluded from statistical calculations on mortality. The content of horse antibodies specific to VT2e in serum from pigs was analyzed in an indirect ELISA. A higher dose of anti-VT2e serum was reflected in higher optical density values in the indirect ELISA. Transient adverse reactions, seen as vomiting, ataxia, and cyanosis, occurred shortly after the injection of horse serum in 1.5% of the pigs, and one pig died. There were no statistically significant differences in mortality due to other causes among the 3 treatment groups in herds A and C. Only pigs from which F18+, VT2e+, ST-, LT- hemolytic E. coli (0139 or O-rough) was isolated were diagnosed as dead due to ED. Deaths due to ED in the control groups were 8.1% and 12.0% in herds A and C, respectively, compared with 0% and 0.7% in the corresponding serum groups. The difference between treatment and control groups was statistically significant (P<0.0001). It was not possible to establish an effect of dose (2, 4, or 6 mL) of anti-VT2e serum, because only one pig died of ED in the treatment groups. It was concluded that passive immunization by intramuscular injection of a VT2e-specific antiserum can be used for protecting piglets against ED.

Animals↗

PubMed2000

Risk factors for acute diarrhoea among inhabitants of Kampala District, Uganda.

OBJECTIVE: To identify modifiable individual and household risk factors for diarrhoea among people of all ages in Kampala district, Uganda. DESIGN: A cross-sectional, analytical study. SETTING: Multi-stage sampling. Four purposively selected parishes, two each from low and high socio-economic residential areas in Kampala district. Two randomly selected zones per parish with 60 households randomly selected from each zone. STUDY GROUP: All members present in each household at time of study. Individual and household information collected by means of personal interview using a questionnaire. MAIN OUTCOME MEASURES: Odds of diarrhoea among individuals or households exposed to a study factor compared with the odds of diarrhoea among those not exposed to the factor. RESULTS: Drinking raw chicken eggs was significantly (P < 0.01) and strongly (odds ratio (OR) = 99) associated with diarrhoea among residents of Kampala district. The odds of diarrhoea in households that 'cooked just enough food per meal' was significantly less (OR = 0.42) than in those that did not. People who used municipal water supplies and those who boiled their drinking water were significantly less likely (OR = 0.27, OR = 0.33, respectively) than those who used other water sources and/or who did not boil drinking water to report an episode of diarrhoea in the 2 weeks preceding the survey. The odds of diarrhoea were 2.6 times greater for individuals who reported a pest problem than for those who did not, while keeping pets was found to be protective (OR = 0.43). The number of income earners was also significantly (P < 0.5) and negatively (OR = 0.59) associated with the occurrence of diarrhoea in a member of the household. CONCLUSIONS: The findings of this study underscore the importance of proper food handling, preparation and eating habits as well as safe water, sanitation practices and socio-economic factors in the epidemiology of diarrhoea in developing countries.

Chi-Square Distribution↗

PubMed1999

A comparison of extracted proteins of isolates of Dermatophilus congolensis by sodium dodecyl sulphate-polyacrylamide gel electrophoresis and Western blotting.

Antigenic diversity within a collection of 18 isolates of Dermatophilus congolensis from different Continents was examined by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and by Western blotting with sera from cattle with clinical dermatophilosis using whole cell extracts obtained by three methods and one extract of extracellular products of D. congolensis. One of the methods involving the release of a lysostaphin-solubilized protein (LSP) of whole cells of D. congolensis revealed a number of discrete and easily-identifiable bands in SDS-PAGE which were found suitable for characterizing protein patterns and was, therefore, subsequently used for a comparative analysis of the proteins of all the D. congolensis isolates. Six electropherotypes (ET) of D. congolensis were identified among the 18 isolates using the protein profiles based on the presence of four protein bands at Molecular weights (MW) 62, 28, 17.4 and 16.4 kDa. The ETs were found among isolates from different animal species and from different sources with ET1 consisting of three bovine and two equine isolates; ET2, two bovine and three ovine isolates; ET3, two bovine isolates; ET4, two bovine isolates; ET5, one bovine and one ovine isolates and ET6, two bovine isolates. Immunoblotting of the extracts of D. congolensis isolates with sera from cattle with clinical dermatophilosis infection demonstrated protein bands of MW ranging from 9 kDa to 188 kDa. Sera from chronic dermatophilosis infection demonstrated a 28 kDa protein which was immunodominant in the LSP extracts of all the 18 isolates of D. congolensis tested while sera from mild infections demonstrated mainly the 62 kDa protein in the same extracts. However, many protein bands were demonstrated in surface membrane (TSMP) and extracellular protein extracts with sera from only mildly infected animals. The protein patterns observed in all isolates of D. congolensis revealed global antigenic similarities and distinct differences among isolates which could not be associated with either geographic, climatic or host factors. Also sera from infected animals from endemic regions of dermatophilosis could not differentiate isolates of D. congolensis. This suggests the possibility that such sera must have come from animals that had been infected by a multitude of D. congolensis strains present in the herd environment and strains an animal could have come across during the 'ritual' annual cross-country migration of the cattle herds.

