Distribution ofEscherichia colistrains harbouring Shiga toxin-producingE. coli(STEC)-associated virulence factors (stx1, stx2, eae, ehxA) from very young calves in the North Island of New Zealand

2014 ◽  
Vol 142 (12) ◽  
pp. 2548-2558 ◽  
Author(s):  
H. IRSHAD ◽  
A. L. COOKSON ◽  
D. J. PRATTLEY ◽  
M. DUFOUR ◽  
N. P. FRENCH
Keyword(s):  
New Zealand ◽  
Shiga Toxin ◽  
Macconkey Agar ◽  
Chain Reaction ◽  
Potassium Tellurite ◽  
E Coli ◽  
The North ◽  
Virulence Markers ◽  
Polymerase Chain ◽  
Common Combination

SUMMARYThe objective of this study was to determine the distribution of Shiga toxin-producingEscherichia coli(STEC) virulence markers (stx1,stx2,eae,ehxA) inE. colistrains isolated from young calves aged fewer than 7 days (bobby calves). In total, 299 recto-anal mucosal swabs were collected from animals at two slaughter plants and inoculated onto tryptone bile X-glucuronide and sorbitol MacConkey agar supplemented with cefixime and potassium tellurite. Isolates were analysed using multiplex polymerase chain reaction to detectstx1,stx2,eaeandehxAgenes. The most common combination of virulence markers wereeae, ehxA(n = 35) followed byeae(n = 9). In total, STEC and atypical enteropathogenicE. coli(aEPEC) were isolated from 8/299 (2·6%) and 37/299 (12·3%) calves, respectively. All the isolates could be assigned to 15 genotype clusters with >70% similarity cut-off usingXbaI pulsed-field gel electrophoresis. It may be concluded that healthy calves from the dairy industry are asymptomatic carriers of a diverse population of STEC and aEPEC in New Zealand.

scholarly journals Prevalence and molecular detection of shiga toxin producing Escherichia coli from diarrheic cattle

2016 ◽  
Vol 14 (1) ◽  
pp. 63-68 ◽  
Author(s):  
MM Akter ◽  
S Majumder ◽  
KH MNH Nazir ◽  
M Rahman
Keyword(s):  
Escherichia Coli ◽  
Shiga Toxin ◽  
Molecular Detection ◽  
Chain Reaction ◽  
Specific Primers ◽  
Fecal Samples ◽  
E Coli ◽  
Uremic Syndrome ◽  
Polymerase Chain ◽  
Positive Isolate

Shiga toxin-producing Escherichia coli (STEC) are zoonotically important pathogen which causes hemorrhagic colitis, diarrhea, and hemolytic uremic syndrome in animals and humans. The present study was designed to isolate and identify the STEC from fecal samples of diarrheic cattle. A total of 35 diarrheic fecal samples were collected from Bangladesh Agricultural University (BAU) Veterinary Teaching Hospital. The samples were primarily examined for the detection of E. coli by cultural, morphological and biochemical characteristics, followed by confirmation of the isolates by Polymerase Chain Reaction (PCR) using gene specific primers. Later, the STEC were identified among the isolated E. coli through detection of Stx-1 and Stx-2 genes using duplex PCR. Out of 35 samples, 25 (71.43%) isolates were confirmed to be associated with E. coli, of which only 7 (28%) isolates were shiga toxin producers, and all of them were positive for Stx-1. However, no Stx-2 positive isolate could be detected. From this study, it may be concluded that cattle can act as a reservoir of STEC which may transmit to human or other animals.J. Bangladesh Agril. Univ. 14(1): 63-68, June 2016

Detection and Quantitation of Enterohemorrhagic Escherichia coli O157, O111, and O26 in Beef and Bovine Feces by Real-Time Polymerase Chain Reaction†

2002 ◽  
Vol 65 (9) ◽  
pp. 1371-1380 ◽  
Author(s):  
VIJAY K. SHARMA
Keyword(s):  
Real Time ◽  
Shiga Toxin ◽  
Enterohemorrhagic Escherichia Coli ◽  
Specific Primer ◽  
Chain Reaction ◽  
Pcr Assay ◽  
E Coli ◽  
Taqman Probes ◽  
Polymerase Chain ◽  
Pcr Assays

