| Use of CRISPR sequence polymorphisms for specific detection of enterohemorrhagic E. coli (EHEC) strains of serotypes O26:H11, O45:H2, O103:H2, O111:H8, O121:H19, O145:H28 and O157:H7 by real time PCR. | |
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MedLine Citation:
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PMID: 23035199 Owner: NLM Status: Publisher |
Abstract/OtherAbstract:
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We explored the genetic diversity of the clustered, regularly interspaced, short palindromic repeats (CRISPR) regions of enterohemorrhagic E. coli (EHEC) to design simplex real-time PCR assays for each of the seven worldwide most important EHEC serotypes. A panel of 958 E. coli investigated for their CRISPR loci by high throughput real-time PCR showed that CRISPR polymorphisms in E. coli strongly correlated with both O:H serotypes and presence of EHEC virulence factors (stx and eae genes). The CRISPR sequences chosen for simplex real-time PCR amplification of EHEC strains belonging to the top7 EHEC serogroups differentiated clearly between EHEC and non-EHEC strains. Specificity estimates of the CRISPR PCR assays varied from 97.5% to 100%. Sensitivity estimates of the assays ranged from 95.7% to 100%. The assays targeting EHEC O145:[H28], O103:[H2], and O45:[H2] displayed 100% sensitivity. The combined usage of two simplex PCR assays targeting different sequences of the O26 CRISPR locus allowed detection of EHEC O26:[H11] with 100% sensitivity. By combining two simplex PCR assays targeting different sequences of the EHEC O157 CRISPR locus, EHEC O157:[H7] was detected with 99.56% sensitivity. EHEC O111:[H8] and EHEC O121:[H19] were detected with 95.9% and 95.7% sensitivity respectively. This study demonstrates that the identification of EHEC serotype specific CRISPR sequences is more specific than the mere identification of O-antigen gene sequences as it is used in current PCR protocols for detection of EHEC strains. |
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Authors:
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Sabine Delannoy; Lothar Beutin; Patrick Fach |
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Publication Detail:
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Type: JOURNAL ARTICLE Date: 2012-10-3 |
Journal Detail:
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Title: Journal of clinical microbiology Volume: - ISSN: 1098-660X ISO Abbreviation: J. Clin. Microbiol. Publication Date: 2012 Oct |
Date Detail:
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Created Date: 2012-10-4 Completed Date: - Revised Date: - |
Medline Journal Info:
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Nlm Unique ID: 7505564 Medline TA: J Clin Microbiol Country: - |
Other Details:
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Languages: ENG Pagination: - Citation Subset: - |
Affiliation:
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Anses (French Agency for Food, Environmental and Occupational Health and Safety), Food Safety Laboratory, 23 Av du Général De Gaulle, Fr-94706 Maisons-Alfort, France. |
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From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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