Indels, structural variation, and recombination drive genomic diversity in Plasmodium falciparum

Abstract

The malaria parasite Plasmodium falciparum has a great capacity for evolutionary adaptation to evade host immunity and develop drug resistance. Current understanding of parasite evolution is impeded by the fact that a large fraction of the genome is either highly repetitive or highly variable and thus difficult to analyze using short-read sequencing technologies. Here, we describe a resource of deep sequencing data on parents and progeny from genetic crosses, which has enabled us to perform the first genome-wide, integrated analysis of SNP, indel and complex polymorphisms, using Mendelian error rates as an indicator of genotypic accuracy. These data reveal that indels are exceptionally abundant, being more common than SNPs and thus the dominant mode of polymorphism within the core genome. We use the high density of SNP and indel markers to analyze patterns of meiotic recombination, confirming a high rate of crossover events and providing the first estimates for the rate of non-crossover events and the length of conversion tracts. We observe several instances of meiotic recombination within copy number variants associated with drug resistance, demonstrating a mechanism whereby fitness costs associated with resistance mutations could be compensated and greater phenotypic plasticity could be acquired.

Culture adaptation of malaria parasites selects for convergent loss-of-function mutants

“…The present study focuses on single nucleotide variants that are robustly called, whereas development of methods to accurately detect novel indels and structural variants in Illumina sequences of P. falciparum cultures might have the potential to identify additional changes40. Although indels might cause frameshifts in other genes, it is nonetheless remarkable that, out of more than 5000 genes in the P. falciparum genome, only five ( Epac, SRPK1 and 3 ApiAP2 genes) are identified to have nonsense SNP mutants causing premature stop codon alleles that have emerged in culture.…”

Fitness Costs and the Rapid Spread of kelch13 -C580Y Substitutions Conferring Artemisinin Resistance

“…falciparum. We recalibrated base quality scores based on a set of verified known variants (33). We first called variants independently for each parasite line using HaplotypeCaller and then merged them by using GenotypeGVCFs, according to default parameters but with –sample_ploidy 1.…”

“…We excluded variants from variable regions (subtelomeric repeats, hypervariable regions, and centromeres) of the P. falciparum genome (33). As the Cas9 protein is transiently expressed (15) during CRISPR and all the parasite lines were originally obtained from single-cell cloning of CRISPR-edited parasites, any SNPs or indels from off-target activity should appear in 100% of the cloned population.…”

An open dataset of Plasmodium falciparum genome variation in 7,000 worldwide samples

“…The predominance of indels in non-coding regions has been previously observed and is most likely a consequence of the extreme AT bias which leads to many short repetitive sequences. 56,57 For the purpose of this analysis, we excluded all variants in subtelomeric and internal hypervariable regions, mitochondrial and apicoplast genomes, and some other regions of the genome where the mapping of short sequence reads is prone to a high error rate due to extremely high rates of variation. 56 A total of 1,838,733 SNPs (of which 1,626,886 were biallelic) and 1,276,027 indels (or SNP/indel combinations) passed all these filters.…”

“…48,49 The data have also been used to develop and test methods for estimating identity by descent 50,51 , imputation 52 , typing structural variants 53 , designing other SNP genotyping platforms 54 and data visualisation 8,55 . In a companion study we performed whole genome sequencing of experimental genetic crosses of P. falciparum, and this provided a benchmark to test the accuracy of our genotyping methods, and to conduct an in-depth analysis of indels, structural variants and recombination events which are complicated to ascertain in these population genetic samples 56 .…”