Open Access Methodology

A rapid and scalable density gradient purification method for Plasmodium sporozoites

Mark Kennedy1, Matthew E Fishbaugher1, Ashley M Vaughan2, Rapatbhorn Patrapuvich3, Rachasak Boonhok34, Narathatai Yimamnuaychok3, Nastaran Rezakhani2, Peter Metzger2, Marisa Ponpuak4, Jetsumon Sattabongkot3, Stefan H Kappe25, Jen CC Hume1 and Scott E Lindner2*

Author Affiliations

1 Center for Mosquito Production and Malaria Infection Research, Seattle Biomedical Research Institute, 307 Westlake Avenue North, Suite 500, Seattle, WA, 98109, USA

2 Malaria Program, Seattle Biomedical Research Institute, 307 Westlake Avenue North, Suite 500, Seattle, WA, 98109, USA

3 Mahidol Vivax Research Center, Faculty of Tropical Medicine, Mahidol University, Bangkok, Thailand

4 Department of Microbiology, Faculty of Science, Mahidol University, Bangkok, Thailand

5 Department of Global Health, University of Washington, Seattle, WA, 98195, USA

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Malaria Journal 2012, 11:421  doi:10.1186/1475-2875-11-421

Published: 17 December 2012

Abstract

Background

Malaria remains a major human health problem, with no licensed vaccine currently available. Malaria infections initiate when infectious Plasmodium sporozoites are transmitted by Anopheline mosquitoes during their blood meal. Investigations of the malaria sporozoite are, therefore, of clear medical importance. However, sporozoites can only be produced in and isolated from mosquitoes, and their isolation results in large amounts of accompanying mosquito debris and contaminating microbes.

Methods

Here is described a discontinuous density gradient purification method for Plasmodium sporozoites that maintains parasite infectivity in vitro and in vivo and greatly reduces mosquito and microbial contaminants.

Results

This method provides clear advantages over previous approaches: it is rapid, requires no serum components, and can be scaled to purify >107 sporozoites with minimal operator involvement. Moreover, it can be effectively applied to both human (Plasmodium falciparum, Plasmodium vivax) and rodent (Plasmodium yoelii) infective species with excellent recovery rates.

Conclusions

This novel method effectively purifies viable malaria sporozoites by greatly reducing contaminating mosquito debris and microbial burdens associated with parasite isolation. Large-scale preparations of purified sporozoites will allow for enhanced in vitro infections, proteomics, and biochemical characterizations. In conjunction with aseptic mosquito rearing techniques, this purification technique will also support production of live attenuated sporozoites for vaccination.

Keywords:
Sporozoite; Purification; Plasmodium; Falciparum; Vivax; Yoelii; Humanized mouse model