Background The emergence of resistance against artemisinin combination treatment is a major concern for malaria control. ACTs are recommended as the rescue treatment, however, there is limited evidence as to whether treatment and re-treatment with ACTs select for drug-resistant P. falciparum parasites. Thus, the present study wanted to investigate the impact of treatment and re-treatment using artesunate-amodiaquine (ASAQ) and artemether-lumefantrine (AL) on the selection of P. falciparum multidrug resistance-1 (pfmdr1) alleles in clinical settings. Methods P. falciparum positive samples were collected from children aged 12-59 months in a clinical trial in DR Congo and Uganda. Pfmdr1 single nucleotide polymorphisms (SNPs) analysis at codons N86Y, Y184F, and D1246Y were performed at baseline and post-treatment with either AL or ASAQ as a rescue treatment using nested PCR followed by restriction fragment length polymorphism (RFLP) assays. Results The pre-treatment prevalence of Pfmdr1 N86 and D1246Y varied significantly between the sites, (p>0.001) and (p=0.013), respectively. There was borderline significant directional selection for Pfmdr1 184F in recurrent infections after treatment with AL in Uganda site (p=0.05). Pfmdr1 NFD haplotype did not significantly change in post-treatment infections after re-treatment with either AL or ASAQ. Comparison between pre-treatment and post-treatment recurrences did not indicate directional selection of Pfmdr1 N86, D1246 alleles in either in the pre-RCT, RCT and post-RCT phases in both AL and ASAQ treatment arms. Pfmdr1 86Y was significantly associated with reduced risk of AL treatment failure (RR=0.34, 95% CI:0.11-1.05, p=0.04) while no evidence for D1246 allele (RR=1.02; 95% CI: 0.42-2.47, p=1.0). Survival estimates showed that the Pfmdr1 alleles had comparable mean-time to PCR-corrected recrudescence and new infections in both AL and ASAQ treatment arms. Conclusion. We found limited impact of treatment and re-treatment with AL or ASAQ on selection for Pfmdr1 variants and haplotypes associated with resistance to partner drugs. These findings further supplement the evidence use of same or alternative ACTs as a rescue therapy for recurrent P.falciparum infections. Continued monitoring of genetic signatures of resistance is warranted to maintain the efficacy of artemisinin-based combination therapies.