The genomes of Plasmodium spp. encode a number of different multigene families that are thought to play a critical role for survival. However, with the exception of the P. falciparum var genes, very little is known about the biological roles of any of the other multigene families. Using the recently developed Selection Linked Integration method, we have been able to activate the expression of a single member of a multigene family of our choice in Plasmodium spp. from its endogenous promoter. We demonstrate the usefulness of this approach by activating the expression of a unique var, rifin and stevor in P. falciparum as well as yir in P. yoelii. Characterization of the selected parasites reveals differences between the different families in terms of mutual exclusive control, co-regulation, and host adaptation. Our results further support the application of the approach for the study of multigene families in Plasmodium and other organisms.

Plasmodium falciparum has developed extensive mechanisms to evade host immune clearance. Currently, most of our understanding is based on in vitro studies of individual parasite variant surface antigens and how this relates to the processes in vivo is not well-understood. Here, we have used a humanized mouse model to identify parasite factors important for in vivo growth. We show that upregulation of the specific PfEMP1, VAR2CSA, provides the parasite with protection from macrophage phagocytosis and clearance in the humanized mice. Furthermore, parasites adapted to thrive in the humanized mice show reduced NK cell-mediated killing through interaction with the immune inhibitory receptor, LILRB1. Taken together, these findings reveal new insights into the molecular and cellular mechanisms that the parasite utilizes to coordinate immune escape in vivo. Identification and targeting of these specific parasite variant surface antigens crucial for immune evasion provides a unique approach for therapy.

Plasmodium falciparum has developed extensive mechanisms to evade host immune clearance. Currently, most of our understanding is based on in vitro studies of individual parasite variant surface antigens and how this relates to the processes in vivo is not well-understood. Here, we have used a humanized mouse model to identify parasite factors important for in vivo growth. We show that upregulation of the specific PfEMP1, VAR2CSA and the RIFIN PF3D7_1254800 provides the parasite with protection from macrophage phagocytosis and natural killer cell mediated killing. Taken together, these findings reveal new insights on the molecular and cellular mechanisms that coordinate the immune escape process the parasite utilizes in vivo. As immune evasion may be particularly important during the establishment of the blood stage infection when parasite numbers are still relatively small, identification of specific parasite variant surface antigens provides targets for developing more effective vaccines by targeting parasite immune evasion.

I would like to offer my deepest gratitude to everybody who helped throughout this PhD and transformed those 4 years into this amazing journey.First and foremost, I would like to thank my Professor and mentor, Professor Peter Preiser, who gave me this opportunity. He provided me with guidance, advice and input on my way. He always allowed and encouraged me to pursue my own ideas, giving me full control of the project. He was always open to new approaches and allowed me to follow up on exciting results, even if it they did not directly involve my main project. I appreciate this freedom, and I never regretted starting this project. Special thanks to all the people from the lab and the malaria community in Singapore in general. Thank you to Dr Omar Sheriff, for giving me the supervision that I needed. Even though it was never his task, he invested hours into helping with experiments, interpretations and troubleshooting. Thank you for teaching me how to keep my parasites happy and how to properly kill them afterwards. Thank you to Ameya Sinha, not only for running the mass spectrometry for me, but for keeping me company, having delightful discussions about god and the world and listening to my occasional complaints. You are, truly, the best. A lot of thanks to Marvin Chew, who unexpectedly stumbled into the world of multigene families, making him my partner in crime and helping me with the immunology aspect of this work.Thanks to my student Chua Zong Sheng and his invaluable help with the recombinant protein expression and binding assays. Thank you Dr Regina Hoo for your help with the microarray and the data analysis. Thank you Jenny for taking care of all the admin things, I would not have been able to keep up with all the rules and regulations without you.