Computer software for testing drug susceptibility of malaria parasites

A computer program is described for the automated analysis of data obtained by flow cytometry for in vitro antimalarial drug susceptibility testing. Samples of malaria-infected red blood cells (RBC), which were cultured in the presence of different concentrations of antimalarial drugs, were stained with Hoechst. The Hoechst fluorescence intensity of infected RBC corresponds to DNA content of the parasites and to their stage of development. After measurement of the samples by a FACStar flow cytometer equipped with a UV laser and an autosampler, FCS 1.0 data files were generated. The HP PAS-CAL program developed for these files identifies five different populations--uninfected RBC, infected RBC, free parasites, leukocytes, and debris--on the basis of their light scatter and fluorescence characteristics. The program calculates the percentage of infected cells, the total number of parasite nuclei, and the average number of nuclei per parasite. The results of each culture are presented as a drug dose-response curve. During data analysis, user interaction is limited to selecting the first file of the first culture. The algorithm then processes each culture automatically. Potential problems or difficulties in analysis are flagged. To date, a total of 862 drug tests have been evaluated and fall into two classes, an extended microtest and the World Health Organization standardized microtest. These tests gave satisfactory results in more than 99% of the cases.