TY - JOUR T1 - The GEN-ERA toolbox: unified and reproducible workflows for research in microbial genomics JF - GigaScience Y1 - 2023 A1 - Luc Cornet A1 - Benoit Durieu A1 - F. Baert A1 - Elizabet D'hooge A1 - David Colignon A1 - Loic Meunier A1 - Valérian Lupo A1 - Ilse Cleenwerck A1 - Heide-Marie Daniel A1 - Rigouts, Leen A1 - Damien Sirjacobs A1 - Stéphane Declerck A1 - Vandamme, Peter A1 - Annick Wilmotte A1 - Denis Baurain A1 - Pierre Becker KW - culture collections KW - Cyanobacteria KW - Genomics KW - Gloeobacterales KW - Metagenomics KW - nextflow KW - phylogenomics KW - Phylogeny KW - Singularity containers KW - Workflow AB -

Background: Microbial culture collections play a key role in taxonomy by studying the diversity of their strains and providing wellcharacterized biological material to the scientific community for fundamental and applied research. These microbial resource centers thus need to implement new standards in species delineation, including whole-genome sequencing and phylogenomics. In this context, the genomic needs of the Belgian Coordinated Collections of Microorganisms were studied, resulting in the GEN-ERA toolbox. The latter is a unified cluster of bioinformatic workflows dedicated to both bacteria and small eukaryotes (e.g., yeasts). Findings: This public toolbox allows researchers without a specific training in bioinformatics to perform robust phylogenomic analyses. Hence, it facilitates all steps from genome downloading and quality assessment, including genomic contamination estimation, to tree reconstruction. It also offers workflows for average nucleotide identity comparisons and metabolic modeling. Technical details: Nextflow workflows are launched by a single command and are available on the GEN-ERA GitHub repository (https: //github.com/Lcornet/GENERA). All the workflows are based on Singularity containers to increase reproducibility. Testing: The toolbox was developed for a diversity of microorganisms, including bacteria and fungi. It was further tested on an empirical dataset of 18 (meta)genomes of early branching Cyanobacteria, providing the most up-to-date phylogenomic analysis of the Gloeobacterales order, the first group to diverge in the evolutionary tree of Cyanobacteria. Conclusion: The GEN-ERA toolbox can be used to infer completely reproducible comparative genomic and metabolic analyses on prokaryotes and small eukaryotes. Although designed for routine bioinformatics of culture collections, it can also be used by all researchers interested in microbial taxonomy, as exemplified by our case study on Gloeobacterales.

VL - 12 M3 - 10.1093/gigascience/giad022 ER - TY - JOUR T1 - Multicentric Analysis of the Species Distribution and Antifungal Susceptibility of Clinical Isolates from Aspergillus Section Circumdati JF - Antimicrobial Agents and Chemotherapy Y1 - 2023 A1 - S. Imbert A1 - A. C. Normand A1 - D. Costa A1 - F. Gabriel A1 - L. Lachaud A1 - C. Schuttler A1 - S. Cassaing A1 - C. Mahinc A1 - L. Hasseine A1 - M. Demar A1 - S Brun A1 - C. Bonnal A1 - A. Moreno-Sabater A1 - Pierre Becker A1 - R. Piarroux A1 - A. Fekkar KW - antifungal susceptibility testing KW - Aspergillus ochraceopetaliformis KW - Aspergillus sclerotiorum KW - Aspergillus section Circumdati KW - Aspergillus westerdijkiae KW - azole resistance KW - MALDI-TOF mass spectrometry AB -

The clinical involvement and antifungal susceptibility of Aspergillus section Circumdati are poorly known. We analyzed 52 isolates, including 48 clinical isolates, belonging to 9 species inside the section Circumdati. The whole section exhibited, by the EUCAST reference method, a poor susceptibility to amphotericin B, but species/ series-specific patterns were observed for azole drugs. This underlines the interest in getting an accurate identification inside the section Circumdati to guide the choice of antifungal treatment in clinical practice.

VL - 67 CP - 4 M3 - 10.1128/aac.01462-22 ER - TY - JOUR T1 - Pulmonary Aspergillosis in Humboldt Penguins—Susceptibility Patterns andMolecular Epidemiology of Clinical and Environmental Aspergillus fumigatus Isolates from a Belgian Zoo JF - Antibiotics Y1 - 2023 A1 - Hanne Debergh A1 - Pierre Becker A1 - Vercammen,F. A1 - K. Lagrou A1 - Roel Haesendonck A1 - Saegerman,C. A1 - Ann Packeu KW - antifungal susceptibility testing KW - Aspergillus fumigatus KW - avian aspergillosis KW - azole resistance KW - cyp51A KW - genotyping KW - MIC KW - microsatellite typing KW - One Health KW - Spheniscus humboldti AB -

Aspergillus fumigatus is the main causative agent of avian aspergillosis and results in significant health problems in birds, especially those living in captivity. The fungal contamination by A. fumigatus in the environment of Humboldt penguins (Spheniscus humboldti), located in a Belgian zoo, was assessed through the analysis of air, water, sand and nest samples during four non-consecutive days in 2021–2022. From these samples, potential azole-resistant A. fumigatus (ARAF) isolates were detected using a selective culture medium. A total of 28 veterinary isolates obtained after necropsy of Humboldt penguins and other avian species from the zoo were also included. All veterinary and suspected ARAF isolates from the environment were characterized for their azole-resistance profile by broth microdilution. Isolates displaying phenotypic resistance against at least one medical azole were systematically screened for mutations in the cyp51A gene. A total of 14 (13.6%) ARAF isolates were identified from the environment (n = 8) and from Humboldt penguins (n = 6). The TR34/L98H mutation was observed in all resistant environmental strains, and in two resistant veterinary strains. To the best of our knowledge, this is the first description of this mutation in A. fumigatus isolates from Humboldt penguins. During the period 2017–2022, pulmonary aspergillosis was confirmed in 51 necropsied penguins, which reflects a death rate due to aspergillosis of 68.0%, mostly affecting adults. Microsatellite polymorphism analysis revealed a high level of diversity among environmental and veterinary A. fumigatus isolates. However, a cluster was observed between one veterinary isolate and six environmental strains, all resistant to medical azoles. In conclusion, the environment of the Humboldt penguins is a potential contamination source of ARAF, making their management even more complex.

