Exploring the evolution and adaptive role of mosaic aneuploidy in a clonalLeishmania donovanipopulation using high throughput single cell genome sequencing

Abstract

14Maintenance of stable ploidy over continuous mitotic events is a paradigm for most higher 15 eukaryotes. Defects in chromosome segregation and/or replication can lead to aneuploidy, a 16 condition often considered deleterious. However, in Leishmania, a Protozoan parasite, 17aneuploidy is a constitutive feature, where variations of somies represent a mechanism of gene 18 expression adaptation, possibly impacting phenotypes. Strikingly, clonal Leishmania 19 populations display cell-to-cell somy variation, a phenomenon named mosaic aneuploidy (MA). 20However, until recently, no method was available for the determination of the complete 21 karyotype of single Leishmania parasites. To overcome this limitation, we used here for the first 22 time a high-throughput single-cell genomic sequencing (SCGS) method to estimate individual 23 karyotypes of 1560 promastigote cells in a clonal population of Leishmania donovani. We 24 identified 128 different karyotypes, of which 4 were dominant. A network analysis revealed that 25 most karyotypes are linked to each other by changes in copy number of a single chromosome 26 and allowed us to propose a hypothesis of MA evolution. Moreover, aneuploidy patterns that 27 were previously described by Bulk Genome Sequencing as emerging during first contact of 28 promastigotes populations with different drugs are already pre-existing in single karyotypes in 29 the SCGS data, suggesting a (pre-)adaptive role of MA. Additionally, the degree of somy 30 variation was chromosome-specific. The SCGS also revealed a small fraction of cells where one 31 or more chromosomes were nullisomic. Together, these results demonstrate the power of SCGS 32to resolve sub-clonal karyotype heterogeneity in Leishmania and pave the way for 33understanding the role of MA in these parasites' adaptability. 34 35

Impact of Genetic Diversity and Genome Plasticity of Leishmania spp. in Treatment and the Search for Novel Chemotherapeutic Targets

“…Interestingly, large-scale research involving a large number of samples and wide geographic range has revealed that genetic diversity is significantly higher than that previously reported ( Bussotti et al., 2018 ; Zackay et al., 2018 ; Franssen et al., 2020 ; Imamura et al., 2020 ; Patino et al., 2020 ; Salloum et al., 2020 ; Zheng et al., 2020 ; Patino et al., 2021 ; Schwabl et al., 2021 ). Recently, single-cell sequencing has demonstrated the presence of several distinct karyotypes within the same Leishmania clone ( Imamura et al., 2020 ; Negreira et al., 2020 ), and multiple-genotype infections have been demonstrated to occur even within the same host and tissue ( Cupolillo et al., 2020 ).…”

DNA Double-Strand Breaks: A Double-Edged Sword for Trypanosomatids

“…Leishmania spp. have remarkably plastic genomes, with genomic alterations such as aneuploidy (Mannaert et al, 2012;Lachaud et al, 2014), and CNVs (Rogers et al, 2011;Bussotti et al, 2018), which seems to be widespread phenomena among the species (Rogers et al, 2011;Lachaud et al, 2014;Negreira et al, 2020). Interspersed throughout the genome of Leishmania spp.…”

Of Drugs and Trypanosomatids: New Tools and Knowledge to Reduce Bottlenecks in Drug Discovery

“…We are however currently revisiting concepts and moving from genome-wide approaches in parasite populations (or clones) to single-cell-based strategies to better understand the plasticity of Leishmania parasites that harbor mosaic aneuploidy—a feature that has impairments in the way the parasite will respond or not to a given drug. Using a single-cell genomic sequencing method, Negreira et al (2020) identified 128 different karyotypes in 1560 L. donovani promastigotes [ 136 ]. They highlight the fact that some karyotypes presented pre-existing adaptations to antimony-based drugs, supporting a hypothesis raised even before this hint [ 137 , 138 ].…”