Évaluation d une technique de rep-PCR semi-automatisée pour le génotypage de souches du complexe Pseudallescheria boydii / Scedosporium apiospermum au cours de la mucoviscidose

Species of the Scedosporium apiospermum complex rank the second among the filamentous fungi colonizing the airways of patients with cystic fibrosis (CF). It is clearly established now that these fungi may disseminate in case of immunodeficiency and that a chronic colonization of the airways by these pathogens may hinder the success of lung transplantation. In this study, we develop a new genotyping method to investigate the epidemiology of the airway colonization by these fungi. 63 multiple and sequential isolates of S. apiospermum collected from 9 CF patients, and selected among those previously studied by random amplification of polymorphic DNA (RAPD) by Defontaine et at. (2002) were analyzed. using the automated typing system DiversiLab® (bioMerieux) based on PCR amplification of repetitive sequences. Two different rep-PCR kits were compared; the DiverSiLab® Aspergillus and the DiversiLab® Fungal kit. The amplification products were separated by capillary electrophoresison; Each obtained profiles were then compared using. the DiversiLab® software, In addition, species identification of these isolates was clarified by sequencing TUB locus of the P-tubulin gene. The Fungal kit appeared to be more discriminating than the Aspergillus kit. Additionally, rep-PCR usually confirmed the colonization patterns described by RAPD. Only one patients showed distinct genotypes; In conclusion, the automated typing system DiversiLab® proved to be an easy and efficient Method to investigate the molecular epidemiology of the airway colonization by P. boydii / S. apiospermum complex species in CF. Moreover, our results confirm the capacity of the different species from this complex to chronically colonize the airways of CF patients despite the repeated antifungal cures.ROUEN-BU Médecine-Pharmacie (765402102) / SudocSudocFranceF

Semi-automated repetitive sequence-based PCR amplification for species of the Scedosporium apiospermum complex

International audiencePurpose: The Scedosporium apiospermum species complex usually ranks second among the filamentous fungi colonizing the airways of patients with cystic fibrosis (CF), but little is known about the molecular epidemiology of the airway colonization.Methods: Polymerase chain reaction (PCR) amplification of repetitive sequences (rep-PCR) was applied to the retrospective analysis of a panel of isolates already studied by random amplification of polymorphic DNA (RAPD) and comprising 63 isolates recovered from sputa from 9 CF patients. Results were compared to those obtained previously by RAPD, and herein by beta-tubulin (TUB) gene sequencing and Multilocus Sequence Typing (MLST).Results: Within the panel of isolates studied, S. apiospermum sensu stricto and Sce-dosporium boydii, as expected, were the predominant species with 21 and 36 isolates, respectively. Four isolates from one patient were identified as Scedosporium auranti-acum, whereas two isolates belonged to the Pseudallescheria ellipsoidea subgroup of S. boydii. rep-PCR analysis of these isolates clearly differentiated the three species and P. ellipsoidea isolates, whatever the rep-PCR kit used, and also permitted strain differentiation. When using the mold primer kit, results from rep-PCR were in close agreement with those obtained by MLST. For both S. apiospermum and S. boydii, 8 genotypes were differentiated by rep-PCR and MLST compared to 10 by RAPD. All S. aurantiacum isolates shared the same RAPD genotype and exhibited the same rep-PCR profile and sequence type.Conclusions: These results illustrate the efficacy of rep-PCR for both species identification within the S. apiospermum complex and genotyping for the two major species of this comple

Do Managers Overreact to Salient Risks? Evidence from Hurricane Strikes

We study how managers respond to hurricane events when their firms are located in the neighborhood of the disaster area. We find that the sudden shock to the perceived liquidity risk leads managers to increase corporate cash holdings and to express more concerns about hurricane risk in 10-Ks/10-Qs, even though the actual risk remains unchanged. Both effects are temporary. Over time, the perceived risk decreases, and the bias disappears. The distortion between perceived and actual risk is large, and the increase in cash is suboptimal. Overall, managerial reaction to hurricanes is consistent with salience theories of choice

Physiopathologie de la colonisation des voies respiratoires par Scedosporium apiospermum au cours de la mucoviscidose

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Ripple Effects of Noise on Corporate Investment

Firms significantly reduce their investment in response to non-fundamental drops in the stock price of their product-market peers. We argue that this result arises because of managers' limited ability to filter out the noise in stock prices when using them as signals about their investment opportunities. The resulting losses of capital investment and shareholders' wealth are economically large, and affect even firms that are not facing severe financing constraints or agency problems. Our findings offer a novel perspective on how stock market inefficiencies can affect the real economy, even in the absence of financing or agency frictions

Epidemiology of airway colonisation by Rasamsonia argillacea during cystic fibrosis

