Exploring the evolution and adaptive role of mosaic aneuploidy in a clonalLeishmania donovanipopulation using high throughput single cell genome sequencing
Authors: Negreira Gabriel H. , Monsieurs Pieter , Imamura Hideo , Maes Ilse , Kuk Nada , Yagoubat Akila , Van den Broeck Frederik , Sterkers Yvon , Dujardin Jean-Claude , Domagalska Malgorzata A. 
Published: March 2020

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, inLeishmania, a Protozoan parasite, aneuploidy is a constitutive feature, where variations of somies represent a mechanism of gene expression adaptation, possibly impacting phenotypes. Strikingly, clonalLeishmaniapopulations 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 singleLeishmaniaparasites. 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 ofLeishmania donovani. 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 inLeishmaniaand pave the way for understanding the role of MA in these parasites’ adaptability.

Update: 25thMay 2021

A revision of the present preprint was released in BioRxiv on 11thMay 2021 (https://www.biorxiv.org/content/10.1101/2021.05.11.443577v2). In the new version, we included two extra samples in our single-cell genome sequencing (SCGS) analysis – the BPK081 cl8 clone (a nearly euploid strain), and a population consisting of a mixture of fourL. donovanistrains which was used as control for high levels of mosaicism in aneuploidy and for estimation of doublets. We also upgraded the bioinformatics pipeline to determine single-cell karyotypes and performed new fluorescence in situ hybridization (FISH) analysis. The new findings observed especially in the BPK081 cl8 led to a reformulation of the text, a new hypothesis for the evolution of mosaicism and a general restructuring of the article. Therefore, the present preprint is obsolete. Please refer to the new preprint entitled “High throughput single cell genome sequencing gives insights in the generation and evolution of mosaic aneuploidy inLeishmania donovani” for more information.