Biology subjects

Houbraken, J.

Publications and source records attributed to Houbraken, J..

6 recordsLinked to original sources

bioRxiv2024

Investigating the genetic relationship of Trichosporon asahii isolates using a newly developed microsatellite typing assay

The basidiomycetous yeast-like fungus Trichosporon asahii is increasingly encountered in the clinic and has been known to cause nosocomial infections. Here, we aim to develop a microsatellite typing tool to investigate the genetic relatedness of Trichosporon asahii isolates. Six microsatellite markers were selected from a nanopore long-read sequencing de novo assembled genome of the T. asahii type-strain CBS 2479. PCR reactions amplified three di-, tri-, or tetranucleotide repeats, respectively. All six microsatellite markers were used to analyze 111 T. asahii isolates, including single and related-patient isolates from clinical and environmental sources. Each marker exhibited between 11 and 37 alleles in this population, resulting in 71 different microsatellite genotypes. The Simpsons diversity index ranged from 0.6452 to 0.8280 for the individual markers, and for the combined set of microsatellites 0.9793, indicating the high discriminatory power of the reported panel. In summary, this panel combines high reproducibility and specificity, making it suitable for use in epidemiological studies of T. asahii and in investigations of potential outbreaks caused by this pathogen.

molecular biology↗

bioRxiv2024

Aspergillus species in Iceland: species distribution and cyp51A mutation testing in a 7.5-year clinical isolate collection

Introduction.Among challenges in diagnosing and treating aspergillosis are Aspergillus species identification, which can inform therapeutic choices, and detection of antifungal resistance. Increasing rates of triazole resistance have been reported for the most common species, Aspergillus fumigatus. Methods.A collection of clinical Aspergillus isolates cultured during 2014-2021, previously identified morphologically, were tested with the AsperGenius qPCR-kit for identification of A. fumigatus and Aspergillus flavus and the presence of a cyp51A mutation encoding azole resistance (TR34/L98H and Y121F/T289A); other species were identified with calmodulin sequencing. Results.Identification of the 312 isolates (224 patients) tested revealed 19 species in nine Aspergillus sections. Rare or cryptic species were identified in 48 isolates (44 patients). Thirty-one isolates had been assigned an incorrect species name when initially cultured. The TR34/L98H and Y121F/T289A mutations were not found in any of the 202 A. fumigatus isolates tested. Discussion.In this first study on clinical Aspergillus isolates in Iceland the proportion of cryptic and rare species was on par with findings from other countries. In contrast, unlike other studies we did not find the most common azole resistance-related mutations in A. fumigatus, possibly due to the absence of significant environmental antifungal pressure in the local agriculture.

microbiology↗

bioRxiv2024

Strain heterogeneity in a non-pathogenic fungus highlights factors contributing to virulence

Fungal pathogens exhibit extensive strain heterogeneity, including variation in virulence. Whether closely related non-pathogenic species also exhibit strain heterogeneity remains unknown. Here, we comprehensively characterized the pathogenic potentials (i.e., the ability to cause morbidity and mortality) of 16 diverse strains of Aspergillus fischeri, a non-pathogenic close relative of the major pathogen Aspergillus fumigatus. In vitro immune response assays and in vivo virulence assays using a mouse model of pulmonary aspergillosis showed that A. fischeri strains varied widely in their pathogenic potential. Furthermore, pangenome analyses suggest that A. fischeri genomic and phenotypic diversity is even greater. Genomic, transcriptomic, and metabolomic profiling identified several pathways and secondary metabolites associated with variation in virulence. Notably, strain virulence was associated with the simultaneous presence of the secondary metabolites hexadehydroastechrome and gliotoxin. We submit that examining the pathogenic potentials of non-pathogenic close relatives is key for understanding the origins of fungal pathogenicity.

evolutionary biology↗

bioRxiv2022

Footprints of domestication in dry-cured meat Penicillium fungi: convergent specific phenotypes and horizontal gene transfers

Some fungi have been domesticated for food production, with genetic differentiation between populations from food and wild environments, and food populations often acquiring beneficial traits through horizontal gene transfers (HGTs). Studying their adaptation to human-made substrates are of fundamental and applied importance, for understanding adaptation processes and for further strain improvement. We studied here the population structures and phenotypes of two distantly related Penicillium species used for dry-cured meat production, P. nalgiovense, the most common species in the dry-cured meat food industry, and P. salamii, used locally by farms. Both species displayed low genetic diversity, lacking differentiation between strains isolated from dry-cured meat and those from other environments. Nevertheless, the strains collected from dry-cured meat within each species displayed slower proteolysis and lipolysis than their wild conspecifics, and those of P. nalgiovense were whiter. Phenotypically, the non-dry-cured meat strains were more similar to their sister species than to their conspecific dry-cured meat strains, indicating an evolution of specific phenotypes in dry-cured meat strains. A comparison of available Penicillium genomes from various environments revealed HGTs, particularly between P. nalgiovense and P. salamii (representing almost 1.5 Mb of cumulative length). HGTs additionally involved P. biforme, also found in dry-cured meat products. We further detected positive selection based on amino-acid changes. Our findings suggest that selection by humans has shaped the P. salamii and P. nalgiovense populations used for dry-cured meat production, which constitutes domestication. Several genetic and phenotypic changes were similar in P. salamii, P. nalgiovense, and P. biforme, indicating convergent adaptation to the same human-made environment. Our findings have implications for fundamental knowledge on adaptation and for the food industry: the discovery of different phenotypes and of two mating types paves the way for strain improvement by conventional breeding, to elucidate the genomic bases of beneficial phenotypes and to generate diversity.

