Effect of Inoculum Size and Culture Age on the Cellular Properties and Host-Pathogen Interactions of Cryptococcus neoformans

Abstract

Aims: Cryptococcus neoformans is a fungal pathogen which infection caused devastating morbidity and mortality in immunocompromised patients. The present study investigated the effect of culture starting inoculum size and culture age towards cellular properties of C. neoformans and its interactions with mammalian host alveolar epithelial cells. Methodology: C. neoformans H99 was cultured at different starting inoculum sizes and collected at varied culture ages to examine the morphology of the yeast cells and agar invasion property. The interaction with host alveolar epithelial cells was assessed in vitro using A549 cells as the host cell model. Results: Visual observation demonstrated that cryptococci cultured with higher starting inoculum sizes and longer incubation periods displayed flocculation properties, aberrant morphologies with lysed cell structure attached to intact yeast cells, release of capsule material to the culture medium, as well as changes in FITC staining of cell surface proteins. The changes in cryptococcal cellular morphology did not affect agar invasion, as the encapsulated cryptococci did not invade agar under all conditions tested. Lysed cell material on cryptococcal cells adhered to host alveolar epithelial cells, which induced localised actin reorganisation at the host-pathogen interface. Conclusion: Our results indicate that the differences in starting inoculum size and culture age of C. neoformans H99 resulted in yeast cells with distinctive morphologies, which affected he pathogen’s association with host alveolar epithelial cells.