Systematic Analysis of Copy Number Variations in the Pathogenic Yeast Candida Parapsilosis Identifies a Gene Amplification in That is Associated with Drug Resistance

We analyzed the genomes of 170 C. parapsilosis isolates and identified multiple copy number variations (CNVs). We identified two genes, () and (), each of which was the target of multiple independent amplification events. Phylogenetic analysis shows that most of these amplifications originated only once. For , which encodes a putative arsenate transporter, 8 distinct CNVs were identified, ranging in size from 2.3 kb to 10.5 kb with 3 to 23 copies. For , 16 distinct CNVs were identified, ranging in size from 0.3 kb to 4.5 kb with 2 to ~50 copies. One unusual amplification resulted in a DUP-TRP/INV-DUP structure similar to some human CNVs. encodes a putative phosphatidylcholine (PC) floppase which is known to regulate the inward translocation of PC in Candida albicans. We found that an increased copy number of correlated with resistance to miltefosine, an alkylphosphocholine drug that affects PC metabolism. Additionally, we conducted an adaptive laboratory evolution experiment in which two C. parapsilosis isolates were cultured in increasing concentrations of miltefosine. Two genes, and , coding for putative PC flippases homologous to S. cerevisiae gained homozygous protein-disrupting mutations in the evolved strains. Overall, our results show that C. parapsilosis can gain resistance to miltefosine, a drug that has recently been granted orphan drug designation approval by the United States Food and Drug Administration for the treatment of invasive candidiasis, through both CNVs or loss-of-function alleles in one of the flippase genes. Copy number variations (CNVs) are an important source of genomic diversity that have been associated with drug resistance. We identify two unusual CNVs in the human fungal pathogen Candida parapsilosis. Both target a single gene ( or ), and they have occurred multiple times in multiple isolates. The copy number of , a putative floppase that controls the inward translocation of lipids in the cell membrane, correlates with resistance to miltefosine, a derivative of phosphatidylcholine (PC) that was originally developed as an anticancer drug. In 2021, miltefosine was designated an orphan drug by the United States Food and Drug Administration for the treatment of invasive candidiasis. Importantly, we find that resistance to miltefosine is also caused by mutations in flippases, which control the outward movement of lipids, and that many C. parapsilosis isolates are prone to easily acquiring an increased resistance to miltefosine.