evolutionary biology↗

bioRxiv2022

High sorbic acid resistance of Penicillium roqueforti is mediated by the SORBUS gene cluster

Penicillium roqueforti is a major food-spoilage fungus known for its high resistance to the food preservative sorbic acid. Here, we demonstrate that the minimum inhibitory concentration of undissociated sorbic acid (MICu) ranges between 4.2 and 21.2 mM when 34 P. roqueforti strains were grown on malt extract broth. A genome-wide association study revealed that the six most resistant strains contained the 180 kbp gene cluster SORBUS, which was absent in the other 28 strains. In addition, a SNP analysis revealed five genes outside the SORBUS cluster that may be linked to sorbic acid resistance. A partial SORBUS knock-out (>100 of 180 kbp) in a resistant strain reduced sorbic acid resistance to similar levels as observed in the sensitive strains. Whole genome transcriptome analysis revealed a small set of genes present in both resistant and sensitive P. roqueforti strains that were differentially expressed in the presence of the weak acid. These genes could explain why P. roqueforti is more resistant to sorbic acid when compared to other fungi, even in the absence of the SORBUS cluster. Together, the MICu of 21.2 mM makes P. roqueforti among the most sorbic acid-resistant fungi, if not the most resistant fungus, which is mediated by the SORBUS gene cluster. Author summaryChemical preservatives, such as sorbic acid, are often used in food to prevent spoilage by fungi, yet some fungi are particularly well-suited to deal with these preservatives. First, we investigated the resistance of 34 Penicillium roqueforti strains to various food preservatives. This revealed that some strains were highly resistant to sorbic acid, while others are more sensitive. Next, we used DNA sequencing to compare the genetic variation between these strains and discovered a specific genetic region (SORBUS) that is unique to the resistant strains. Through comparative analysis with other fungal species the SORBUS region was studied in more detail and with the use of genetic engineering tools we removed this unique region. Finally, the mutant lacking the SORBUS region was confirmed to have lost its sorbic acid resistance. This finding is of particular interest as it suggests that only some, not all, P. roqueforti strains are potent spoilers and that specific genetic markers could help in the identification of resistant strains.

genomics↗

bioRxiv2020

Biosynthetic gene clusters, secondary metabolite profiles, and cards of virulence in the closest nonpathogenic relatives of Aspergillus fumigatus

Aspergillus fumigatus is a major human pathogen that causes hundreds of thousands of infections yearly with high mortality rates. In contrast, Aspergillus fischeri and the recently described Aspergillus oerlinghausenensis, the two species most closely related to A. fumigatus, are not known to be pathogenic. Some of the "cards of virulence" that A. fumigatus possesses are secondary metabolites that impair the host immune system, protect from host immune cell attacks, or acquire key nutrients. Secondary metabolites and the biosynthetic gene clusters (BGCs) that typically encode them often vary within and between fungal species. To gain insight into whether secondary metabolism-associated cards of virulence vary between A. fumigatus, A. oerlinghausenensis, and A. fischeri, we conducted extensive genomic and secondary metabolite profiling analyses. By analyzing multiple A. fumigatus, one A. oerlinghausenensis, and multiple A. fischeri strains, we identified both conserved and diverged secondary metabolism-associated cards of virulence. For example, we found that all species and strains examined biosynthesized the major virulence factor gliotoxin, consistent with the conservation of the gliotoxin BGC across genomes. However, species differed in their biosynthesis of fumagillin and pseurotin, both contributors to host tissue damage during invasive aspergillosis; these differences were reflected in sequence divergence of the intertwined fumagillin/pseurotin BGCs across genomes. These results delineate the similarities and differences in secondary metabolism-associated cards of virulence between a major fungal pathogen and its nonpathogenic closest relatives, shedding light into the genetic and phenotypic changes associated with the evolution of fungal pathogenicity. ImportanceThe major fungal pathogen Aspergillus fumigatus kills tens of thousands each year. In contrast, the two closest relatives of A. fumigatus, namely Aspergillus fischeri and Aspergillus oerlinghausenensis, are not considered pathogenic. A. fumigatus virulence stems, partly, from its ability to produce small molecules called secondary metabolites that have potent activities during infection. In this study, we examined whether A. fumigatus secondary metabolites and the metabolic pathways involved in their production are conserved in A. oerlinghausenensis and A. fischeri. We found that the nonpathogenic close relatives of A. fumigatus produce some, but not all, secondary metabolites thought to contribute to the success of A. fumigatus in causing human disease and that these similarities and differences were reflected in the underlying metabolic pathways involved in their biosynthesis. Compared to its nonpathogenic close relatives, A. fumigatus produces a distinct cocktail of secondary metabolites, which likely contributes to these organisms vastly different potentials to cause human disease. More broadly, the study of nonpathogenic organisms that have virulence-related traits, but are not currently considered agents of human disease, may facilitate the prediction of species capable of posing future threats to human health.

evolutionary biology↗