Facilities

The Plasmodium berghei research model

A malaria parasite of rodents, Plasmodium berghei, is used as a research model. We mainly use parasites of the ANKA strain of P. berghei. The ANKA strain and its clones have been obtained from the Prince Leopold Institute for Tropical Medicine in Antwerp. Parasites are maintained and analyzed either in culture in vitro or in vivo in laboratory animals (rodents). 

Infections of mosquitoes

Facilities are available for breeding of Anopheles stephensi mosquitoes and for mosquito transmission of P. berghei. Mosquitoes are reared and maintained in climate incubators. Mosquitoes are infected via membrane feeding of infected blood.

In vitro cultivation of the blood stages of the parasite

In our laboratory technologies have been developed for the short- and long-term in vitro culture of the blood stages. Standardized cultures with synchronized development of parasites are used for a wide range of applications, such as drug sensitivity testing, drug screening, phenotype analysis, production of parasites for genetic modification and isolation of the different parasite forms for molecular analysis.

In vitro cultivation of mosquito stages of the parasite

We have developed standardized techniques for in vitro gamete production, fertilization and zygote development. This development of the parasite occurs normaly in the midgut of mosquitoes, which is less accesible for research purposes. We are able to produce large numbers of ookinetes, that are highly infectious to mosquitoes when fed using membrane feeding procedures.

Molecular biology techniques

All the standard technologies for analysis of DNA and RNA are used routinely in our laboratory. Specialized assays for the analysis of transcription have been developed. These activities have generated resources that can be requested and provided a technology base that continues to be developed. For example, in situ RNA hybridisation, RNA interference, malaria parasite transfection and DNA microarrays.

Genome and Post-Genome Technologies

Genome and high throughput post-genomic expression studies are being performed on P. berghei; including genome analysis, transcriptome and proteome. Genome studies, in collaboration with the Sanger Centre, Hinxton, UK, has involved the generation of syntenic maps which describe the positional relationship of genes on chromosomes between different species of Plasmodium.
Detailed transcriptome analysis of various developmental stages (distinct time-points during asexual and sexual development in blood) of the P. berghei have been conducted using microarray chips. Differential transcription profiles have been generated and are available here. Proteome analysis studies have been performed in collaboration with Prof Matthias Mann (Odense, Denmark) and Dr. Edwin Lasonder (Nijmegen, The Netherlands), on uniquely acquired life cycle stages (i.e. separated male and female mature P. berghei gametocytes, purified merozoites of different species of Plasmodium; sporozoites). This analysis has been performed using Liquid Chromatography Tandem Mass Spectrometry (LC-MSMS) and the resulting spectra analysed using the MASCOT algorithm. Proteomes generated from different samples have informed targeted gene disruption studies aimed at looking at function of proteins at different points of parasite development; lists of proteins identified by this analysis can be found here.

Genetic modification of malaria parasites

Technologies for genetic modification of P. berghei have been developed in our laboratory (Transient and stable transfection; Expression of transgenes; Gene knock-out; Luciferase and GFP expression), providing a broad range of genetic tools for the study of malaria parasite biology. New tools are continously under development and a number of well defined gene knockout mutant parasites are available.

Mutant and transgenic lines of P. berghei

We have generated a number of mutant, transgenic and knock-out parasite lines which are used in our studies. For example parasite lines that express transgenes such as Green Fluorescence Protein (GFP) and Luciferase. These paraites are used to investigate specific interactions of the parasite with cells of their hosts.

In vivo imaging of parasite-host interactions

Advances in genetic modification of malaria parasites and imaging technologies for visualizing cells expressing reporter genes have significantly broadened the possibilities for in vivo studies of host parasite interactions. Transgenic parasites expressing bioluminescent reporter proteins (i.e. luciferase) are used in real-time imaging experiments with either live animals or in isolated organs. Imaging is performed using intensified charge-coupled device (I-CCD) photon-counting video camera which is part of the IVIS100 system (Xenogen)

Technologies to study phenotype and cellular aspects of malaria development

 The combination of in vitro cultivation of the parasite and in vivo infections permit the detailed analysis of the phenotype of the parasites. For example, gamete fertilisation, zygote development, synchronous bloodstage development, gametocyte formation. In addition: FACS analysis of growth characteristics of blood stages; Drug screening assays; Light- and fluorescence microscopy; Visualisation of live, GFP expressing parasites.