title: Targeted gene deletion and in vivo analysis of putative virulence gene function in the pathogenic dermatophyte Arthroderma benhamiae.
abstract: Dermatophytes cause the majority of superficial mycoses in humans and animals. However, little is known about the pathogenicity of this specialized group of filamentous fungi, for which molecular research has been limited thus far. During experimental infection of guinea pigs by the human pathogenic dermatophyte Arthroderma benhamiae, we recently detected the activation of the fungal gene encoding malate synthase AcuE, a key enzyme of the glyoxylate cycle. By the establishment of the first genetic system for A. benhamiae, specific ΔacuE mutants were constructed in a wild-type strain and, in addition, in a derivative in which we inactivated the nonhomologous end-joining pathway by deletion of the A. benhamiae KU70 gene. The absence of AbenKU70 resulted in an increased frequency of the targeted insertion of linear DNA by homologous recombination, without notably altering the monitored in vitro growth abilities of the fungus or its virulence in a guinea pig infection model. Phenotypic analyses of ΔacuE mutants and complemented strains depicted that malate synthase is required for the growth of A. benhamiae on lipids, major constituents of the skin. However, mutant analysis did not reveal a pathogenic role of the A. benhamiae enzyme in guinea pig dermatophytosis or during epidermal invasion of the fungus in an in vitro model of reconstituted human epidermis. The presented efficient system for targeted genetic manipulation in A. benhamiae, paired with the analyzed infection models, will advance the functional characterization of putative virulence determinants in medically important dermatophytes.
authors.affiliation: Leibniz Institute for Natural Product Research and Infection Biology, Hans Knoell Institute, Junior Research Group Fundamental Molecular Biology of Pathogenic Fungi, Beutenbergstr. 11a, D-07745 Jena, Germany.
authors.forename: Maria
authors.initials: M
authors.lastname: Grumbt
authors.forename: Valérie
authors.initials: V
authors.lastname: Defaweux
authors.forename: Bernard
authors.initials: B
authors.lastname: Mignon
authors.forename: Michel
authors.initials: M
authors.lastname: Monod
authors.forename: Anke
authors.initials: A
authors.lastname: Burmester
authors.forename: Johannes
authors.initials: J
authors.lastname: Wöstemeyer
authors.forename: Peter
authors.initials: P
authors.lastname: Staib
mesh_terms: D000505:Alopecia / Q000382:microbiology; D000818:Animals; D003883:Arthrodermataceae / Q000201:enzymology / Q000235:genetics / Q000472:pathogenicity*; D003881:Dermatomycoses / Q000382:microbiology*; D004890:Erythema / Q000382:microbiology; D005260:Female; D005656:Fungal Proteins / Q000235:genetics* / Q000378:metabolism; D017353:Gene Deletion; D006168:Guinea Pigs; D006197:Hair / Q000382:microbiology; D018859:Hair Follicle / Q000382:microbiology / Q000473:pathology; D006801:Humans; D008292:Malate Synthase / Q000235:genetics / Q000378:metabolism; D008297:Male; D045522:Recombinases / Q000235:genetics* / Q000378:metabolism; D012867:Skin / Q000382:microbiology / Q000473:pathology; D017768:Skin, Artificial / Q000382:microbiology; D037521:Virulence Factors / Q000235:genetics*
chemical_list: D005656:Fungal Proteins; D045522:Recombinases; D037521:Virulence Factors; D008292:Malate Synthase