Actinomycetales↗

PubMed1999

Temporal and geographical distributions of reported cases of Escherichia coli O157:H7 infection in Ontario.

The distribution of 3001 cases of verocytotoxigenic Escherichia coli (VTEC) reported in the Province of Ontario, Canada, were examined to describe the magnitude of this condition geographically and to evaluate the spatial relationship between livestock density and human VTEC incidence using a Geographical Information System. Incidence of VTEC cases had a marked seasonal pattern with peaks in July. Areas with a relatively high incidence of VTEC cases were situated predominantly in areas of mixed agriculture. Spatial models indicated that cattle density had a positive and significant association with VTEC incidence of reported cases (P = 0.000). An elevated risk of VTEC infection in a rural population could be associated with living in areas with high cattle density. Results of this study suggested that the importance of contact with cattle and the consumption of contaminated well water or locally produced food products may have been previously underestimated as risk factors for this condition.

Agriculture↗

PubMed1999

Associations between virulence factors of Shiga toxin-producing Escherichia coli and disease in humans.

Associations between known or putative virulence factors of Shiga toxin-producing Escherichia coli and disease in humans were investigated. Univariate analysis and multivariate logistic regression analysis of a set of 237 isolates from 118 serotypes showed significant associations between the presence of genes for intimin (eae) and Shiga toxin 2 (stx2) and isolates from serotypes reported in humans. Similar associations were found with isolates from serotypes reported in hemorrhagic colitis and hemolytic-uremic syndrome. The enterohemorrhagic E. coli (EHEC) hemolysin gene was significantly associated with isolates from serotypes found in severe diseases in univariate analysis but not in multivariate logistic regression models. A strong association between the intimin and EHEC-hemolysin genes may explain the lack of statistical significance of EHEC hemolysin in these multivariate models, but a true lack of biological significance of the hemolysin in humans or in disease cannot be excluded. This result warrants further investigations of this topic. Multivariate analysis revealed an interaction between the eae and stx2 genes, thus supporting the hypothesis of the synergism between the adhesin intimin and Shiga toxin 2. A strong statistical association was observed between the stx2 gene and severity of disease for a set of 112 human isolates from eight major serotypes. A comparison of 77 isolates of bovine origin and 91 human isolates belonging to six major serotypes showed significant associations of the genes for Shiga toxin 1 and EspP protease with bovine isolates and an increased adherence on HEp-2 cell cultures for human isolates, particularly from diarrheic patients and healthy persons.

Adhesins, Bacterial↗

PubMed1999

Virulence markers in Shiga toxin-producing Escherichia coli isolated from cattle.

This study identified potential virulence markers in 93 eae-positive and 179 eae-negative Shiga toxin-producing Escherichia coli (STEC), isolated from a random sampling of healthy cattle in southwestern Ontario. PCR amplification was used to identify genes for enterohemorrhagic E. coli (EHEC)-hemolysin, the EAF plasmid, and bundle-forming pili (Bfp); adherence to HEp-2 cells and to bovine colonocytes, and the fluorescent actin staining (FAS) test were used to characterize interaction of the bacteria with epithelial cells. The EHEC-hemolysin sequences were detected in 98% of eae-positive isolates compared with 34% of eae-negative isolates. All isolates were negative for EAF and bfp sequences. There was 100% correlation between localized adherence (LA) to HEp-2 cells and the FAS test. Forty-eight (52%) of the eae-positive isolates were LA/FAS-positive, whereas none of the 179 eae-negative isolates was positive in either test. Among the eae-negative isolates, 20 (11%) showed diffuse adherence and 5 (2.8%) showed enteroaggregative adherence to HEp-2 cells. Seventy-three percent of the eae-positive isolates adhered to bovine colonocytes, whereas only 26% of 120 eae-negative isolates that were tested adhered. All 13 O157:H7 isolates were positive for eae and EHEC-hemolysin gene sequences, LA/FAS, and adherence to bovine colonocytes. It is concluded that possession of genes for eae and EHEC hemolysin is correlated with the serotype of STEC, that production of EHEC hemolysin was highly correlated with serotypes implicated in human disease, and that none of the potential markers that were examined can be used to predict the potential virulence of an isolate.

Acyltransferases↗

PubMed1998

Localization of potential binding sites for the edema disease verotoxin (VT2e) in pigs.