VL - 12 CP - 3 M3 - https://doi.org/10.3390/antibiotics12030584 ER - TY - JOUR T1 - Pulmonary Aspergillosis in Humboldt Penguins—Susceptibility Patterns and Molecular Epidemiology of Clinical and Environmental Aspergillus fumigatus Isolates from a Belgian Zoo, 2017–2022 JF - Antibiotics Y1 - 2023 A1 - Hanne Debergh A1 - Pierre Becker A1 - Vercammen, Francis A1 - Lagrou, Katrien A1 - Roel Haesendonck A1 - Saegerman, Claude A1 - Ann Packeu VL - 12 CP - 3 M3 - 10.3390/antibiotics12030584 ER - TY - JOUR T1 - Remarkable fungal biodiversity on northern Belgium bats and hibernacula JF - Mycologia Y1 - 2023 A1 - Pierre Becker A1 - Claudia Van den Eynde A1 - F. Baert A1 - Elizabet D’hooge A1 - Robby De Pauw A1 - Normand, Anne-Cécile A1 - Piarroux, Renaud A1 - Dirk Stubbe KW - Mycobiome KW - Pseudogymnoascus cavicola KW - Pseudogymnoascus destructans KW - white-nose syndrome AB -

Bats can be affected by fungal pathogens such as Pseudogymnoascus destructans, the causative agent
of the white-nose syndrome. Their body surface can also be colonized by fungal commensals or carry
transient fungal species and participate in their dispersal. In this study, 114 bat specimens belonging
to seven species were sampled from various locations in northern Belgium. Culture-based methods
revealed an important mycological diversity, with a total of 209 different taxa out of the 418 isolates.
Overall, a mean of 3.7 taxa per bat was recorded, but significant differences were observed between
sampling sites and seasons. The mycobiomes were dominated by cosmopolitan and plant-associated
species, in particular from the genera Cladosporium, Penicillium, and Aspergillus. Other species known
to be related to bats or their environment, such as Apiotrichum otae, were also retrieved. Sampling of
hibernacula indicated that diverse fungal species can inhabit these sites, including a yet undescribed
Pseudogymnoascus species, distinct from Ps. destructans, namely, Ps. cavicola.

VL - 115 CP - 4 M3 - 10.1080/00275514.2023.2213138 ER - TY - Generic T1 - Towards Characterization and genotoxicity assessment of mycotoxin mixtures present in food. Y1 - 2023 A1 - Julie Sanders A1 - George Johnson A1 - Roel Anthonissen A1 - Pierre Becker A1 - Emmanuel Tangni A1 - Julien Masquelier A1 - Tamara Vanhaecke A1 - Birgit Mertens KW - additivity KW - benchmark dose modelling KW - genotoxicity KW - mixtures KW - Mycotoxins JF - BelTox Annual Meeting CY - Brussels, Belgium ER - TY - JOUR T1 - High diversity of yeast species and strains responsible for vulvovaginal candidiasis in South-East Gabon JF - Journal of Medical Mycology Y1 - 2022 A1 - M. Bignoumba A1 - R. Onanga A1 - B.S. Kumulungui A1 - R.F. Kassa Kassa A1 - Y. Mouanga Ndzime A1 - K. Mbombe Moghoa A1 - Dirk Stubbe A1 - Pierre Becker KW - Africa KW - Candida nivariensis KW - Genotyping Techniques KW - MALDI-TOF MS KW - vulvovaginal candidiasis M3 - 10.1016/j.mycmed.2022.101354 ER - TY - JOUR T1 - High diversity of yeast species and strains responsible for vulvovaginal candidiasis in South-East Gabon. JF - J Mycol Med Y1 - 2022 A1 - M Bignoumba ED - R Onanga ED - B S Kumulungui ED - R F Kassa Kassa ED - Y Mouanga Ndzime ED - K Mbombe Moghoa AB -

OBJECTIVES: Candida albicans generally remains the principal pathogenic yeast responsible for vulvovaginal candidiasis (VVC), although with variable prevalence. In this study, we evaluated the evolution of the prevalence of the non-Candida albicans Candida (NCAC) species and investigated the genotypic diversity and the population genetic structure of the circulating C. albicans strains associated with VVC in the vicinity of Franceville (Gabon).

METHODS: A total of 110 independent isolates were identified using both MALDI-TOF MS and conventional techniques. The population genetic structure of the C. albicans strains was determined by multiple locus variable-number tandem repeat analysis using 4 microsatellite markers.

RESULTS: The mean and median age of the patients was 31 years. Seven patients had a mixed infection. C. albicans accounted for 62 % (n=68) of the total isolates. NCAC were dominated by C. glabrata, followed by P. kudriavzevii, C. parapsilosis, C. tropicalis, M. guilliermondii, and C. nivariensis. The cluster analysis revealed a high diversity, with a total of 50 different genotypes. The most represented genotype was shared by only four strains, while the vast majority (39 strains) had a unique MLVA pattern. Geographic clusters were not detected.

CONCLUSION: The study provides information on species distribution and possible changing epidemiology while reporting for the first time C. nivariensis in VVC in Africa. This study is also the first to investigate the genotypic diversity of the circulating C. albicans strains associated with VVC in Central Africa. Such analyses would help understand the molecular epidemiology of C. albicans.

