• Genomic Epidemiology of the Malaria Parasite Plasmodium falciparum: Implications for Whole-Organism Malaria Vaccine Development

      Moser, Kara; Carneiro da Silva, Joana; Plowe, Christopher V. (2018)
      Whole-organism malaria vaccines have shown great promise in initial clinical trials against controlled human malaria infections (CHMI) with homologous P. falciparum strains. However, efficacy against heterologous CHMI and against natural infection is somewhat lower. While the reason for lower efficacy against non-vaccine strains is unknown, several knowledge gaps, if addressed, would assist in developing highly-efficacious whole-organism malaria vaccines. First, regions of the parasite genome responsible for protection have not been identified. Such identification could be achieved by comparing breakthrough P. falciparum infections in vaccinated individuals with the vaccine strain; however, it is not clear how best to genetically characterize breakthrough infections, in vaccinated individuals. Second, vaccine and challenge strains have not been characterized and compared at the genomic level to circulating strains from parasite populations in malaria endemic regions. Finally, a rigorous investigation of allele frequency fluctuations between temporally isolated parasite populations would help predict if such changes would affect vaccine efficacy. Using next- and third-generation sequencing technologies, new reference assemblies for whole-organism malaria vaccine and CHMI strains were generated, along with reference assemblies for 19 clinical isolates to use for improved read mapping and characterization of clinical P. falciparum isolates. Using a geographic-specific reference assembly improved the ability to characterize clinical isolates through increased read coverage, and so these references may be helpful to characterize breakthrough infections (particularly for studies in Southeast Asian populations). In addition, assemblies for vaccine and CHMI strains reveal thousands of variants between these parasites. CHMI strains also have variants which convey differences in immunological potential, thus confirming their appropriateness as heterologous CHMI strains. Vaccine and CHMI strains were also shown to representative of their respective geographic origins when compared to extant parasite populations from malaria endemic regions. Finally, we show that while there were fluctuations in allele frequencies between West and East African parasite populations over almost a decade, none affected the frequency of vaccine candidate alleles in a way that would meaningfully impact vaccine efficacy. These results will greatly assist in the interpretation of whole-organism malaria vaccine trials, and will pave the way for the design of next-generation whole-organism malaria vaccines.
    • Theileria infections in African cattle and buffalo: understanding genetic variation and speciation

      Palmateer, Nicholas; Carneiro da Silva, Joana; 0000-0002-4307-8049 (2021)
      East Coast fever, caused by the apicomplexan parasite Theileria parva, has an estimated annual death toll of over a million cattle in endemic sub-Saharan regions. The African Cape buffalo is the natural reservoir of T. parva and rarely exhibits clinical symptoms when infected, but transmits the parasite to cattle via a tick vector. Previous studies based on a few genetic markers showed that buffalo-derived T. parva subpopulations contain greater antigenic diversity than those from cattle. Interestingly, cattle are infected and killed by T. parva of buffalo origin, but cannot transmit those parasites, suggesting that a degree of host specificity exists. The characterization of genetic variation within and between cattle- and buffalo-derived T. parva is critical to understand the molecular mechanism(s) of host specificity. To overcome obstacles in T. parva biology that prevent the straightforward acquisition of sufficient DNA for whole genome sequencing (WGS), we adapted a DNA capture approach to select T. parva from a mix of parasite and bovine DNA obtained from T. parva-infected bovine lymphocyte cultures. To gain access to variable genomic regions that cannot be characterized through read mapping approaches, we assembled the captured reads de novo. From starting material of <1%-4% parasite DNA in a mixed sample from host and parasite, >98% of sequence reads post-capture are of parasite origin and >97% of the genome is recovered, reflecting the method’s high specificity and sensitivity. We used this whole genome DNA capture followed by sequencing to generate WGS data from 15 cattle- and 24 buffalo-derived T. parva isolates. This resulted in the generation of the first assembly of a buffalo-derived T. parva isolate. Furthermore, we determined that cattle- and buffalo-derived T. parva isolates differ in various measures at levels consistent with speciation. Finally, capture and analysis of members of the T. parva repeat (Tpr) multigene family, which encode some of the most variable antigen families in the species, enabled the study of Tpr evolution and initial inferences of its possible involvement in parasite-host interactions. These results have greatly advanced the study of the T. parva genome and improved our understanding of the evolution of this parasite population.