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Master Thesis

Recherche de mutants de Francisella tularensis résistants aux fluoroquinolones chez les patients atteints de tularémie

Abstract : In Europe, fluoroquinolones are considered as a first-line therapy of tularaemia. However, 10-15% of treated patients experience relapses, suggesting in vivo selection of fluoroquinolone resistant mutants in F. tularensis. The aim of the present study was to develop a molecular tool allowing detection of fluoroquinolone resistance in F. tularensis, to tentatively detect fluoroquinolone-resistant mutants in clinical samples. We designed a real time PCR assay for detection of mutations occurring in the Quinolone Resistance Determining Region (QRDR) of the gyrA gene, encoding subunit A of DNA gyrase, the main target of fluoroquinolones in F. tularensis. The assay was validated using fluoroquinolone-susceptible F. tularensis strains, and fluoroquinolone-resistant strains with gyrA mutations previously selected in our laboratory. The real-time PCR test allowed accurate differentiation of the wild-type strains from resistant mutants according to their respective melting temperatures. The specificity of our assay was confirmed using 32 clinical isolates of Francisella tularensis, characterized as wild type by sequencing of gyrA QRDR. We didn't find any gyrA mutant from clinical samples. We have elaborated a new real-time PCR assay allowing accurate detection of gyrA mutations in F. tularensis. This test can now be used on a routine basis to detect fluoroquinolone resistance either in isolated F. tularensis strains or directly in clinical samples. Our preliminary experience, however, indicates that in vivo selection of mutants after fluoroquinolone therapy is probably a rare event, and may not explain the frequent relapses observed in tularemia patients.
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Master Thesis
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Submitted on : Tuesday, July 2, 2013 - 2:00:12 PM
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  • HAL Id : dumas-00840391, version 1

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Gautier Hoarau. Recherche de mutants de Francisella tularensis résistants aux fluoroquinolones chez les patients atteints de tularémie. Médecine humaine et pathologie. 2012. ⟨dumas-00840391⟩

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