VL - 33 CP - 2 M3 - 10.1016/j.mycmed.2022.101354 ER - TY - JOUR T1 - Identification of Molds with Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry: Performance of the Newly Developed MSI-2 Application in Comparison with the Bruker Filamentous Fungi Database and MSI-1. JF - J Clin Microbiol Y1 - 2021 A1 - Normand, Anne-Cécile A1 - Marion Blaize A1 - Sébastien Imbert A1 - Arnaud Fekkar A1 - Piarroux, Renaud KW - Aspergillus KW - Databases, Factual KW - Fungi KW - Fusarium KW - Humans KW - Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization AB -

Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) represents a promising tool for the rapid and efficient identification of molds, but improvements are still necessary to achieve satisfactory results when identifying cryptic species. Here, we aimed to validate a new web application, MSI-2, which replaces MSI-1, an application that was built and deployed online in 2017. For the evaluation, we gathered 633 challenging isolates obtained from daily hospital practice that were first identified with DNA-based methods, and we submitted their corresponding mass spectra to three identification programs (Bruker, MSI-1, and MSI-2). The MSI-2 application had a better identification performance at the species level than MSI-1 and Bruker, reaching 83.25% correct identifications, compared with 63.19% (MSI-1), 38.07% (Bruker with a 1.7 threshold), and 21.8% (Bruker with a 2.0 threshold). The MSI-2 application performed especially well for Aspergillus and Fusarium species, including for many cryptic species, reaching 90% correct identifications for Aspergillus species and 78% for Fusarium species compared to 69% and 43% with MSI-1. Such an improvement may have a positive impact on patient management by facilitating the identification of cryptic species potentially associated with a specific antifungal resistance profile.

VL - 59 CP - 10 M3 - 10.1128/JCM.01299-21 ER - TY - JOUR T1 - Identification of Molds with Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry: Performance of the Newly Developed MSI-2 Application in Comparison with the Bruker Filamentous Fungi Database and MSI-1ABSTRACT JF - Journal of Clinical Microbiology Y1 - 2021 A1 - Normand, Anne-Cécile A1 - Marion Blaize A1 - Sébastien Imbert A1 - Ann Packeu A1 - Pierre Becker A1 - Arnaud Fekkar A1 - Dirk Stubbe A1 - Piarroux, Renaud KW - Fungi KW - identification KW - MALDI-TOF MS KW - pathogen VL - 59 CP - 10 M3 - 10.1128/JCM.01299-21 ER - TY - JOUR T1 - The taxonomy of the Trichophyton rubrum complex: a phylogenomic approach JF - Microbial Genomics Y1 - 2021 A1 - Luc Cornet A1 - Elizabet D'hooge A1 - Nicolas Magain A1 - Dirk Stubbe A1 - Ann Packeu A1 - Denis Baurain A1 - Pierre Becker KW - Dermatophytes KW - Fungi KW - Genomics KW - Phylogeny KW - Taxonomy VL - 7 CP - 11 M3 - 10.1099/mgen.0.000707 ER - TY - JOUR T1 - Blastomycosis in Africa and the Middle East: A Comprehensive Review of Reported Cases and Reanalysis of Historical Isolates Based on Molecular DataAbstractBackgroundMethodsResultsConclusions JF - Clinical Infectious Diseases Y1 - 2020 A1 - Ilan S Schwartz A1 - Jose F Muñoz A1 - Chris R Kenyon A1 - Nelesh P Govender A1 - Lisa McTaggart A1 - Tsidiso G Maphanga A1 - Susan Richardson A1 - Pierre Becker A1 - Christina A Cuomo A1 - Juan G McEwen A1 - Lynne Sigler KW - Blastomycosis KW - Fungi VL - 73 CP - 7 M3 - 10.1093/cid/ciaa1100 ER - TY - JOUR T1 - Screening strategy targeting the presence of food enzyme-producing fungi in food enzyme preparations JF - Food Control Y1 - 2020 A1 - Marie Deckers A1 - Kevin Vanneste A1 - Raf Winand A1 - Marijke Hendrickx A1 - Pierre Becker A1 - Sigrid C.J. De Keersmaecker A1 - Deforce, Dieter A1 - Marie-Alice Fraiture A1 - Nancy Roosens KW - Food enzymes KW - Fungi VL - 117 M3 - 10.1016/j.foodcont.2020.107295 ER - TY - JOUR T1 - Superficial mycoses in Belgium: Burden, costs and antifungal drugs consumption JF - Mycoses Y1 - 2020 A1 - Pierre Becker A1 - Pauline Lecerf A1 - Julie Claereboudt A1 - Brecht Devleesschauwer A1 - Ann Packeu A1 - Marijke Hendrickx KW - antifungal drugs consumption KW - Belgium KW - burden of disease KW - Candidiasis KW - Cost KW - dermatophytosis KW - Disability-Adjusted Life Years KW - superficial mycosis AB -

Background

Monitoring of superficial mycoses requires more attention due to their important incidence, health costs and antifungal drugs consumption.

Objectives

The objectives were to estimate the burden of superficial mycoses in Belgium and to assess trends in associated antifungal consumption.

Methods

The burden of dermatophytoses (including onychomycosis), as well as skin and genital candidiasis, was estimated using disability‐adjusted life years (DALY). Moreover, trends in systemic and topical antifungal consumption in ambulatory care were examined for the period 2010‐2017, together with their associated costs.

