title: Identification and Characterization of VNI/VNII and Novel VNII/VNIV Hybrids and Impact of Hybridization on Virulence and Antifungal Susceptibility Within the C. neoformans/C. gattii Species Complex.
abstract: Cryptococcus neoformans and C. gattii are pathogenic basidiomycetous yeasts and the commonest cause of fungal infection of the central nervous system. Cryptococci are typically haploid but several inter-species, inter-varietal and intra-varietal hybrids have been reported. It has a bipolar mating system with sexual reproduction occurring normally between two individuals with opposite mating types, α and a. This study set out to characterize hybrid isolates within the C. neoformans/C. gattii species complex: seven unisexual mating intra-varietal VNI/VNII (αAAα) and six novel inter-varietal VNII/VNIV (aADα). The URA5-RFLP pattern for VNII/VNIV (aADα) differs from the VNIII (αADa) hybrids. Analysis of the allelic patterns of selected genes for AD hybrids showed 79% or more heterozygosis for the studied loci except for CBS132 (VNIII), which showed 50% of heterozygosity. MALDI-TOF MS was applied to hybrids belonging to different sero/mating type allelic patterns. All hybrid isolates were identified as belonging to the same hybrid group with identification scores ranging between 2.101 to 2.634. All hybrids were virulent when tested in the Galleria mellonella (wax moth) model, except for VNII/VNIV (aADα) hybrids. VNI/VGII hybrids were the most virulent hybrids. Hybrids recovered from larvae manifested a significant increase in capsule and total cell size and produced a low proportion (5-10%) of giant cells compared with the haploid control strains. All strains expressed the major virulence factors-capsule, melanin and phospholipase B-and grew well at 37°C. The minimal inhibitory concentration of nine drugs was measured by micro-broth dilution and compared with published data on haploid strains. MICs were similar amongst hybrids and haploid parental strains. This is the first study reporting natural same sex αAAα intra-varietal VNI/VNII hybrids and aADα inter-varietal VNII/VNIV hybrids.
authors.affiliation: Molecular Mycology Research Laboratory, Centre for Infectious Diseases and Microbiology, Sydney Medical School - Westmead Hospital, Marie Bashir Institute for Infectious Diseases and Biosecurity, The University of Sydney, Westmead Institute for Medical Research, Sydney, Australia.
authors.forename: Mojgan
authors.initials: M
authors.lastname: Aminnejad
authors.affiliation: Laboratory Micologia Medica, Dip. Scienze Biomediche per la Salute, Università degli Studi di Milano, Milano, Italy.
authors.forename: Massimo
authors.initials: M
authors.lastname: Cogliati
authors.affiliation: Molecular Mycology Research Laboratory, Centre for Infectious Diseases and Microbiology, Sydney Medical School - Westmead Hospital, Marie Bashir Institute for Infectious Diseases and Biosecurity, The University of Sydney, Westmead Institute for Medical Research, Sydney, Australia.
authors.forename: Shuyao
authors.initials: S
authors.lastname: Duan
authors.affiliation: Mycology Research Laboratory, Department of Microbiology, Medical School, National Kapodistrian University of Athens, Athens, Greece.
authors.forename: Michael
authors.initials: M
authors.lastname: Arabatzis
authors.affiliation: Mycology Group, Robert Koch Institute Berlin, Berlin, Germany.
authors.forename: Kathrin
authors.initials: K
authors.lastname: Tintelnot
authors.affiliation: Grupo de Microbiología, Instituto Nacional de Salud, Bogotá, Colombia.
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Castañeda
authors.affiliation: Mycology Laboratory, National Institute of Infectious Diseases, Oswaldo Cruz Foundation, Rio de Janeiro, Brazil.
authors.forename: Marcia
authors.initials: M
authors.lastname: Lazéra
authors.affiliation: Mycology Research Laboratory, Department of Microbiology, Medical School, National Kapodistrian University of Athens, Athens, Greece.
authors.forename: Aristea
authors.initials: A
authors.lastname: Velegraki
authors.affiliation: School of Molecular & Biomedical Sciences, University of Adelaide, Adelaide, SA 5005, Australia.
authors.forename: David
authors.initials: D
authors.lastname: Ellis
authors.affiliation: Molecular Mycology Research Laboratory, Centre for Infectious Diseases and Microbiology, Sydney Medical School - Westmead Hospital, Marie Bashir Institute for Infectious Diseases and Biosecurity, The University of Sydney, Westmead Institute for Medical Research, Sydney, Australia.
authors.forename: Tania C
authors.initials: TC
authors.lastname: Sorrell
authors.affiliation: Molecular Mycology Research Laboratory, Centre for Infectious Diseases and Microbiology, Sydney Medical School - Westmead Hospital, Marie Bashir Institute for Infectious Diseases and Biosecurity, The University of Sydney, Westmead Institute for Medical Research, Sydney, Australia.
authors.forename: Wieland
authors.initials: W
authors.lastname: Meyer
mesh_terms: D000935:Antifungal Agents / Q000494:pharmacology*; D056285:Cryptococcus gattii / Q000235:genetics / Q000502:physiology*; D003455:Cryptococcus neoformans / Q000235:genetics / Q000502:physiology*; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism; D049770:Genes, Mating Type, Fungal / Q000235:genetics; D006238:Haploidy; D006824:Hybridization, Genetic / Q000235:genetics*; D008245:Lysophospholipase / Q000378:metabolism; D008543:Melanins / Q000378:metabolism; D008826:Microbial Sensitivity Tests; D060885:Multiplex Polymerase Chain Reaction; D016533:Mycological Typing Techniques; D065288:Serogroup; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014774:Virulence / Q000187:drug effects* / Q000235:genetics; D037521:Virulence Factors / Q000032:analysis / Q000235:genetics / Q000378:metabolism
chemical_list: D000935:Antifungal Agents; C087781:CAP59 protein, Cryptococcus neoformans; D005656:Fungal Proteins; D008543:Melanins; D037521:Virulence Factors; D008245:Lysophospholipase