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Master Thesis

Apport du séquençage nouvelle génération au typage MLST de Pneumocystis jirovecii dans le cadre d'une épidémie chez des patients transplantés d'organe solide

Abstract : Multilocus Sequence Typing (MLST) allows accurate strain characterization, but Sanger sequencing presents limitations for subpopulations determination. Yet, mixed strains coinfections are frequent in Pneumocystis jirovecii Pneumonia (PCP). Next Generation Sequencing (NGS) allows both majority and minority variant studies. We evaluated the added value of NGS in an epidemiological MLST study of a PCP cluster in solid organ transplant (SOT) recipients. Thirty-two patients (12 grouped cases SOT and 20 controls) presenting with PCP at Grenoble Hospital, France, were included between 2012 and 2015. A three loci scheme was used for MLST analysis (MIT26S, CYTB, SOD). New primers were designed to obtain ~ 750 bp amplicons (GS +). Sequencing was achieved on 32 samples with a NGS technology (GS junior +, Roche), and with Sanger technology for 12 of them. Among the 12 SOT, 5 shared a major genotype C2a. Proportion of mixed genotypes infections was significantly higher with NGS analysis (65%) compared to Sanger (25%). NGS approach revealed infections with more than 2 subpopulations (53%) and the presence of ultra-minority strains (1%) among which the C2a genotype. Transmission map confirmed the probable nosocomial PCP acquisition. The 732bp MIT26S sequence revealed three new SNPs and a new MNP which increased significantly the discriminatory power of MIT26S locus. As expected, in this context of PCP, NGS outperforms Sanger approach for sub-population determination and revealed new discriminating SNPs. These findings bring new insight for future epidemiological studies on this non cultivable opportunistic fungus.
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Master Thesis
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https://dumas.ccsd.cnrs.fr/dumas-01564760
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Submitted on : Wednesday, July 19, 2017 - 10:46:47 AM
Last modification on : Wednesday, July 15, 2020 - 8:56:03 AM

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  • HAL Id : dumas-01564760, version 1

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Eléna Charpentier. Apport du séquençage nouvelle génération au typage MLST de Pneumocystis jirovecii dans le cadre d'une épidémie chez des patients transplantés d'organe solide. Sciences pharmaceutiques. 2016. ⟨dumas-01564760⟩

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