Understanding Developmental Decision Making in Malaria Parasites using Single-Cell Transcriptomics

Single-cell sequencing has the potential to revolutionise our understanding of malaria parasites. Malaria is caused by single-celled parasitic organisms of the genus Plasmodium which display remarkable cellular plasticity during their complex life cycle with large variations in size (1.2 to 50 μm) and the environments they inhabit in mammalian and mosquito hosts. Clinical symptoms of malaria result from asexual replication within red blood cells, whereas transmission to new hosts relies on fertilisation and replication in the mosquito. Both disease development and transmission are therefore underpinned by the parasite’s ability to serially differentiate into morphologically distinct forms. This versatility is orchestrated by tight regulation of a compact genome where, despite the importance of malaria in global health, the function of ~40% of genes remains unknown. Better understanding of gene use and gene function throughout the parasite’s life cycle is needed to inform the development of much needed new drugs, vaccines, and transmission-blocking strategies. The dearth in gene function knowledge is largely due to the inadequacy of bulk RNA-seq in profiling dynamic stages where there are fast transitions and insufficient methods to synchronize the population. In addition, some species of parasites such as P. malariae remain unprofiled at the transcriptional level due to the fact it is usually found at a low parasitemia in humans and is commonly co-infected with another species. Profiling of Plasmodium parasites using scRNA-seq would overcome these limitations. During my PhD, I used scRNA-seq to build a Malaria Cell Atlas which profiled all stages in the P. berghei life cycle using a modified Smart-seq2 protocol. In addition, I profiled the blood stages responsible for malaria symptoms at higher-cell-coverage using droplet-based scRNA-Seq in P. berghei, P. falciparum, and P. knowlesi. This work was followed by specifically profiling the sexual development of wild-type and single knock-out mutant P. berghei parasites using both droplet- and plate-based scRNA-seq. This has led to the identification of novel essential genes in gametocytogenesis and a greater understanding of the genes expressed during this developmental process which will be important in developing novel transmission-blocking strategies. I then developed and applied a scRNA-seq preservation protocol in order to profile circulating forms of P. malariae and P. falciparum from in vivo infections of volunteers in Mbita, Kenya.

Name dc:type.qualificationname

Doctor of Philosophy (PhD)

Level dc:type.qualificationlevel

Doctoral

Grantor dc:publisher.institution

University of Cambridge

Year dc:date.issued

2020

Author dc:creator

• Russell, Andrew

Advisors dc:contributor.advisor

• Voet, Thierry
• Billker, Oliver
• Lawniczak, Mara

• scRNA-seq
• infectious diseases
• single-cell
• genomics
• malaria

Licence

• https://apollo8-f-pro.lib.cam.ac.uk/bitstreams/4f69b28e-20d5-428e-955b-6be6f96eec7c/download
• https://www.rioxx.net/licenses/all-rights-reserved/

Language dc:language

eng

Author Identifier

0000-0001-5411-2807

OAI identifier oai:identifier

oai:www.repository.cam.ac.uk:1810/319601