Enterohemorrhagic Escherichia coli (EHEC) O157:H7 and certain non-O157 EHEC serotypes (such as O26:H11, O26: NM, O111:H8, and O111:NM) have emerged as significant causes of human disease throughout the world. Important virulence attributes of EHEC are the intimin protein (encoded by the eae gene) and Shiga toxins 1 and 2 (encoded by the stx1 and stx2 genes, respectively). Two sets of real-time polymerase chain reaction (R-PCR) assays were developed for the simultaneous detection and quantitation of EHEC through the monitoring of the presence of the eae and stx genes, and these assays were evaluated. In the eaeR-PCR assay, three sets of primers and TaqMan probes were designed for the amplification and real-time detection of a portion of the eae gene specific to the EHEC O26, O111, and O157 serotypes. In the stxR-PCR assay, two sets of primers and TaqMan probes were used to amplify and detect the stx1 and stx2 genes. DNA prepared from 67 bacterial strains carrying known virulence markers was tested to determine the specificities of the two assays. In the eaeR-PCR assay, eaeO157- and eaeO111-specific primer-probe sets identified only EHEC O157 and O111 strains, respectively. The eaeO26-specific primer-probe set identified all EHEC O26 isolates and some Shiga toxin–negative serotypes of enteropathogenic E. coli and rabbit diarrheagenic E. coli. The stxR-PCR assay was able to identify only those strains carrying either or both of the Shiga toxin–encoding genes. The detection range of both R-PCR assays was linear over DNA concentrations corresponding to 103 to 108 CFU/ml of an EHEC strain. Both assays were able to detect and quantify very low levels (1 to 10 CFU/g of food or feces) of EHEC in feces and ground beef enriched for 16 h in a modified Trypticase soy broth. In conclusion, eae- and stx-based R-PCR assays are reliable and sensitive methods for the rapid screening and specific and quantitative detection of important serotypes of EHEC in cattle and in foods of bovine origin.

Prevalence and Characteristics of Shiga Toxin-Producing Escherichia coli in Beef Cattle Slaughtered on Prince Edward Island

2000 ◽  
Vol 63 (11) ◽  
pp. 1583-1586 ◽  
Author(s):  
R. DOUGLAS SCHURMAN ◽  
HARRY HARIHARAN ◽  
SUSAN B HEANEY ◽  
KRIS RAHN
Keyword(s):  
Escherichia Coli ◽  
Shiga Toxin ◽  
Prince Edward Island ◽  
Toxin Production ◽  
Positive Sample ◽  
Chain Reaction ◽  
Antimicrobial Drugs ◽  
E Coli ◽  
Human Illness ◽  
Polymerase Chain

Fecal swabs obtained from a random sample of 1,000 beef slaughter steers and heifers from 123 Prince Edward Island (P.E.I.) farms were examined for the presence of Shiga toxin-producing Escherichia coli (STEC) using a Vero cell assay (VCA). Multiple isolates from each positive sample were tested similarly. VCA-positive isolates were confirmed as E. coli biochemically, tested for drug resistance, serotyped, and tested by polymerase chain reaction (PCR). Animals were classified as positive when an isolate was positive on VCA and the presence of the gene responsible for toxin production was confirmed by PCR. The prevalence of STEC in beef slaughter steers and heifers on P.E.I. was 4% (40 of 1,000). The total number of isolates was 43, and these comprised 26 serotypes, including 13 isolates belonging to 6 serotypes known to be associated with human illness. The most frequently isolated STEC serotype was E. coli O157 (5 isolates out of 43). Of the five E. coli O157 isolates, four were E. coli O157:H7, a serious human pathogen. The majority of STEC isolates, including all O157:H7, isolates, were susceptible to 16 commonly used antimicrobial drugs. According to PCR, 65% of the STEC isolates had the gene for Stx1. Four of these isolates, including two O157: H7, had genes for Shiga toxin (Stx)1 and Stx2.

scholarly journals Fallow Deer (Dama dama) as a Reservoir of Shiga Toxin-Producing Escherichia coli (STEC)

Animals ◽  
2020 ◽  
Vol 10 (5) ◽  
pp. 881
Author(s):  
Anna Szczerba-Turek ◽  
Bernard Kordas
Keyword(s):  
Escherichia Coli ◽  
Polymerase Chain Reaction ◽  
Foodborne Pathogens ◽  
Shiga Toxin ◽  
Fallow Deer ◽  
Dama Dama ◽  
Chain Reaction ◽  
E Coli ◽  
Deer Population ◽  
Polymerase Chain

Shiga toxin-producing Escherichia (E.) coli (STEC) are responsible for the outbreaks of serious diseases in humans. Only a few reports on fallow deer as a reservoir of foodborne pathogens have been published to date. The purpose of this study was to determine the occurrence of STEC strains in the fallow deer population in Poland. In all, 94 fallow deer swabs were tested. Polymerase chain reaction (PCR) was performed to detect the virulence profile of stx1, stx2 and eae or aggR genes, to identify the subtypes of stx1 and stx2 genes and to perform O and H serotyping. STEC and attaching and effacing (AE)-STEC were identified in 13 isolates (13.83%). The most hazardous virulence profile was detected in three strains, namely stx2d serotype O103:HNM, eae/stx1a serotype O26:HNM and eae/stx1a serotype O157:H7. The predominant stx gene was stx2, which was identified in 76.92% of isolates. E. coli O157 was detected in 4/94 (4.26%). Other E. coli serogroups, O26, O103, O111 and O145, were identified in 14/94 fallow deer (14.89%). The present findings suggest that fallow deer are carriers of STEC/AE-STEC that are potentially pathogenic to humans.

