Applying next generation sequencing of genomes and transcriptomes to investigate the population structure and biology of Plasmodium species

Plasmodium parasites, the causative agents of malaria, are responsible for a significant burden of disease worldwide. Understanding populations and parasite biology is critical to developing effective control mechanisms and questions relating to these have been addressed in this thesis using whole genome and transcriptome sequencing. In Southeast Asia, the zoonotic species Plasmodium knowlesi now causes a significant amount of malarial disease, particularly in Malaysia. Clinical isolates of P. knowlesi from peninsular Malaysia were whole genome sequenced and single nucleotide polymorphisms were used to inform population genomic analyses. This work confirmed the existence of a third, divergent population of P. knowlesi in peninsular Malaysia and uncovered evidence of selection acting on these parasites. In sub-Saharan Africa, P. falciparum is responsible for virtually all malarial disease. Clinical isolates sampled from West Africa were investigated using whole transcriptome sequencing of ex vivo parasites, focusing on a gene known as mspdbl2, which is a possible marker of gametocytogenesis. Schizonts from clinical isolates were assessed for MSPDBL2 expression by immunofluorescence assays and were whole transcriptome sequenced. Analysis of gene expression was carried out correlating to expression of MSPDBL2, revealing enrichment of genes with known or suspected involvement in gametocytogenesis. A limiting factor of this investigation was the very low amount of material available from ex vivo culture. An alternative avenue for investigating limiting material is through single-cell sequencing. One method for single-cell transcriptomics was trialled in this thesis and extensive optimisation resulted in the sequencing of transcriptomes from a small number of P. falciparum schizonts, offering a future methodology for investigating gene expression using very few parasites.