Exploring the evolution and adaptive role of mosaic aneuploidy in a clonal Leishmania donovani population using high throughput single cell genome sequencing

Maintenance of stable ploidy over continuous mitotic events is a paradigm for most higher eukaryotes. Defects in chromosome segregation and/or replication can lead to aneuploidy, a condition often considered deleterious. However, in , a Protozoan parasite, aneuploidy is a constitutive feature, where variations of somies represent a mechanism of gene expression adaptation, possibly impacting phenotypes. Strikingly, clonal populations display cell-to-cell somy variation, a phenomenon named mosaic aneuploidy (MA). However, until recently, no method was available for the determination of the complete karyotype of single parasites. To overcome this limitation, we used here for the first time a high-throughput single-cell genomic sequencing (SCGS) method to estimate individual karyotypes of 1560 promastigote cells in a clonal population of . We identified 128 different karyotypes, of which 4 were dominant. A network analysis revealed that most karyotypes are linked to each other by changes in copy number of a single chromosome and allowed us to propose a hypothesis of MA evolution. Moreover, aneuploidy patterns that were previously described by Bulk Genome Sequencing as emerging during first contact of promastigotes populations with different drugs are already pre-existing in single karyotypes in the SCGS data, suggesting a (pre-)adaptive role of MA. Additionally, the degree of somy variation was chromosome-specific. The SCGS also revealed a small fraction of cells where one or more chromosomes were nullisomic. Together, these results demonstrate the power of SCGS to resolve sub-clonal karyotype heterogeneity in and pave the way for understanding the role of MA in these parasites’ adaptability.