Detection of Viable Shiga Toxin–Producing Escherichia coli by Quantitative Competitive Polymerase Chain Reaction

2003 ◽  
Vol 66 (7) ◽  
pp. 1277-1282 ◽  
Author(s):  
W. LI ◽  
M. A. DRAKE
Keyword(s):  
Escherichia Coli ◽  
Polymerase Chain Reaction ◽  
Cell Viability ◽  
Shiga Toxin ◽  
Ground Beef ◽  
Multiple Time ◽  
Chain Reaction ◽  
Competitive Polymerase Chain Reaction ◽  
E Coli ◽  
Polymerase Chain

With the use of Escherichia coli O157:H7 as a model, a procedure for the quantitative detection of viable Shiga toxin–producing E. coli (STEC) in broth and cooked ground beef enrichments with multiple–time point quantitative competitive polymerase chain reaction (QC-PCR) was developed. The A subunit (a 401-bp fragment) of the stx2 gene was chosen as a target sequence. Immunomagnetic separation (IMS) was used to isolate and concentrate cells from ground beef enrichments. Cell viability was confirmed on the basis of the quantitative increase in the signal of target bands from QC-PCR across multiple time points. The application of IMS increased detection limits relative to those for QC-PCR without IMS. E. coli O157:H7 inoculated at 0.20 CFU/g of cooked ground beef (25 g of ground beef plus 225 ml of Bacto modified EC medium plus novobiocin) was detected and confirmed to be viable in <15 h. A DNA-based molecular approach can be used to determine cell viability.

Sorbitol non-fermenting shiga toxin-producingEscherichia coliin cattle on smallholdings

2014 ◽  
Vol 143 (1) ◽  
pp. 94-103 ◽  
Author(s):  
M. Z. ISLAM ◽  
J. P. CHRISTENSEN ◽  
P. K. BISWAS
Keyword(s):  
Escherichia Coli ◽  
Polymerase Chain Reaction ◽  
Shiga Toxin ◽  
Gel Electrophoresis ◽  
Virulence Genes ◽  
Pulsed Field Gel Electrophoresis ◽  
Chain Reaction ◽  
E Coli ◽  
Faecal Samples ◽  
Polymerase Chain

SUMMARYWe investigated faecal samples collected from the rectum of 518 cattle on 371 randomly selected smallholdings in Bangladesh for the presence of sorbitol non-fermenting (SN-F) shiga toxin-producingEscherichia coli(STEC). The SN-F isolates were tested for the presence ofrfbO157,stx1, stx2, eaeandhlyAgenes by polymerase chain reaction (PCR). Seven SN-F isolates lacking these genes were profiled by pulsed-field gel electrophoresis (PFGE) to verify their clonality. SN-FE. coliwas identified in 44 [8·5%, 95% confidence interval (CI) 6·4–11·2] samples; of these, 28 (5·4%, 95% CI 3·8–7·7) had shiga toxin-producing strains, although only two carried therfbO157 gene. Thirteen isolates carried thehlyAgene while 18 harboured theeaegene. Based on PFGE, six pulsotypes were observed among the seven isolates that had no virulence genes. To the best of our knowledge this is the first report on shiga toxin-producingE. colifrom direct rectal faecal samples of cattle on smallholdings.

scholarly journals Molecular Characterization of ESBL-Producing Enterobacteriaceae in Northern Portugal

2014 ◽  
Vol 2014 ◽  
pp. 1-6 ◽  
Author(s):  
Rúben Fernandes ◽  
Paula Amador ◽  
Carla Oliveira ◽  
Cristina Prudêncio
Keyword(s):  
Klebsiella Oxytoca ◽  
Enterobacter Aerogenes ◽  
Generation Cephalosporin ◽  
Fourth Generation ◽  
Chain Reaction ◽  
E Coli ◽  
The North ◽  
Genetic Patterns ◽  
Polymerase Chain

Extended-spectrumβ-lactamases (ESBLs) prevalence was studied in the north of Portugal, among 193 clinical isolates belonging to citizens in a district in the boundaries between this country and Spain from a total of 7529 clinical strains. In the present study we recovered some members of Enterobacteriaceae family, producing ESBL enzymes, includingEscherichia coli(67.9%),Klebsiella pneumoniae(30.6%),Klebsiella oxytoca(0.5%),Enterobacter aerogenes(0.5%), andCitrobacter freundii(0.5%).β-lactamases genes blaTEM, blaSHV, and blaCTX-M were screened by polymerase chain reaction (PCR) and sequencing approaches. TEM enzymes were among the most prevalent types (40.9%) followed by CTX-M (37.3%) and SHV (23.3%). Among our sample of 193 ESBL-producing strains 99.0% were resistant to the fourth-generation cephalosporin cefepime. Of the 193 isolates 81.3% presented transferable plasmids harboringblaESBLgenes. Clonal studies were performed by PCR for the enterobacterial repetitive intragenic consensus (ERIC) sequences. This study reports a high diversity of genetic patterns. Ten clusters were found forE. coliisolates and five clusters forK. pneumoniaestrains by means of ERIC analysis. In conclusion, in this country, the most prevalent type is still the TEM-type, but CTX-M is growing rapidly.

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