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Infection and Immunity, February 2002, p. 655-660, Vol. 70, No. 2
0019-9567/01/$04.00+0     DOI: 10.1128/IAI.70.2.655-660.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Plasmodium knowlesi Provides a Rapid In Vitro and In Vivo Transfection System That Enables Double-Crossover Gene Knockout Studies

Clemens H. M. Kocken,1 Hastings Ozwara,1,2 Annemarie van der Wel,1 Annette L. Beetsma,1 Jason M. Mwenda,2 and Alan W. Thomas1*

Department of Parasitology, Biomedical Primate Research Centre, 2280 GH Rijswijk, The Netherlands,1 Departments of Infectious Diseases and Reproductive Biology, Institute of Primate Research, National Museums of Kenya, Karen, Nairobi, Kenya2

Received 26 July 2001/ Returned for modification 19 September 2001/ Accepted 31 October 2001

Transfection technology for malaria parasites provides a valuable tool for analyzing gene function and correlating genotype with phenotype. Transfection models are even more valuable when appropriate animal models are available in addition to complete in vitro systems to be able to fully analyze parasite-host interactions. Here we describe the development of such a model by using the nonhuman primate malaria Plasmodium knowlesi. Blood-stage parasites were adapted to long-term in vitro culture. In vitro-adapted parasites could readapt to in vivo growth and regain wild-type characteristics after a single passage through an intact rhesus monkey. P. knowlesi parasites, either in vitro adapted or in vivo derived, were successfully transfected to generate circumsporozoite protein (CSP) knockout parasites by double-crossover mechanisms. In vitro-transfected and cloned CSP knockout parasites were derived in a time span of only 18 days. Microscopic evaluation of developing oocysts from mosquitoes that had fed on CSP knockout parasites confirmed the impairment of sporozoite formation observed in P. berghei CSP knockout parasites. The P. knowlesi model currently is the only malaria system that combines rapid and precise double-crossover genetic manipulation procedures with complete in vitro as well as in vivo possibilities. This allows for full analysis of P. knowlesi genotype-phenotype relationships and host-parasite interactions in a system closely related to humans.

* Corresponding author. Mailing address: BPRC, Department of Parasitology, Lange Kleiweg 139, 2280 GH Rijswijk, The Netherlands. Phone: (31) 15-2842-538. Fax: (31) 15-2843-986. E-mail: thomas{at}bprc.nl.

Editor: J. M. Mansfield

Infection and Immunity, February 2002, p. 655-660, Vol. 70, No. 2
0019-9567/01/$04.00+0     DOI: 10.1128/IAI.70.2.655-660.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Conway, D. J. (2007). Molecular Epidemiology of Malaria. Clin. Microbiol. Rev. 20: 188-204 [Abstract] [Full Text]  
  • Ozwara, H., Langermans, J. A. M., Kocken, C. H. M., van der Wel, A., van der Meide, P. H., Vervenne, R. A. W., Mwenda, J. M., Thomas, A. W. (2003). Transfected Plasmodium knowlesi Produces Bioactive Host Gamma Interferon: a New Perspective for Modulating Immune Responses to Malaria Parasites. Infect. Immun. 71: 4375-4381 [Abstract] [Full Text]  
  • OZWARA, H., LANGERMANS, J. A. M., MAAMUN, J., FARAH, I. O., YOLE, D. S., MWENDA, J. M., WEILER, H., THOMAS, A. W. (2003). EXPERIMENTAL INFECTION OF THE OLIVE BABOON (PAPIO ANUBIS) WITH PLASMODIUM KNOWLESI: SEVERE DISEASE ACCOMPANIED BY CEREBRAL INVOLVEMENT. Am J Trop Med Hyg 69: 188-194 [Abstract] [Full Text]  


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