Department of Microbiology

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Browsing Department of Microbiology by browse.metadata.advisor "Boekhout, Teun"

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    Molecular characterisation of Cryptococcus neoformans and Cryptococcus gattii present in tree hollows, their antifungal susceptibility, and chemical factors influencing their growth on woody debris
    (Stellenbosch : Stellenbosch University, 2020-03) Vreulink, Jo-Marie; Botha, Alfred; Vismer, Hester; Boekhout, Teun; Stellenbosch University. Faculty of Science. Dept. of Microbiology.
    ENGLISH ABSTRACT: Members of the Cryptococcus neoformans/Cryptococcus gattii species complex causes cryptococcosis; a disease that is a major burden among HIV/AIDS patients in South Africa. Infection is likely obtained via inhalation of basidiospores from the environment. Basidiospores are produced during opposite-sex mating that occurs between MATα and MATa strains or through same-sex mating. Although it is well established that the habitats of pathogenic cryptococci can serve as infection sources, little was known about the prevalence of environmental pathogenic cryptococci and their relation to clinical isolates in South Africa. It was suggested that environmental factors could affect mating type occurrence, yet nothing was known about this aspect. Furthermore, it was unestablished if South African environmental isolates presented different antifungal susceptibilities than clinical isolates. To address these facets, pathogenic cryptococci were first isolated from the South African environment. Following molecular characterisation, the epidemiological relationship between the environmental and clinical isolates was investigated. Subsequently, the effect of chemical factors on the growth of strains found to colonise trees differentially was examined. Lastly, the antifungal susceptibility of the environmental isolates was compared to that of clinical isolates. Pathogenic cryptococci were isolated from woody debris samples collected on three sampling dates from trees situated in a public park in Cape Town (PPCT) and on one sampling date from trees located on a privately-owned game farm in the Northern Cape (GFNC), whereafter the isolates were molecularly characterised. Since trees were not permanently colonised by C. gattii sensu stricto MATa, it was estimated whether chemical factors in the wood could have affected the occurrence of C. gattii s.s. MATa. A positive correlation was detected between the numbers of C. gattii s.s. MATa and the ammonium-nitrogen (NH₄-N) concentration. Subsequently, the effect of low and high ammonium chloride (NH₄Cl) concentrations on the growth of C. gattii s.s. MATa and MATα was investigated in woody microcosms. Antifungal susceptibility differences between environmental and clinical isolates were evaluated by determining the amphotericin B (AmB) and fluconazole (FLU) susceptibility of the isolates. The pathogenic cryptococci isolated from the PPCT trees were predominantly C. gattii s.s. ST230 followed by C. neoformans s.s. genotype AFLP1/VNI ST23. Since ST23 was previously isolated from infected patients, the PPCT trees might be an infection source. Growth studies revealed that C. gattii s.s. MATa isolates were able to compete better with MATα isolates in microcosms supplemented with a high NH₄Cl concentration, which provided a possible explanation for the absence of MATa in PPCT trees after the first sampling date. All of the environmental isolates were more susceptible to AmB than clinical isolates. In contrast, environmental C. neoformans s.s. genotype AFLP/VNI and C. gattii s.s. isolates were less susceptible to FLU than corresponding clinical isolates, indicating that FLU resistance can develop among environmental isolates. Furthermore, temporal variation in C. gattii s.s. ST230 isolates’ FLU susceptibility was potentially linked to variations in iron, NH₄-N and sodium concentrations within their original woody habitat. This study indicates that environmental factors can affect the numbers and antifungal susceptibility of pathogenic cryptococci in the South African environment.