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Identification of virulotypes and serotypes of enteropathogenic (EPEC) and Shigatoxigenic (STEC) Escherichia coli from healthy cattle at slaughterhouses in Wallonia.
Takaki, Shino; Duprez, Jean-Noël; Fakih, Ibrahim et al.
2016AESA international congress “25 years of animal epidemiology: past, present and future”
 

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Abstract :
[en] Escherichia coli producing the attachment-effacement (AE) lesion (EPEC) and/or Shiga toxins (STEC) cause enteritis and (bloody) diarrhoea in young calves and in humans, and are also present in the intestines of healthy cattle. Besides the O157:H7 serotype, which is the main serotype causing STEC outbreaks in the world EPEC and STEC can belong to dozens of O serogroups. Of them, 9 have been frequently identified worldwide: O5, O26, O103, O104, O111, O118, O121, O145 and O165. The aim of this study is to identify the virulotypes and the O serotypes of EPEC and STEC isolated from healthy cattle at slaughterhouses in Wallonia. A total of 245 faeces (216 <1year-old bulls, 25 cows and 4 heifers) were sampled between April and June 2014 in 2 slaughterhouses in Wallonia and grown overnight at 37°C in Lauryl sulfate Enterobacteriaceae selective broth. The enrichment broths were assayed with an stx1, stx2 (Shiga toxin) and eae (AE lesion) triplex PCR and positive broths were inoculated onto 4 agar media: McConkey’s, Chromagar ES, Chromagar ES with tellurite and Chromagar STEC. Up to ten colonies per plate were picked up, sub-cultured and tested by the colony hybridization assay with gene probes targeting the stx1, stx2 and eae genes. The triplex PCR was again performed on all probe-positive isolates. The PCR-positive E. coli were subsequently assayed with two pentaplex PCR targeting the specific genes coding for the ten O serogroups listed above. Of the 2563 sub-cultured isolates, 744 isolates (29%) from 62 animals (25%) tested positive with the colony hybridization assay. Of them, 687 isolates (92%) from 59 animals were positive with the triplex PCR and the results of both tests were in agreement for 617 isolates (83%). One to 29 isolates per animal were probe- and PCR-positive. The positive isolates grew on Chromagar STEC (379; 55%), on Chromagar ES with tellurite (189; 28%), on Chromagar ES (62; 9%) or on McConkey’s agar (57; 8%). The most frequent virulotypes were eae+ (EPEC: 372 isolates; 54%), eae+stx1+ (AE_STEC: 119 isolates; 17%) and stx2+ (STEC: 118 isolates; 17%). In some animals different virulotypes were identified. The serogrouping with the two pentaplex PCR is in progress. AE-STEC, EPEC and STEC are excreted by 25% of the healthy cattle at slaughterhouses in Wallonia and different virulotypes can be excreted by the same animal. Conversely the methodology followed gives no precise idea of the actual level of excretion since the hybridization and PCR were performed after enrichment in selective broth. Therefore multiple isolates belonging to the same virulotype might represent the same clone. Identification of the serogroups and comparison by Pulsed Field Gel Electrophoresis should help to clarify that point. Quantitative (q)PCR is today the best method to quantify bacterial excretion, but is more expensive. The results of the hybridization and PCR correspond to between 80 and 90%. Though the colony hybridization is still useful for large-scale surveillance it needs radioactive probes for highest sensitivity and is more time-consuming than PCR. Therefore the PCR should be the first routine choice if it can be automatized at large scale. Further steps are the confirmation of the PCR results of the 70 isolates with different hybridization and PCR results and the identification of the serogroups with the two pentaplex PCR and later with PCR for the other serogroups, to compare them with isolates from young diarrhoeic calves and from humans.
Disciplines :
Veterinary medicine & animal health
Author, co-author :
Takaki, Shino
Duprez, Jean-Noël  ;  Université de Liège > Département des maladies infectieuses et parasitaires (DMI) > Département des maladies infectieuses et parasitaires (DMI)
Fakih, Ibrahim
Korsak Koulagenko, Nicolas ;  Université de Liège > Département de sciences des denrées alimentaires (DDA) > Département de sciences des denrées alimentaires (DDA)
Thiry, Damien ;  Université de Liège > Département des maladies infectieuses et parasitaires (DMI) > Bactériologie et pathologie des maladies bactériennes
Mainil, Jacques ;  Université de Liège > Département des maladies infectieuses et parasitaires (DMI) > Bactériologie et pathologie des maladies bactériennes
Language :
English
Title :
Identification of virulotypes and serotypes of enteropathogenic (EPEC) and Shigatoxigenic (STEC) Escherichia coli from healthy cattle at slaughterhouses in Wallonia.
Publication date :
September 2016
Event name :
AESA international congress “25 years of animal epidemiology: past, present and future”
Event place :
Liège, Belgium
Event date :
07/09/2016 - 09/09/2016
Audience :
International
Available on ORBi :
since 12 September 2016

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