1. Rapid and iterative genome editing in the malaria parasite Plasmodium knowlesi provides new tools for P. vivax research.
- Author
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Mohring F, Hart MN, Rawlinson TA, Henrici R, Charleston JA, Diez Benavente E, Patel A, Hall J, Almond N, Campino S, Clark TG, Sutherland CJ, Baker DA, Draper SJ, and Moon RW
- Subjects
- Animals, Antigens, Protozoan genetics, Antigens, Protozoan immunology, Antigens, Protozoan metabolism, Biomedical Research methods, Biomedical Research trends, Humans, Malaria immunology, Malaria parasitology, Malaria prevention & control, Malaria Vaccines administration & dosage, Malaria Vaccines immunology, Malaria, Vivax immunology, Parasites immunology, Parasites physiology, Plasmodium knowlesi immunology, Plasmodium knowlesi physiology, Protozoan Proteins genetics, Protozoan Proteins immunology, Protozoan Proteins metabolism, Receptors, Cell Surface genetics, Receptors, Cell Surface immunology, Receptors, Cell Surface metabolism, Gene Editing methods, Malaria, Vivax genetics, Parasites genetics, Plasmodium knowlesi genetics
- Abstract
Tackling relapsing Plasmodium vivax and zoonotic Plasmodium knowlesi infections is critical to reducing malaria incidence and mortality worldwide. Understanding the biology of these important and related parasites was previously constrained by the lack of robust molecular and genetic approaches. Here, we establish CRISPR-Cas9 genome editing in a culture-adapted P. knowlesi strain and define parameters for optimal homology-driven repair. We establish a scalable protocol for the production of repair templates by PCR and demonstrate the flexibility of the system by tagging proteins with distinct cellular localisations. Using iterative rounds of genome-editing we generate a transgenic line expressing P. vivax Duffy binding protein (PvDBP), a lead vaccine candidate. We demonstrate that PvDBP plays no role in reticulocyte restriction but can alter the macaque/human host cell tropism of P. knowlesi . Critically, antibodies raised against the P. vivax antigen potently inhibit proliferation of this strain, providing an invaluable tool to support vaccine development., Competing Interests: FM, MH, TR, RH, JC, ED, AP, JH, NA, SC, TC, CS, DB, SD, RM No competing interests declared, (© 2019, Mohring et al.)
- Published
- 2019
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