Results

Due to their high incidence and long treatment duration, dermatophytoses represented the bulk of the burden, accounting for 92.2% of the total DALYs of superficial mycoses. Terbinafine was the most prescribed antifungal in terms of doses (35.4% of the total doses) while fluconazole was the most delivered drug in terms of packages (29.1% of the total packages). More than 70% of the prescriptions were made by general practitioners while consumption varied according to age and gender of the patients. A global 12% decrease in antifungal prescriptions was observed between 2011 and 2017. However, this reduction would result mainly from packaging changes and increased self‐medication. A significant decrease in itraconazole treatments was notably compensated by an increased prescription of fluconazole packages.

Conclusion

This study emphasises that dermatological presentations of superficial mycoses are the most important in terms of both burden and antifungal consumption in Belgium. Further reduction in antifungals use can be achieved by applying the adequate treatment after identification of the causative agent.

VL - 63 CP - 5 M3 - 10.1111/myc.13063 ER - TY - JOUR T1 - Unexpected mould diversity in clinical isolates from French Guiana and associated identification difficulties. JF - Med Mycol Y1 - 2020 A1 - C Nabet A1 - S Imbert A1 - A C Normand A1 - D Blanchet A1 - R Chanlin A1 - Pierre Becker A1 - M Demar A1 - R Piarroux KW - Fungi KW - identification KW - MALDI-TOF MS KW - pathogen AB -

New mold species are increasingly reported in invasive fungal infections. However, these fungi are often misdiagnosed or undiagnosed due to the use of inappropriate laboratory diagnostic tools. Tropical countries, such as French Guiana, harbor a vast diversity of environmental fungi representing a potential source of emerging pathogens. To assess the impact of this diversity on the accuracy of mold-infection diagnoses, we identified mold clinical isolates in French Guiana during a five-month follow-up using both microscopy and matrix-assisted laser desorption ionization time-of-flight mass spectrometry. In total, 38.8% of the 98 obtained molds isolates could not be identified and required a DNA-based identification. Fungal diversity was high, including 46 species, 26 genera, and 13 orders. Fungal ecology was unusual, as Aspergillus species accounted for only 27% of all isolates, and the Nigri section was the most abundant out of the six detected Aspergillus sections. Macromycetes (orders Agaricales, Polyporales, and Russulales) and endophytic fungi accounted for respectively 11% and 14% of all isolates. Thus, in tropical areas with high fungal diversity, such as French Guiana, routine mold identification tools are inadequate. Molecular identifications, as well as morphological descriptions, are necessary for the construction of region-specific mass spectrum databases. These advances will improve the diagnosis and clinical management of new fungal infections.

LAY SUMMARY: In French Guiana, environmental fungal diversity may be a source of emerging pathogens. We evaluated microscopy and mass spectrometry to identify mold clinical isolates. With 39% of unidentified isolates, a region-specific mass spectrum database would improve the diagnosis of new fungal infections.

VL - 59 M3 - 10.1093/mmy/myaa091 ER - TY - JOUR T1 - Identification of fungal isolates by MALDI-TOF mass spectrometry in veterinary practice: validation of a web application. JF - J Vet Diagn Invest Y1 - 2019 A1 - Pierre Becker A1 - Normand, Anne-Cécile A1 - Gerty Vanantwerpen A1 - Vanrobaeys, Mia A1 - Roel Haesendonck A1 - Vercammen, Francis A1 - Dirk Stubbe A1 - Piarroux, Renaud A1 - Marijke Hendrickx KW - Animals KW - Animals, Zoo KW - Cattle KW - Databases, Factual KW - Fungi KW - Multilocus Sequence Typing KW - Online Systems KW - Pets KW - Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization KW - Veterinary Medicine KW - yeasts AB -

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) has emerged as a reliable method to identify fungal isolates. The success of this approach relies on the availability of exhaustive databases, but the latter were built with a focus on human pathogens. We assessed a large in-house database of reference spectra and a dedicated web application for their suitability for use in veterinary laboratories. A panel of 290 mold and yeast isolates representing 69 different fungal species was isolated from various animals (including pets, cattle, and zoo animals) and identified using both MALDI-TOF MS and conventional techniques. The performance of the 2 methods was compared, and identifications were confirmed by DNA sequencing. MALDI-TOF MS allowed distinction between some closely related species and achieved 89% correct identification at the species level. In comparison, only 60% of the isolates were correctly identified with conventional approaches. Using this online application, MALDI-TOF MS thus appears to be a relevant alternative for the identification of fungal isolates encountered by animal health professionals.

VL - 31 CP - 3 M3 - 10.1177/1040638719835577 ER - TY - JOUR T1 - Public microbial resources centres: key hubs for FAIR microorganisms and genetic materials. JF - Appl Environ Microbiol Y1 - 2019 A1 - Pierre Becker A1 - M Bosschaerts A1 - P Chaerle A1 - H-M Daniel A1 - Hellemans, A A1 - A Olbrechts A1 - Rigouts, L A1 - A Wilmotte A1 - Marijke Hendrickx AB -

In the context of open science, the availability of research materials is essential for knowledge accumulation and to maximize the impact of scientific research. In microbiology, microbial domain biological resource centres (mBRCs) have a long-standing experience in preserving and distributing authenticated microbial strains and genetic materials (e.g., recombinant plasmids, DNA libraries) to support new discoveries and follow-on studies. These culture collections play a central role in the conservation of microbial biodiversity and have expertise in cultivation, characterization and taxonomy of microorganisms. Information associated with preserved biological resources are recorded in databases and accessible through online catalogues. Legal expertise developed by mBRCs guarantees end users with the traceability and legality of the acquired material, notably with respect to the Nagoya Protocol. Yet, the awareness of the advantages to deposit biological materials in professional repositories remains low and the necessity to secure strains and genetic resources for future research must be emphasised. This review describes the unique position of mBRCs in microbiology and molecular biology through their history, evolving roles, expertise, services, challenges and international collaborations. It also calls for an increased deposit of strains and genetic resources, a responsibility shared by scientists, funding agencies and publishers. Journal policies requesting deposit during submission of a manuscript is already one of the measures to make more biological materials available to the broader community, hence fully releasing their potential and improving openness and reproducibility in scientific research.

M3 - 10.1128/AEM.01444-19 ER - TY - JOUR T1 - Systemic antifungal drug use in Belgium-One of the biggest antifungal consumers in Europe. JF - Mycoses Y1 - 2019 A1 - Berdieke Goemaere A1 - Lagrou, Katrien A1 - Spriet, Isabel A1 - Marijke Hendrickx A1 - Eline Vandael A1 - Pierre Becker A1 - Boudewijn Catry KW - Antifungal Agents KW - Belgium KW - Drug Utilization KW - Female KW - Humans KW - intensive care units KW - Male KW - Mycoses AB -

BACKGROUND: Reports on the consumption of systemic antifungal drugs on a national level are scarce although of high interest to compare trends and the associated epidemiology in other countries and to assess the need for antifungal stewardship programmes.

OBJECTIVES: To estimate patterns of Belgian inpatient and outpatient antifungal use and provide reference data for other countries.

METHODS: Consumption records of antifungals were collected in Belgian hospitals between 2003 and 2016. Primary healthcare data were available for the azoles for the period 2010-2016.

RESULTS: The majority of the antifungal consumption resulted from prescriptions of fluconazole and itraconazole in the ambulatory care while hospitals were responsible for only 6.4% of the total national consumption and echinocandin use was limited. The annual average antifungal consumption in hospitals decreased significantly by nearly 25% between 2003 and 2016, due to a decrease solely in non-university hospitals. With the exception of specialised burn centres, antifungals are mostly consumed at ICUs and internal medicine wards. A significant decline was also observed in the consumption of azoles in primary health care, attributed to itraconazole. The major part of azoles was prescribed by generalists followed by dermatologists.

CONCLUSIONS: In spite of the downward trend in annual use of systemic antifungal drugs, Belgium remains one of the biggest consumers in Europe.

VL - 62 CP - 6 M3 - 10.1111/myc.12912 ER - TY - JOUR T1 - Black aspergilli: A remaining challenge in fungal taxonomy? JF - Med Mycol Y1 - 2018 A1 - Elizabet D'hooge A1 - Pierre Becker A1 - Dirk Stubbe A1 - Normand, Anne-Cécile A1 - Piarroux, Renaud A1 - Marijke Hendrickx KW - antifungal susceptibility KW - Aspergillus sect. Nigri KW - MALDI-TOF MS KW - Phylogeny AB -

Aspergillus section Nigri is a taxonomically difficult but medically and economically important group. In this study, an update of the taxonomy of A. section Nigri strains within the BCCM/IHEM collection has been conducted. The identification accuracy of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) was tested and the antifungal susceptibilities of clinical isolates were evaluated. A total of 175 strains were molecularly analyzed. Three regions were amplified (ITS, benA, and caM) and a multi-locus phylogeny of the combined loci was created by using maximum likelihood analysis. The in-house MALDI-TOF MS reference database was extended and an identification data set of 135 strains was run against a reference data set. Antifungal susceptibility was tested for voriconazole, itraconazole, and amphotericin B, using the EUCAST method. Phylogenetic analysis revealed 18 species in our data set. MALDI-TOF MS was able to distinguish between A. brasiliensis, A. brunneoviolaceus, A. neoniger, A. niger, A. tubingensis, and A. welwitschiae of A. sect. Nigri. In the routine clinical lab, isolates of A. sect. Nigri are often identified as A. niger. However, in the clinical isolates of our data set, A. tubingensis (n = 35) and A. welwitschiae (n = 34) are more common than A. niger (n = 9). Decreased antifungal susceptibility to azoles was observed in clinical isolates of the /tubingensis clade. This emphasizes the importance of identification up to species level or at least up to clade level in the clinical lab. Our results indicate that MALDI-TOF MS can be a powerful tool to replace classical morphology.

M3 - 10.1093/mmy/myy124 ER - TY - JOUR T1 - Clonal Spread of Candida glabrata Bloodstream Isolates and Fluconazole Resistance Affected by Prolonged Exposure: a 12-Year Single-Center Study in Belgium. JF - Antimicrob Agents Chemother Y1 - 2018 A1 - Berdieke Goemaere A1 - Lagrou, Katrien A1 - Spriet, Isabel A1 - Marijke Hendrickx A1 - Pierre Becker KW - antifungal resistance KW - Candida glabrata KW - Gene Expression KW - genotypic identification AB -

is a major cause of candidemia in immunocompromised patients and is characterized by a high-level of fluconazole resistance. In the present study, the acquisition of antifungal resistance and potential clonal spread of were explored at a single center over a 12-year period by analyzing 187 independent clinical bloodstream isolates. One strain was found to be micafungin resistant due to a mutation in the gene. Fluconazole resistance remained stable throughout the period and was observed in 20 (10.7%) of the isolates. An analysis of the antifungal consumption data revealed that recent prior exposure to fluconazole increased the risk to be infected by a resistant strain. In particular, the duration of the treatment was significantly longer for patients infected by a resistant isolate, while the total and mean daily doses received did not impact the acquisition of resistance in No link between genotype and resistance was found. However, multilocus variable-number tandem-repeat analyses indicated a potential intrahospital spread of some isolates between patients. These isolates shared the same genetic profiles, and infected patients were hospitalized in the same unit during an overlapping period. Finally, quantitative real-time PCR analyses showed that, unlike that for other ABC efflux pumps, the expression of CgCDR1 was significantly greater in resistant strains, suggesting that it would be more involved in fluconazole (FLC) resistance. Our study provides additional evidence that the proper administration of fluconazole is required to limit resistance and that strict hand hygiene is necessary to avoid the possible spreading of isolates between patients.

VL - 62 CP - 8 M3 - 10.1128/AAC.00591-18 ER - TY - JOUR T1 - Decision criteria for MALDI-TOF MS-based identification of filamentous fungi using commercial and in-house reference databases JF - BMC Microbiology Y1 - 2017 A1 - A-C Normand A1 - C Cassagne A1 - M. Gautier A1 - Pierre Becker A1 - S. Ranque A1 - Marijke Hendrickx A1 - R. Piarroux KW - Database KW - Fungi KW - MALDI-TOF MS KW - mould AB -

BACKGROUND: Several Matrix-Assisted Laser Desorption/Ionization Time-of-Flight mass spectrometry protocols, which differ in identification criteria, have been developed for mold and dermatophyte identification. Currently, the most widely used approach is Bruker technology, although no consensus concerning the log(score) threshold has been established. Furthermore, it remains unknown how far increasing the number of spots to compare results might improve identification performance. In this study, we used in-house and Bruker reference databases as well as a panel of 422 isolates belonging to 126 species to test various thresholds. Ten distinct identification algorithms requiring one to four spots were tested.

RESULTS: Our findings indicate that optimal results were obtained by applying a decisional algorithm in which only the highest score of four spots was taken into account with a 1.7 log(score) threshold. Testing the entire panel enabled identification of 87.41% (in-house database) and 35.15% (Bruker database) of isolates, with a positive predictive value (PPV) of 1 at the genus level for both databases as well as 0.89 PPV (in-house database) and 0.72 PPV (Bruker database) at the species level. Applying the same rules to the isolates for which the species were represented by at least three strains in the database enabled identification of 92.1% (in-house database) and 46.6% (Bruker database) of isolates, with 1 PPV at the genus level for both databases as well as 0.95 PPV (in-house database) and 0.93 PPV (Bruker database) at the species level.

CONCLUSIONS: Depositing four spots per extract and lowering the threshold to 1.7, a threshold which is notably lower than that recommended for bacterial identification, decreased the number of unidentified specimens without altering the reliability of the accepted results. Nevertheless, regardless of the criteria used for mold and dermatophyte identification, commercial databases require optimization.

VL - 17 CP - 1 M3 - 10.1186/s12866-017-0937-2 ER - TY - JOUR T1 - Increasing candidemia incidence from 2004 to 2015 with a shift in epidemiology in patients pre-exposed to antifungals. JF - Mycoses Y1 - 2017 A1 - Berdieke Goemaere A1 - Pierre Becker A1 - Van Wijngaerden, Eric A1 - Maertens, Johan A1 - Spriet, Isabel A1 - Marijke Hendrickx A1 - Lagrou, Katrien KW - Antifungal Agents KW - Candida spp. KW - candidaemia KW - epidemiology KW - incidence AB -

BACKGROUND: Candidemia is an important health problem in immunocompromised patients with an epidemiology varying with region, period and patient population involved.

OBJECTIVES: The occurrence of candidemia and the associated species distribution over a 12-year period at a large tertiary care centre in Belgium were analysed. The trend in incidence in the intensive care units (ICUs) and non-ICUs was investigated as well as the influence of antifungal exposure on the species distribution.

PATIENTS/METHODS: Candidemia incidence was expressed per 10,000 patient days (PD). Epidemiological parameters and antifungal consumption data were extracted from the hospital database and delivered by the hospital pharmacy, respectively.

RESULTS: From 2004 until 2015, 865 candidemia episodes occurred in 826 patients at the University Hospitals Leuven. Candida albicans (59%) remained the most important cause of candidemia, followed by C. glabrata (22.4%) and C. parapsilosis (8%). The mean incidence in the whole hospital was 1.48 per 10,000 PD. The incidence in ICUs increased reaching up to 10.7 per 10,000 PD whereas in the non-ICUs, the incidence decreased. Prior exposure to fluconazole and echinocandins was associated with candidemia caused by less susceptible species.

CONCLUSIONS: Candidemia incidence increased in the whole hospital, driven by ICUs. Surveillance of candidemia epidemiology on a local scale is of high value to guide empirical treatment strategies. This article is protected by copyright. All rights reserved.