The purpose of this study was to identify organs and cells to which the edema disease verotoxin (VT2e) could bind in pigs. Frozen 4-5 microns thick sections of organs usually affected in edema disease (colon, spinal cord, cerebellum and eyelid) and organs not usually affected (liver, ileum) from two 5- to 6-week-old weaned pigs were permeabilized with acetone, then exposed to VT2e. Unbound VT2e was removed by washing and bound VT2e was detected by immunohistochemistry. In the eyelid, double-label immunofluorescence was used to identify the cells to which VT2e bound. VT2e was shown to bind to all six organs that were examined. The toxin bound to arteries in all organs, to veins in all organs except the liver, and to enterocytes in the ileal crypts. Double labelling of eyelid with monoclonal antibodies specific for von Willebrand factor or alpha-smooth actin and VT2e showed that the toxin bound to endothelial and vascular smooth muscle cells. The binding of VT2e to endothelium is consistent with findings for other verotoxins but binding to vascular smooth muscle has not been reported for other verotoxins. It is concluded that i) factors other than the presence of receptors for VT2e influence the development of lesions in edema disease, and ii) smooth muscle necrosis, which is characteristic of the vascular lesions in edema disease, may be due to a direct action of toxin on smooth muscle cells.

Animals↗

PubMed1997

A comparison of procedures involving single step Salmonella, 1-2 Test, and modified semisolid Rappaport-Vassiliadis medium for detection of Salmonella in ground beef.

Procedures involving Single Step Salmonella (SSS), 1-2 Test, and Modified Semisolid Rappaport-Vassiliadis Medium (MSRV) were compared for their speed and sensitivity in detection of Salmonella in ground beef contaminated with one isolate of each of five Salmonella serotypes. Inocula of 10, 10(2) and 10(3) CFU/g of ground beef were used. When pre-enrichment in buffered peptone water and selective enrichment in tetrathionate broth were used, SSS and MSRV detected all five species of Salmonella at all levels of contamination, whereas the 1-2 Test was positive in 0, 12, and 15 of 15 tests at 10, 10(2) and 10(3) CFU/g, respectively. When only pre-enrichment was used, the results with MSRV were unchanged but the SSS test failed to detect S. typhimurium. Thus, with pre-enrichment the MSRV and SSS procedures were equally sensitive and both produced a result on the third day. The 1-2 Test was less sensitive and slower, with results available on day-4. The MSRV protocol was best overall.

Animals↗

PubMed1997

Hemolysin phenotypes and genotypes of eaeA-positive and eaeA-negative bovine verotoxigenic Escherichia coli.

Intimin or EaeA protein has been implicated in the attaching/effacing lesion caused by entero-hemorrhagic Escherichia coli (EHEC) in the intestine but it is not produced by all EHEC and is therefore not adequate as a marker for EHEC. Hemolysins are produced by a high percentage of verotoxigenic E. coli (VTEC) and could be a marker for EHEC, but their distribution and relation to virulence are not known. We used PCR amplification to determine the presence or absence of eaeA sequences in 281 VTEC isolates from the feces of healthy cattle. There were 101 eaeA-positive isolates, which belonged to O groups 5, 26, 69, 80, 84, 98, 103, 111, 119, 145, 157 and 108 eaeA-negative isolates, which belonged to O groups 8, 22, 38, 113, 119, 116, 132, 153, 156 or untypable. All isolates were tested for hemolysis on horse blood agar and on washed sheep blood agar. PCR amplification was used to test for EHEC hemolysin, Ehly1, Ehly2 and alpha-hemolysin D sequences. Among eaeA-positive isolates 98% were positive for EHEC hemolysin sequences and were hemolytic on washed sheep red blood cell agar; the corresponding percentage for eaeA-negative isolates was 36%. Ehly1 and Ehly2 sequences were present in only 11 isolates (O groups 26, 84, 119 and 132). None of the eaeA-positive and 13 of the eaeA-negative isolates (including all 11 isolates of O group 132) were positive for alpha-hemolysin D gene sequences by PCR and alpha-hemolysin production on horse blood agar. We conclude that since Ehly1, Ehly2 and alpha-hemolysin occur at low frequency among bovine VTEC and serotypes implicated in human disease they are unlikely to be significant virulence factors. In contrast, EHEC hemolysin was present in almost all eaeA-positive VTEC isolates and in approximately one-third the eaeA-negative ones; it may be both a virulence marker and virulence factor but further testing is required. The study identified isolates with all combinations of eaeA and EHEC hly, which may be useful for further testing.

Adhesins, Bacterial↗

PubMed1997

Evaluation of an aroA mutant Salmonella typhimurium vaccine in chickens using modified semisolid Rappaport Vassiliadis medium to monitor faecal shedding.

In groups of chickens vaccinated orally or intramuscularly with a live aroA mutant Salmonella typhimurium vaccine strain and then experimentally inoculated with 10(8) CFU of wild type S. typhimurium or 10(9) CFU of S. enteritidis, faecal shedding of the vaccine and wild type strains was monitored by the buffered peptone water-modified semisolid Rappaport Vassiliadis medium method, which detected less than 10(2) CFU per gram of faeces. The vaccine strain was shed in the faeces for up to 26 days. Vaccination failed to reduce the faecal shedding of wild type S. typhimurium or S. enteritidis. The variation in the shedding patterns of chickens within each group was greater than between treatment groups.

3-Phosphoshikimate 1-Carboxyvinyltransferase↗