International audienceBackground :Species of the Rasamsonia argillacea complex are emerging fungi in patients with cystic fibrosis (CF) or chronic granulomatous disease (CGD). It was clearly established now that these fungi may disseminate in immunodeficient patients. The aim of this work was to evaluate the automated typing system DiversiLab for investigating the molecular epidemiology of airway colonization by species of the R. argillacea complex.Methods :142 isolates of the R. argillacea complex collected from 33 CF patients and one CGD patient from 10 European hospital centers were analyzed, together with 2 environmental isolates. The recently described automated typing system DiversiLab (bioMérieux)was based on PCR amplification of repetitive sequences using the pan-fungus DiversiLab Fungal kit. The amplification products were separated by capillary electrophoresis, resulting in a single profile for each isolate which were analyzed using the DiversiLab software. All isolates were identified at the species level by beta-tubulin sequencing.Results :Among the 144 isolates studied, 34 different genotypes were identified. Most isolates belonged to three clusters in close agreement with species identification. Some epidemiologically unrelated isolates from distinct patients, exhibited the same genotype. Moreover, R. argillacea isolates were recovered from sequential sputum samples from 15 CF patients, suggesting chronic colonization. This was confirmed for 13/15 patients with a unique genotype (8/13), or a largely dominant genotype (5/13). Different genotypes were found for sequential isolates collected from 2/15 patients. Four sequential isolates of the same were collected from one CGD patient.Conclusion :The DiversiLab system proved to be an easy and efficient method to investigate the molecular epidemiology of R. argillacea in CF or CGD patients. Results confirm the capacity of R. argillacea species to chronically colonize CF patient airways.</p

Rep-PCR (DiversiLab, bioMérieux), une méthode automatisée pour des études épidémiologiques de la colonisation fongique des voies respiratoires au cours de la mucoviscidose

International audienc

Sequential variations of Rasamsonia spp. isolate genotypes in cystic fibrosis patients

International audienceRasamsonia species have been recently described as emergent pathogens in human and animals. Recent multilocus phylogenetic studies have shown that the genus Rasamsonia (R.) includes at least four phylogenetically related species forming the R. argillacea complex, R. argillacea sensu stricto, R. piperina, R. eburnea and R. aegroticola, which have been mostly reported in the clinical context of chronic granulomatous disease (CGD) and cystic fibrosis (CF). Differences in sensitivity to antifungal therapies which have been noted within the genus Rasamsonia underline the importance of early identification of clinical isolates for optimal treatment during the early stages of infection. In this context, the aim of this work was to obtain genotypic characterization of clinical Rasamsonia spp. isolates from 26 CF patients and one acutely infected CGD patient using PCR amplification of repetitive DNA sequences (rep-PCR) and to compare species distributionsdetermined by β tubulin sequencing with rep-PCR genotypes/clusters.One hundred and ten isolates were collected sequentially or simultaneously from 01/09/1998 to 08/2012 from sputum samples of 26 CF patients. One CGD patient (4 isolates) was studied in parallel. Two references strains (CBS #408.73 (R. piperina) and #432.62 (R. cylindrospora) were obtained from CBS. After extraction, PCR amplifications of repetitive DNA sequences were realized for each sample using the DiversiLab Fungal kit (bioMérieux). Amplicons were separated by capillary electrophoresis in an Agilent 2100 Bioanalyzer (Massy, France). Single electrophoretic profiles obtained for each isolate were compared using the DiversiLab Healthcare software (version 3.41).As previously reported in CF patients, beta-tubulin gene sequencing of 110 isolates identified more frequently R. aegroticola (72 isolates from 19 CF patients) than R. argillacea stricto sensu (30 isolates from 8 CF patients), and R. piperina (8 isolates from 5 CF patients), with 15, 6, and 7 distinct profiles, respectively. From the CGD patient, all isolates were identified as R. argillacea stricto sensu. Without exception, species identification according to rep-PCR genotypes was entirely consistent with beta-tubulin sequencing results.Present data provide the first description of sequential Rasamsonia spp. variations with time in CF patients. Rasamsonia argillacea sensu stricto isolates were found persistent for years in all sequentially sampled CF patients except 1 who presented 4 different successive genotypes. In contrast to R. argillacea sensustricto, sequential detection of R. aegroticola and R. piperina revealed frequent successive changes in isolate genotypes in most CF patients.</p

Efficacité de la micafungine dans une infection à Geosmithia argillacea chez une patiente atteinte de mucoviscidose

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Clinical and microbiological efficacy of micafungin on Geosmithia argillacea infection in a cystic fibrosis patient

International audienceCystic fibrosis (CF) patients are at high risk of colonization of the airways by a number of fungi, including the emerging opportunistic fungus Geosmithia argillacea. We report the eradication of respiratory G. argillacea associated with clinical resolution of severe symptoms by high-dose and prolonged micafungin therapy in a young CF patient.</p