Abstract
Whole genome sequencing of human fungal pathogens has revolutionized the speed and accuracy in which sequence variants that cause antifungal resistance can be identified. Genome rearrangements resulting in copy number variation (CNV) are a significant source of acquired antifungal drug resistance across diverse fungal species. Some CNVs are transient in nature, while other CNVs are stable and well tolerated even in the absence of antifungal drugs. By visualizing whole genome sequencing read depth as a function of genomic location, CNVs and CNV breakpoints (genomic positions where the copy number changes occur relative to the rest of the genome) are rapidly identified. A similar analysis can be used to visualize allele ratio changes that occur across the genomes of heterozygous fungal species, both in the presence and absence of CNVs. This protocol walks through the bioinformatic analysis of CNVs and allele ratios utilizing free, open-source visualization tools. We provide code to use with an example dataset (matched antifungal drug-sensitive and drug-resistant Candida albicans isolates) and notes on how to expand this protocol to other fungal genomes.
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Acknowledgements
We are grateful to Petra Vande Zande, Natthapon Soisangwan, and Taylor Crooks for providing helpful feedback on the manuscript and to Robin Dowell and her team for providing free and accessible short-read sequence alignment workshops. Funding was provided by the National Institute of Health (R01AI143689) and the Burroughs Wellcome Fund Investigators in the Pathogenesis of Infectious Diseases (no. 1020388) to A.S.
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Todd, R.T., Selmecki, A. (2023). Copy Number Variation and Allele Ratio Analysis in Candida albicans Using Whole Genome Sequencing Data. In: Krysan, D.J., Moye-Rowley, W.S. (eds) Antifungal Drug Resistance. Methods in Molecular Biology, vol 2658. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-3155-3_8
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DOI: https://doi.org/10.1007/978-1-0716-3155-3_8
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