Carbon Metabolism and iCryptococcus neoformans/i Virulence

Cryptococcus neoformans is an important fungal pathogen of immunocompromised individuals, with a close relative - C. gattii - emerging as a serious threat for the immunocompetent. During active infection, C. neoformans migrates to the brain and persists in the cerebrospinal fluid (CSF). Since CSF is a nutrient-limited environment, we sought to understand fungal carbon utilization in CSF. Prior studies have established the role of phosphoenolpyruvate carboxykinase ( PCK1 , links 3-carbon utilization with gluconeogenesis) in virulence using a inhalational mouse model. We evaluated a pck1delta mutant using a rabbit CSF model of virulence and found no defect in virulence for this mutant. Isocitrate lyase ( ICL1 ), a key enzyme in the glyoxylate cycle (2-carbon utilization), was also previously examined by gene deletion and shown to have no impact on virulence in either the mouse or rabbit models of cryptococcal disease. In order to evaluate the use of various carbon sources by C. neoformans in the CSF environment, we created a pck1delta/icl1delta double mutant to evaluate the impairment of both 2- and 3- carbon utilization in a rabbit model of cryptococcosis. Furthermore, a pyruvate kinase ( PYK1 ) mutant was made to examine the importance of glycolysis on growth and virulence in vivo. These studies serve to determine the relative roles of different carbon metabolism pathways in the persistence of C. neoformans in the CSF environment of the host.