Antimicrob Agents Chemother

A strain of the opportunistic pathogenic yeast was genetically modified for use as a cellular model for assessing by allele replacement the impact of lanosterol C14α-demethylase mutations on azole resistance. was chosen because it is susceptible to azole antifungals, it belongs to the CTG clade of yeast, which includes most of the species pathogenic for humans, and it is haploid and easily amenable to genetic transformation and molecular modeling. In this work, allelic replacement is targeted at the locus by the reconstitution of a functional auxotrophic marker in the 3' intergenic region of Homologous and heterologous alleles are expressed from the resident promoter of , allowing accurate comparison of the phenotypic change in azole susceptibility. As a proof of concept, we successfully expressed in different alleles, either bearing or not bearing mutations retrieved from a clinical context, from two phylogenetically distant yeasts, and constitutes a high-fidelity expression system, giving specific Erg11p-dependent fluconazole MICs very close to those observed with the donor strain. This work led us to characterize the phenotypic effect of two kinds of mutation: mutation conferring decreased fluconazole susceptibility in a species-specific manner and mutation conferring fluconazole resistance in several yeast species. In particular, a missense mutation affecting amino acid K143 of Erg11p in species, and the equivalent position K151 in , plays a critical role in fluconazole resistance

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

Distribution dans la mucoviscidose et écologie des différentes espèces du complexe Scedosporium apiospermum

The Pseudallescheria boydii / Scedosporium apiospermum species complex ranks second among the filamentous fungi isolated from sputum samples of patients with cystic fibrosis, the most common genetic inherited disease in the Caucasian population and whose prognosis is closely related to respiratory functional deterioration. Distribution of the five different species of the complex was investigated in French patients with cystic fibrosis. Our results suggest that P. boydii is the predominant species (62%), followed by S. apiospermum (24%), Scedosporium aurantiacum (10%) and Pseudallescheria minutispora (4%). We also demonstrated the ability of these two species to chronically colonize the respiratory tract of patients with cystic fibrosis. Natural niches and reservoirs of these species that are potential sources of human contamination were then studied in the Pays-de-la-Loire region, France. The highest densities of these species were found in human-impacted areas. Scedosporium dehoogii is the most abundant species; S. aurantiacum, P. boydii and S. apiospermum were mainly found in agricultural areas, seashores and playgrounds, respectively. As PCR techniques are faster and more sensitive than cultural methods and allow semi-quantification, we finally developed a molecular method for detection and quantification of the P. boydii / S. apiospermum species complex. However, an optimization of the DNA extraction step is required for application on soil samples.Le complexe Pseudallescheria boydii / Scedosporium apiospermum se situe au deuxième rang des champignons filamenteux isolés des expectorations de patients atteints de mucoviscidose, maladie génétique la plus fréquente dans la population caucasienne et dont le pronostic est étroitement lié à l'atteinte respiratoire. La distribution des cinq espèces constituant ce complexe a été étudiée au sein d'une cohorte de patients français atteints de mucoviscidose. Nos résultats suggèrent que P. boydii est l'espèce prédominante (62%), suivie de S. apiospermum (24%), Scedosporium aurantiacum (10%) et Pseudallescheria minutispora (4%). Nous avons également démontré la capacité de ces deux dernières espèces à coloniser de manière chronique les voies respiratoires de ces patients. Les niches naturelles et les réservoirs de ces espèces constituant des sources potentielles de contamination humaine ont ensuite été étudiées dans la région Pays-dela- Loire. Les densités les plus importantes de ces espèces ont été retrouvées dans les zones impactées par les activités humaines. Scedosporium dehoogii est l'espèce la plus abondante ; S. aurantiacum, P. boydii et S. apiospermum ont été principalement retrouvées dans les zones agricoles, les plages et les aires de jeux, respectivement. Les techniques de PCR étant plus rapides et plus sensibles que les méthodes culturales et permettant une semi-quantification, nous avons enfin développé une méthode moléculaire pour la détection et la quantification du complexe P. boydii / S. apiospermum. Cependant, une optimisation de l'étape d'extraction d'ADN génomique est nécessaire pour permettre l'application de cette méthode à des échantillons de sol

Distribution dans la mucoviscidose et écologie des différentes espèces du complexe Scedosporium apiospermum

