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Title: Investigations into the within-host genomic diversity and phenotypic variation of Plasmodium falciparum
Author: Murray, L.
ISNI:       0000 0004 6349 6891
Awarding Body: London School of Hygiene & Tropical Medicine
Current Institution: London School of Hygiene and Tropical Medicine (University of London)
Date of Award: 2017
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Clinical isolates of Plasmodium falciparum frequently consist of multiple genotypes, particularly within areas of high infection endemicity. Understanding these infections is important, as they have been linked to traits including drug resistance and virulence. I compared interpretations on within-host diversity from the allele-frequency based FWS index on whole-genome sequence data within a set of clinical isolates with microsatellite genotyping. Whilst there is significant correlation between the two methodologies, over half of isolates consist predominantly of a single genotype by FWS, yet possess multiple genotypes through the microsatellite typing, thus providing correlative and complementary information. Furthermore, genomic diversity within an infection can be produced by meiotic recombination between heterozygous haploid gametes, classical superinfection, or arise through de novo mutation. I applied FWS as a genome-wide scan of diversity within four West African populations. Through two approaches, utilising gene and bootstrapped SNP scores of FWS, I attempted to identify loci with consistently high levels of within-host homozygosity relative to their withinpopulation heterozygosity, that were posited to be under within-host directional selection. These approaches did not consistently identify outlier loci across the four populations, but I discuss alternative approaches to identify targets of within-host selection. Experimental work performed within rodent malaria species has reported that growth and gametocytogenesis are two plastic phenotypes affected by the presence of a competitor genotype. Utilising qPCR assays, comparison of the expression levels of the gametocyte commitment markers AP2-G and GDV-1, in addition to the sex-specific markers Pfs25 and Pfs230p, between two West African populations of differing transmission intensity, were all not significantly different. In addition, I designed a multicycle low starting density growth assay, initially using four laboratory clones, finding mean parasite multiplication rate (PMR) of these clones to range from 7.59 to 10.53 over the assay. When applied to fourteen recently adapted ex vivo clinical isolates, mean PMR was much lower at 2.90, with a range of 1.97 to 4.10. In combination with allele-specific qPCR, the growth assay was applied to assess competitive growth of the four laboratory clones, with no evidence of a significant induced plastic growth response of any clone. Together, these approaches will enable competitive growth studies of clinical isolates. The baseline quantitative data provided in this thesis supports the investigation of plastic phenotypes of P. falciparum.
Supervisor: Conway, D. J. Sponsor: Biotechnology and Biological Sciences Research Council
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral