Characterization of Phenotypic Switching in Cryptococcus neoformans Biofilms

Abstract

Cryptococcus neoformans is an encapsulated yeast-like fungus that is a relatively frequent cause of meningoencephalitis in immunocompromised patients and also occasionally causes disease in apparently healthy individuals. This fungus collectively forms biofilms on polystyrene plates and medical devices, whereas individually can undergo phenotypic switching. Both events have profound consequences in the establishment of fungal infection and are associated with persistent infection due to increase resistance to antimicrobial therapy. In this study, we characterized switch phenotypes in C. neoformans biofilms. Smooth, mucoid, and wrinkled switch phenotypes of various switching C. neoformans strains were examined for their adhering and biofilm-forming ability on 96-well plates using cell counts and 2,3-bis (2-methoxy-4-nitro-5-sulfophenyl)-5-[(phenylamino) carbonyl]-2H-tetrazolium hydroxide (XTT) reduction assay, respectively. Both assays showed that C. neoformans strains with the parent smooth phenotype adhered and formed stronger biofilms than their mucoid and wrinkled counterparts. Furthermore, the phenotypic switching frequencies of the individual colony types grown in biofilms or as planktonic cells were investigated. For the parent smooth variant of most strains, we found enhanced phenotypic switching in cryptococcal biofilms when compared to switching rates of planktonic cells. In contrast, the back-switching rate of mucoid to smooth variant was significantly higher in planktonic cells of seven strains of C. neoformans strains. These results suggested that phenotypic switching can occur in cryptococcal biofilms and extend our understanding of the relationship of both phenomena.

Variability of Phenotypic Traits in Cryptococcus Varieties and Species and the Resulting Implications for Pathogenesis

“…Both mucoid switch variants (RC2-MC and NP1-MC) have altered cell walls and exhibit increased sensitivity to lysing enzyme. Cell charge and melanization are not affected by PS in the two switch systems; however, biofilm formation can differ in the switch variants [104]. …”

Central venous catheter-related bloodstream infection and Cryptococcus neoformans

“…Cryptococcus neoformans collectively forms biofilms on polystyrene plates and medical devices [6]. Martinez and Casadevall demonstrated that fungal biofilm formation is dependent on support surface characteristics and that growth in the biofilm state makes fungal cells less susceptible to potential environmental stresses, including antibiotics [7;8].…”

Fungal Biofilms: Relevance in the Setting of Human Disease

“…The capsule can be altered by phenotypic switching, which can also change the ability of C. neoformans to generate biofilms [94]. Conditions that mimic the host environment have been suggested to be permissive for biofilm formation [95].…”

“…Quorum sensing is also described for C. neoformans [103], but its impact on biofilm formation is still unknown. There is evidence that certain cell variants exhibit higher switching rates when grown as biofilm-associated cells rather than planktonic cells [94]. …”