The drug susceptibility of the Plasmodium falciparum parasite can be assessed with in vitro testing to measure fundamental parasite antimalarial sensitivity. Determining parasite growth outside of the human host eliminates host effects such as drug metabolism and immunity. Key roles for the In vitro Module include providing support to the research community to standardise in vitro methods, and generating tools and methodology for data analysis and upload into the WWARN Data Repository.
Another important role for the In vitro Module is to strengthen the practical value of information on the in vitro susceptibility of Plasmodium falciparum isolates to different antimalarial drugs by providing up-to-date summary information at the country and regional levels, and by improving communication among researchers, malaria control programs and policy makers.
What we do
The In vitro Module curates the in vitro data in the WWARN Data Repository — primarily drug susceptibility results from malaria parasites collected from infected patients — for display in WWARN Explorer, a web-based visualisation tool. View In vitro data in the WWARN Explorer preview.
The In vitro Data Management and Statistical Analytical Plan (DMSAP) provides a transparent description of the methodology used to handle and analyse in vitro data. WWARN has developed the In Vitro Analysis and Reporting Tool (IVART) using an algorithm based on the INSERM-developed ICEstimator (see below) to generate reproducible IC50 estimates. IVART produces an automated report documenting the data upload, transformation and analysis process, including the results generated.
The In vitro Module also supports researchers in generating quality, cross-comparable data by providing Standard Operating Procedures (SOPs), shared test reagents (see QA/QC), and agreed standard strains of Plasmodium to use as controls.
In May 2009, WWARN held its first In vitro Technical Advisory Group (TAG) meeting in Paris, France. This meeting identified a clear need to focus on internal quality controls and quality assurance, in order to normalise data from different methods and ensure reliable results. In response, the In vitro Module hosted a technical workshop aimed at harmonising methodology across in vitro laboratories, particularly by encouraging the use of the culture adapted clone 3D7 as a common control. The ratio of a test isolate IC50 value will be compared with the response of the 3D7 clone.
Encouraging common key parameters of in vitro tests and associated measures of quality control, for example the use of standard reference clones, will allow comparison of in vitro susceptibilities between laboratories and within a single laboratory over time. The results will contribute to the understanding of parasite resistance and its geographic and temporal patterns.
For more information, contact invitro@wwarn.org.
Collaborations
Surveillance of imported malaria
The In vitro Module is currently involved with a collaborative project undertaking the surveillance malaria cases that are imported to non-endemic countries by travellers returning to Europe (France, UK and tropical networks) and North America (US and Canada). The goal is to determine whether this surveillance data can contribute to the detection of emerging antimalarial drug resistance in endemic areas, where accurate information is not otherwise available.
In vitro tools
Epidemiological surveillance of drug susceptibility of malaria parasites is a major component of disease control. Measurement of in vitro response of the malaria parasites to drugs requires robust and precise methods. The usual parameter for monitoring in vitro susceptibility is the concentration that inhibits 50% of the parasite’s activity (IC50 value). Two tools have been developed by other groups to assist researchers with in vitro studies:
ICEstimator
Developed by the Inserm U738 team, the ICEstimator estimates IC50 concentration with the associated 95% confidence interval (95%CI). A precise determination of IC50 is needed for comparison of data among populations of malaria parasites. Confidence Interval determination may help to discard estimates with insufficient precision. The concentrations that inhibit 90% and 99% of the parasite’s activity (IC90 and IC99) can also be needed for comparison of data and are displayed in version 1.2. ICEstimator version 1.2 allows simple management of data measuring the effect of clones, strains, or isolates of malaria parasites resulting from uptake of labelled nutrient (isotopic test), production of proteins (ELISA tests), or growth (schizont maturation). The software estimates the drug concentration that inhibits growth by 50% (IC50) using the nls function of R, which is freely available on the R project website.
HN-NonLin
Developed by Harald Noedl, HN-NonLin is a basic, free tool developed to facilitate the analysis of malaria in vitro drug sensitivity test results. This software allows for the entry of raw data obtained from drug sensitivity assays (such as isotopic assays, colorimetric assays, or schizont maturation tests), calculates the principal parameters of drug sensitivity (inhibitory concentrations), and plots the dose-response curves for individual isolates, strains or clones.
