New Assays to Characterise Growth-Related Phenotypes of Plasmodium falciparum Reveal Variation in Density-Dependent Growth Inhibition between Parasite Lines

PLoS One. 2016 Oct 25;11(10):e0165358. doi: 10.1371/journal.pone.0165358. eCollection 2016.

Abstract

The growth phenotype of asexual blood stage malaria parasites can influence their virulence and also their ability to survive and achieve transmission to the next host, but there are few methods available to characterise parasite growth parameters in detail. We developed a new assay to measure growth rates at different starting parasitaemias in a 96-well format and applied it to characterise the growth of Plasmodium falciparum lines 3D7-A and 3D7-B, previously shown to have different invasion rates and to use different invasion pathways. Using this simple and accurate assay we found that 3D7-B is more sensitive to high initial parasitaemia than 3D7-A. This result indicates that different parasite lines show variation in their levels of density-dependent growth inhibition. We also developed a new assay to compare the duration of the asexual blood cycle between different parasite lines. The assay is based on the tight synchronisation of cultures to a 1 h parasite age window and the subsequent monitoring of schizont bursting and formation of new rings by flow cytometry. Using this assay we observed differences in the duration of the asexual blood cycle between parasite lines 3D7 and HB3. These two new assays will be useful to characterise variation in growth-related parameters and to identify growth phenotypes associated with the targeted deletion of specific genes or with particular genomic, transcriptomic or proteomic patterns. Furthermore, the identification of density-dependent growth inhibition as an intrinsic parasite property that varies between parasite lines expands the repertoire of measurable growth-related phenotypic traits that have the potential to influence the outcome of a malarial blood infection.

MeSH terms

  • Erythrocytes / parasitology
  • Flow Cytometry
  • Genomics
  • Humans
  • Life Cycle Stages
  • Malaria, Falciparum / diagnosis
  • Malaria, Falciparum / parasitology
  • Merozoites / physiology
  • Microarray Analysis
  • Parasitemia / diagnosis
  • Parasitemia / parasitology*
  • Phenotype
  • Plasmodium falciparum / growth & development*
  • Plasmodium falciparum / physiology
  • Proteomics
  • Schizonts / growth & development

Grants and funding

This research was supported by Spanish Ministry of Economy and Competitiveness (MINECO, http://www.mineco.gob.es/portal/site/mineco/idi) grant SAF2013-43601-R to AC, co-funded by the European Regional Development Fund (ERDF, European Union, http://ec.europa.eu/regional_policy/en/funding/erdf/), and grant 2014 SGR 485 to AC from the Secretary for Universities and Research under the Department of Economy and Knowledge of the Government of Catalonia (http://universitatsirecerca.gencat.cat/ca/inici/). ET-F is supported by fellowship BES-2014-067901 by MINECO, co-funded by the European social fund (EU). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.