U1 - http://www.ncbi.nlm.nih.gov/pubmed/29024057?dopt=Abstract M3 - 10.1111/myc.12714 ER - TY - JOUR T1 - Validation of a New Web Application for Identification of Fungi by Use of Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry JF - Journal of Clinical Microbiology Y1 - 2017 A1 - A. C. Normand A1 - Pierre Becker A1 - F. Gabriel A1 - C. Cassagne A1 - I. Accoceberry A1 - M. Gari-Toussaint A1 - L. Hasseine A1 - D. De Geyter A1 - D. Pierard A1 - I. Surmont A1 - F. Djenad A1 - J.L. Donnadieu A1 - M. Piarroux A1 - S. Ranque A1 - Marijke Hendrickx A1 - R. Piarroux ED - David W. Warnock VL - 55 CP - 9 M3 - 10.1128/JCM.00263-17 ER - TY - Generic T1 - Screening cyanobacteria to discover new antifungal compounds Y1 - 2016 A1 - Pierre Becker A1 - Wilmotte,A. A1 - Philippe Szternfeld A1 - Mirjana Andjelkovic A1 - R. Anthonissen A1 - Luc Verschaeve A1 - Marijke Hendrickx ED - IPP KW - Activity KW - an KW - Analyses KW - analysi KW - analysis KW - Antibiotic KW - AS KW - DRUG KW - drugs KW - effect KW - effects KW - evidence KW - growth KW - identify KW - incidence KW - Increase KW - Institute KW - IS KW - metabolites KW - methanol extract KW - Methanol extracts KW - Mycology KW - ON KW - Order KW - protein KW - resistance KW - SCREENING KW - Search KW - Secondary KW - Test KW - variety KW - work AB - Antifungal resistance is a growing concern in clinical settings with an increase in the incidence of multidrug resistant strains. The constant search for new antifungal compounds is therefore necessary. Cyanobacteria are known to produce a variety of secondary metabolites, out of which some appear to have antibiotic, anticancer or antifungal effects. In this work, a screening of cyanobacteria from the Antarctica was performed in order to discover potential new antifungal drugs. The analysis of a first set of methanol extracts from 15 different strains put in evidence the antifungal activity of one of them. Indeed, the extract of one JF - Departemental Retreat of the Mycology Department of the Pasteur Institute 2016 CY - Paris CP - Département de Mycologie de l'Institut Pasteur de Paris U1 - 37164 U2 - 29/06/2016 - 30/06/2016 ER - TY - Generic T1 - Identifying fungal strains by MALDI-TOF MS: examples of applications Y1 - 2015 A1 - Pierre Becker A1 - Ann Packeu A1 - Martiny,D. A1 - A. De Bel A1 - Ranque,S. A1 - Piarroux,R. A1 - Marijke Hendrickx KW - application KW - applications KW - MALDI-TOF KW - MALDI-TOF MS KW - medical KW - Medical mycology KW - MS KW - Mycology KW - strain KW - trend KW - trends JF - 7th Trends in Medical Mycology PB - NA CY - NA CP - EORTC, ESCMID U1 - 38047 U2 - 9-12/10/2015 ER - TY - Generic T1 - A modern tool for a conservative purpose: the use of MALDI-TOF MS in a fungal culture collection Y1 - 2015 A1 - Pierre Becker A1 - Dirk Stubbe A1 - Claessens,J. A1 - Roesems,S. A1 - Bastin,Y A1 - Chantal Planard A1 - C Cassagne A1 - Piarroux,R. A1 - Marijke Hendrickx ED - d'Enfert,C KW - a KW - accuracy KW - alternative KW - an KW - Analyses KW - AS KW - at KW - community KW - Control KW - conventional KW - Cost effectiveness KW - Cost-effectiveness KW - cryopreservation KW - culture KW - Dna KW - expertise KW - filamentous fungi KW - Fungi KW - Genetic KW - identification KW - IS KW - LEVEL KW - Long-term KW - MALDI-TOF KW - MALDI-TOF MS KW - Mass KW - Mass Spectrometry KW - method KW - Microscopy KW - MS KW - Mycology KW - objectives KW - ON KW - performance KW - preservation KW - result KW - results KW - routine KW - Species KW - strain KW - time KW - time of flight KW - use KW - viability AB - One of the main objectives of the BCCM/IHEM fungal culture collection is the long-term preservation of interesting strains to ensure their availability for the scientific community at large. The viability, purity and identity of a strain is verified before entering the collection, but also after lyophilisation or cryopreservation. For filamentous fungi, these routine identity checks are traditionally done based on morphological analyses, and DNA sequencing if necessary. This requires extensive taxonomical expertise, and is time consuming and costly. The BCCM/IHEM collection evaluated matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) as an alternative method for the routine post-preservation identity controls. A total of 481 controls were carried out using MALDI-TOF MS in parallel with the conventional procedure. The overall performance of the MALDI-TOF MS reached 84% of correct identifications at species level. Moreover, misidentification at entry in the collection could be corrected for 14 strains by mass spectrometry, which was confirmed by DNA sequencing. Out of these 14, only eight had been detected by the traditional method. Considering these results, a workflow combining MALDI-TOF MS, microscopy and genetic analyses is proposed to enhance accuracy and time- and cost-effectiveness of routine identity controls in fungal culture collections. JF - First departemental retreat Department of Mycology of the Institut Pasteur CP - d'Enfert,C U1 - 39131 U2 - 29-30/06/2015 ER - TY - Generic T1 - New developments in the BCCM/IHEM fungal strains collection Y1 - 2015 A1 - Pierre Becker A1 - Dirk Stubbe A1 - Berdieke Goemaere A1 - David Triest A1 - Marijke Hendrickx KW - Development KW - International KW - Network KW - strain KW - symposium JF - International Scientific Symposium of the Institut Pasteur International Network PB - NA CY - NA CP - Institut Pasteur U1 - 39132 U2 - 14-16/102015 ER - TY - JOUR T1 - Quality control in culture collections: confirming identity of filamentous fungi by MALDI-TOF MS JF - Mycoscience Y1 - 2015 A1 - Pierre Becker A1 - Dirk Stubbe A1 - Claessens,J. A1 - Roesems,S. A1 - Bastin,Y. A1 - Chantal Planard A1 - C Cassagne A1 - Piarroux,R. A1 - Marijke Hendrickx KW - a KW - alternative KW - an KW - Analyses KW - AS KW - at KW - Control KW - conventional KW - culture KW - Dna KW - evidence KW - filamentous fungi KW - Fungi KW - Genetic KW - identification KW - IS KW - KNOWLEDGE KW - LEVEL KW - MALDI-TOF KW - MALDI-TOF MS KW - Mass KW - Mass Spectrometry KW - method KW - methods KW - Microscopy KW - MS KW - Mycology KW - Objective KW - performance KW - preservation KW - Quality KW - Quality Control KW - result KW - results KW - routine KW - Species KW - strain KW - study KW - time of flight KW - viability KW - work AB -

In culture collections, strains are controlled after preservation to guarantee their viability, purity and identity. For filamentous fungi, the identity is traditionally verified by performing morphological analyses with the support of DNA sequencing if required. These methods are particularly time-consuming and require extensive knowledge of mycology. In this study, matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) was evaluated as an alternative method for fast, robust and objective identity controls in the routine work of the BCCM/IHEM fungal collection. A total of 481 controls were carried out using mass spectrometry and compared to the results obtained by the conventional procedure. The overall performance of the MALDI-TOF MS reached 84% of correct identification at species level. Moreover, misidentification at entry in the collection was put in evidence for 14 strains by mass spectrometry and confirmed by DNA sequencing. Out of these, only eight were detected by the traditional method. Considering these results, a workflow combining MALDI-TOF MS, microscopy and genetic analyses is proposed to speed up and objectify identity controls in fungal culture collections.

