Impact of antimalarial treatment and chemoprevention on the drug sensitivity of malaria parasites isolated from ugandan children

Abstract

Changing treatment practices may be selecting for changes in the drug sensitivity of malaria parasites. We characterized ex vivo drug sensitivity and parasite polymorphisms associated with sensitivity in 459 Plasmodium falciparum samples obtained from subjects enrolled in two clinical trials in Tororo, Uganda, from 2010 to 2013. Sensitivities to chloroquine and monodesethylamodiaquine varied widely; sensitivities to quinine, dihydroartemisinin, lumefantrine, and piperaquine were generally good. Associations between ex vivo drug sensitivity and parasite polymorphisms included decreased chloroquine and monodesethylamodiaquine sensitivity and increased lumefantrine and piperaquine sensitivity with pfcrt 76T, as well as increased lumefantrine sensitivity with pfmdr1 86Y, Y184, and 1246Y. Over time, ex vivo sensitivity decreased for lumefantrine and piperaquine and increased for chloroquine, the prevalences of pfcrt K76 and pfmdr1 N86 and D1246 increased, and the prevalences of pfdhfr and pfdhps polymorphisms associated with antifolate resistance were unchanged. In recurrent infections, recent prior treatment with artemether-lumefantrine was associated with decreased ex vivo lumefantrine sensitivity and increased prevalence of pfcrt K76 and pfmdr1 N86, 184F, and D1246. In children assigned chemoprevention with monthly dihydroartemisinin-piperaquine with documented circulating piperaquine, breakthrough infections had increased the prevalence of pfmdr1 86Y and 1246Y compared to untreated controls. The noted impacts of therapy and chemoprevention on parasite polymorphisms remained significant in multivariate analysis correcting for calendar time. Overall, changes in parasite sensitivity were consistent with altered selective pressures due to changing treatment practices in Uganda. These changes may threaten the antimalarial treatment and preventive efficacies of artemether-lumefantrine and dihydroartemisinin-piperaquine, respectively.

Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Figures

FIG 1

Association between P. falciparum genetic polymorphisms and ex vivo drug sensitivity. Geometric mean IC50s are shown for parasites with wild-type (WT), mixed (Mix), or mutant (Mut) genotypes at the indicated alleles. The number of samples in each category is indicated by N. Values for each allele were compared between wild-type and mixed or mutant genotypes using t tests, and comparisons with P values of <0.05 (*), <0.01 (**), and <0.001 (***) are labeled.

FIG 2

Ex vivo sensitivity of P. falciparum over time. For longitudinal assessments of IC50 data, drug sensitivities were plotted as IC50s over time. Each point represents a single isolate. A representation of IC50 over time was plotted using the lowess smoothing function in Stata.

FIG 3

Allele prevalences over time. Prevalences of wild-type (WT), mixed (Mix), and mutant (Mut) alleles during the indicated intervals are shown. The number of samples in each category is indicated by N. Changes in wild-type allele prevalences compared to those in 2010 that were significant in univariate analyses are labeled for P values of <0.05 (*), <0.01 (**), and <0.001 (***). With multivariate analysis adjusting for the amount of time since last AL treatment and chemoprevention arms, changes in the prevalences of pfmdr1 86 and 184 mutations remained significant.

FIG 4

Impact of prior therapy with AL on ex vivo drug sensitivities. Samples were categorized based on the time since patients had received a prior therapy with AL, and geometric mean IC50s were calculated. The number of samples in each category is indicated by N. Values for each drug were compared with those for children who did not receive prior therapy within 70 days (over 70), and comparisons with P values of <0.05 (*), <0.01 (**), and <0.001 (***) in univariate analysis are labeled. With multivariate analysis, only the median piperaquine IC50 for 51 to 70 days since prior treatment was statistically significant.

FIG 5

Impact of prior therapy with AL on polymorphism prevalence. Samples were categorized based on the time since patients had received a prior therapy with AL, and prevalences of wild-type (WT), mixed (Mix), and mutant (Mut) alleles were determined. The number of samples in each category is indicated by N. Prevalences for each polymorphism were compared with those for children who did not receive prior therapy within 70 days (over 70), and comparisons with P values of <0.05 (*), <0.01 (**), and <0.001 (***) in univariate analysis are labeled. With multivariate analysis, the labeled comparisons remained statistically significant.

FIG 6

Impact of chemoprevention regimen on polymorphisms. The prevalences of wild-type (WT), mixed (Mix), and mutant (Mut) alleles are shown for parasites isolated from subjects in the different study arms. The number of samples in each category is indicated by N. For the DP arm, results for all subjects (DPall) and those classified based on drug-level analysis as DPlow (PQ levels, <10 ng/ml), DPmed (PQ levels, 10 to <20 ng/ml), and DPhigh (PQ levels, >20 ng/ml) are shown. Studied polymorphisms with significant differences in prevalences of WT alleles between samples from children receiving the indicated chemoprevention regimen and those not receiving chemoprevention are labeled for P values of <0.05 (*) and <0.01 (**). With multivariate analysis, the labeled comparisons remained statistically significant.