One of the leading fungal pathogen of humans, Candida albicans, does not appear to engage in canonical sexual recombination. This is surprising given C. albicans’ proclivity to effectively switch between being a commensal and an opportunistic pathogen and rapidly evolve drug resistances. The molecular chaperone Hsp90, a central regulator of protein homeostasis, governs virulence and drug resistance in C. albicans. This then raises the question: how does an ‘obligate’ diploid generate genetic variations? Here, we address this question by identifying if and how Hsp90 regulates genome integrity in C. albicans. Forche et al. (2011) demonstrated that different stresses elevated Loss-of-Heterozygosity (LoH) rates across the C. albicans genome. We hypothesize that alterations of either Hsp90 level or function result in LoH and thus promote genome diversification. In line with this hypothesis, we conducted fluctuation assay showing that multiple modes of Hsp90 perturbation increase LoH rate, suggesting that Hsp90 is required for the maintenance of a diploid, heterozygous genome but compromised Hsp90 function promotes the genomic rearrangement. More importantly, our SNP-RFLP analyses revealed that the types of LoH events are contingent on the mechanisms used to abrogate Hsp90 function. Pharmacological inhibition increased frequency of local recombination events such as gene conversion or break-induced recombination, whereas high temperature stress caused whole-chromosome homozygosis. This very novel observation suggests that C. albicans may utilize distinct forms of LoH to deal with disparate stressors. This concept is supported by our in vitro fungal drug resistance test which showing a biological relevant pattern. Hence, our work provides evidence that Hsp90 governing LoH rates and types is one of principles underpinning C. albicans genome plasticity, which aids in the generation of mosaic karyotypes with novel phenotypes. This indicates that Hsp90 affects a pathogen’s virulence through genome modulations, eventually imparting both theoretical and clinic novel knowledge.