Le complexe Pseudallescheria boydii / Scedosporium apiospermum se situe au deuxième rang des champignons filamenteux isolés des expectorations de patients atteints de mucoviscidose, maladie génétique la plus fréquente dans la population caucasienne et dont le pronostic est étroitement lié à l'atteinte respiratoire. La distribution des cinq espèces constituant ce complexe a été étudiée au sein d'une cohorte de patients français atteints de mucoviscidose. Nos résultats suggèrent que P. boydii est l'espèce prédominante (62%), suivie de S. apiospermum (24%), Scedosporium aurantiacum (10%) et Pseudallescheria minutispora (4%). Nous avons également démontré la capacité de ces deux dernières espèces à coloniser de manière chronique les voies respiratoires de ces patients. Les niches naturelles et les réservoirs de ces espèces constituant des sources potentielles de contamination humaine ont ensuite été étudiées dans la région Pays-dela- Loire. Les densités les plus importantes de ces espèces ont été retrouvées dans les zones impactées par les activités humaines. Scedosporium dehoogii est l'espèce la plus abondante ; S. aurantiacum, P. boydii et S. apiospermum ont été principalement retrouvées dans les zones agricoles, les plages et les aires de jeux, respectivement. Les techniques de PCR étant plus rapides et plus sensibles que les méthodes culturales et permettant une semi-quantification, nous avons enfin développé une méthode moléculaire pour la détection et la quantification du complexe P. boydii / S. apiospermum. Cependant, une optimisation de l'étape d'extraction d'ADN génomique est nécessaire pour permettre l'application de cette méthode à des échantillons de sol.The Pseudallescheria boydii / Scedosporium apiospermum species complex ranks second among the filamentous fungi isolated from sputum samples of patients with cystic fibrosis, the most common genetic inherited disease in the Caucasian population and whose prognosis is closely related to respiratory functional deterioration. Distribution of the five different species of the complex was investigated in French patients with cystic fibrosis. Our results suggest that P. boydii is the predominant species (62%), followed by S. apiospermum (24%), Scedosporium aurantiacum (10%) and Pseudallescheria minutispora (4%). We also demonstrated the ability of these two species to chronically colonize the respiratory tract of patients with cystic fibrosis. Natural niches and reservoirs of these species that are potential sources of human contamination were then studied in the Pays-de-la-Loire region, France. The highest densities of these species were found in human-impacted areas. Scedosporium dehoogii is the most abundant species; S. aurantiacum, P. boydii and S. apiospermum were mainly found in agricultural areas, seashores and playgrounds, respectively. As PCR techniques are faster and more sensitive than cultural methods and allow semi-quantification, we finally developed a molecular method for detection and quantification of the P. boydii / S. apiospermum species complex. However, an optimization of the DNA extraction step is required for application on soil samples.ANGERS-BU Médecine-Pharmacie (490072105) / SudocSudocFranceF

Scedo-Select III: a new semi-selective culture medium for detection of the Scedosporium apiospermum species complex

International audienceThe Scedosporium apiospermum complex is responsible for a large variety of infections in human. Members of this complex have become emerging fungal pathogens with an increasing ocurrence in patients with underlying conditions such as immunosuppression or cystic fibrosis. A better knowledge of these fungi and of the sources of contamination of the patients is required and more accurate detection methods from the environment are needed. In this context, a highly selective culture medium was developed in the present study. Thus, various aliphatic, cyclic, or aromatic compounds were tested as the sole carbon source, in combination with some inorganic nitrogen sources and fungicides. The best results were obtained with 4-hydroxy-benzoate combined with ammonium sulfate and the fungicides dichloran and benomyl. This new culture medium called Scedo-Select III was shown to support growth of all species of the S. apiospermum complex. Subsequently, this new culture medium was evaluated successfully on water and soil samples, exhibiting higher sensitivity and selectivity than the previously described SceSel+ culture medium. Therefore, this easy-to-prepare and synthetic semi-selective culture medium may be useful to clarify the ecology of these fungi and to identify their reservoirs in patients’ environment.</p

Trichosporon faecale invasive infection in a patient with severe aplastic anemia: Efficacy of voriconazole and liposomal amphotericin B before neutrophil recovery

We report a case of a 51-year old man with a severe aplastic anemia who developed an invasive trichosporonosis to Trichosporon faecale with fungemia and skin lesions during severe neutropenia. The treatment was successful before neutrophil recovery with a combination of voriconazole and liposomal amphotericin B

Saprochaete clavata invasive infection in a patient with severe aplastic anemia: Efficacy of voriconazole and liposomal amphotericin B with adjuvant granulocyte transfusions before neutrophil recovery following allogeneic bone marrow transplantation

We report a case of a 27-year old man with severe aplastic anemia who developed a Saprochaete clavata (Geotrichum clavatum) disseminated invasive infection shortly prior a scheduled allogeneic bone marrow transplantation. Treatment with a combination of voriconazole, liposomal amphotericin B and adjuvant granulocyte transfusions was successful before neutrophil recovery

Draft Genome Sequence of the Environmental Fungus Scedosporium dehoogii

International audienceToday, the genus Scedosporium comprises at least ten species with four of them, Scedosporium apiospermum, Scedosporium boydii, Scedosporium aurantiacum and Scedosporium minutisporum capable of colonizing the lungs of patients with cystic fibrosis. Scedosporium dehoogii, which is also common in the soil, has never been reported as causing human pulmonary infections. Here we report the first genome sequence for S. dehoogii, an invaluable resource to understand the genetic bases of pathogenesis in the genus Scedosporium

Detection of novel astroviruses among rodents of Gabon, Central Africa

International audienc