VL - 56 U1 - 39130 M3 - http://dx.doi.org/10.1016/j.myc.2014.08.002 ER - TY - Generic T1 - Trends in candidaemia and antifungal consumption in Belgium Y1 - 2015 A1 - Berdieke Goemaere A1 - Pierre Becker A1 - A Ingenbleek A1 - Boudewijn Catry A1 - Marijke Hendrickx A1 - K. Lagrou KW - Belgium KW - CONSUMPTION KW - medical KW - Medical mycology KW - Mycology KW - trend KW - trends JF - 7th Trends in Medical Mycology PB - NA CY - NA CP - EORTC, ECMM U1 - 38209 U2 - 09-12/10/2015 ER - TY - JOUR T1 - Identification of filamentous fungi isolates by MALDI-TOF mass spectrometry: clinical evaluation of an extended reference spectra library. JF - Med Mycol Y1 - 2014 A1 - Pierre Becker A1 - de Bel, Annelies A1 - Martiny, Delphine A1 - Ranque, Stéphane A1 - Piarroux, Renaud A1 - C Cassagne A1 - Detandt, Monique A1 - Marijke Hendrickx KW - Databases, Factual KW - Fungi KW - Humans KW - Mycological Typing Techniques KW - Mycoses KW - Reproducibility of Results KW - Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization AB -

The identification of filamentous fungi by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) relies mainly on a robust and extensive database of reference spectra. To this end, a large in-house library containing 760 strains and representing 472 species was built and evaluated on 390 clinical isolates by comparing MALDI-TOF MS with the classical identification method based on morphological observations. The use of MALDI-TOF MS resulted in the correct identification of 95.4% of the isolates at species level, without considering LogScore values. Taking into account the Brukers' cutoff value for reliability (LogScore >1.70), 85.6% of the isolates were correctly identified. For a number of isolates, microscopic identification was limited to the genus, resulting in only 61.5% of the isolates correctly identified at species level while the correctness reached 94.6% at genus level. Using this extended in-house database, MALDI-TOF MS thus appears superior to morphology in order to obtain a robust and accurate identification of filamentous fungi. A continuous extension of the library is however necessary to further improve its reliability. Indeed, 15 isolates were still not represented while an additional three isolates were not recognized, probably because of a lack of intraspecific variability of the corresponding species in the database.

VL - 52 CP - 8 U1 - http://www.ncbi.nlm.nih.gov/pubmed/25349253?dopt=Abstract M3 - 10.1093/mmy/myu064 ER - TY - JOUR T1 - MALDI-TOF mass spectrometry: revolutionising clinical laboratory diagnosis of mould infections JF - Clin.Microbiol.Infect. Y1 - 2014 A1 - Gautier,M. A1 - Ranque,S. A1 - A-C Normand A1 - Pierre Becker A1 - Ann Packeu A1 - C Cassagne A1 - L'ollivier,C. A1 - Marijke Hendrickx A1 - Piarroux,R. KW - a KW - accuracy KW - ALL KW - an KW - article KW - AS KW - at KW - challenge KW - Clinical KW - criteria KW - data KW - de KW - Diagnosis KW - diversity KW - Dna KW - electronic KW - France KW - identification KW - implementation KW - Improvement KW - Increase KW - INFECTION KW - infections KW - IS KW - journal KW - Laboratories KW - LEVEL KW - MALDI-TOF KW - MALDI-TOF MS KW - Mass KW - Mass Spectrometry KW - Morphology KW - MS KW - Mycology KW - ON KW - parasitology KW - Patient KW - period KW - Prognosis KW - Reduction KW - result KW - results KW - routine KW - Species KW - strain KW - Technique AB -

Clinical diagnosis of mould infections currently involves complex species identification based on morphological criteria, which is often prone to error. Employing an extensive mould species reference spectral library (up to 2,832 reference spectra, corresponding to 708 strains from 347 species), we assessed the extent to which MALDI-TOF mass spectrometry (MALDI-TOF MS) enhanced the accuracy of species identification. MALDI-TOF MS data were validated against morphology- and DNA sequencing-based results using 262 clinical isolates collected over a four-month period in 2013. The implementation of MALDI-TOF MS displayed a dramatic improvement in mould identification at the species level (increased from 78.2% to 98.1%) and a marked reduction in misidentification rate (i.e., 9.8% to 1.2%). We then compared the mould identification results obtained before (i.e., 2011) and after (i.e., 2013) the implementation of MALDI-TOF MS in routine identification procedures, which increased from 64.57% to 100%. Re-assessment of a set of isolates from 2011 using this procedure, including MALDI-TOF MS, yielded an increase in species diversity from 16 to 42 species. Finally, applying this procedure during a 16-month period (2012-2013) enabled the identification of 1,094/1,107 (98.8%) clinical mould isolates corresponding to 107 distinct species. MALDI-TOF MS-based mould species identification may soon challenge traditional techniques in the clinical laboratory, as patient prognosis is largely contingent on rapid and accurate diagnosis. This article is protected by copyright. All rights reserved

VL - 20 CP - 12 U1 - 39167 M3 - 10.1111/1469-0691.12750 [doi] ER -