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© Institut Pasteur/Antoinette Ryter
Salmonella spp. Bactéries à Gram négatif, aérobies ou anaérobies facultatifs à transmission orofécale. Les salmonelles majeures (sérotype typhi et sérotype paratyphi) sont responsables des fièvres typhoïde et paratyphoïde chez l'homme uniquement ; les salmonelles mineures (sérotype typhimurium et sérotype enteritidis) sont impliquées dans 30 à 60 % des gastroentérites et toxiinfections d'origine alimentaire. Image colorisée.
Publication : PLoS neglected tropical diseases

CRISPR is an optimal target for the design of specific PCR assays for salmonella enterica serotypes Typhi and Paratyphi A

Scientific Fields
Diseases
Organisms
Applications
Technique

Published in PLoS neglected tropical diseases - 30 Jan 2014

Fabre L, Le Hello S, Roux C, Issenhuth-Jeanjean S, Weill FX

Link to Pubmed [PMID] – 24498453

PLoS Negl Trop Dis 2014;8(1):e2671

BACKGROUND: Serotype-specific PCR assays targeting Salmonella enterica serotypes Typhi and Paratyphi A, the causal agents of typhoid and paratyphoid fevers, are required to accelerate formal diagnosis and to overcome the lack of typing sera and, in some situations, the need for culture. However, the sensitivity and specificity of such assays must be demonstrated on large collections of strains representative of the targeted serotypes and all other bacterial populations producing similar clinical symptoms.

METHODOLOGY: Using a new family of repeated DNA sequences, CRISPR (clustered regularly interspaced short palindromic repeats), as a serotype-specific target, we developed a conventional multiplex PCR assay for the detection and differentiation of serotypes Typhi and Paratyphi A from cultured isolates. We also developed EvaGreen-based real-time singleplex PCR assays with the same two sets of primers.

PRINCIPAL FINDINGS: We achieved 100% sensitivity and specificity for each protocol after validation of the assays on 188 serotype Typhi and 74 serotype Paratyphi A strains from diverse genetic groups, geographic origins and time periods and on 70 strains of bacteria frequently encountered in bloodstream infections, including 29 other Salmonella serotypes and 42 strains from 38 other bacterial species.

CONCLUSIONS: The performance and convenience of our serotype-specific PCR assays should facilitate the rapid and accurate identification of these two major serotypes in a large range of clinical and public health laboratories with access to PCR technology. These assays were developed for use with DNA from cultured isolates, but with modifications to the assay, the CRISPR targets could be used in the development of assays for use with clinical and other samples.

https://www.ncbi.nlm.nih.gov/pubmed/24498453