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Background: In 2006, the National Malaria Control Program (NMCP) in Mali recommended artemisinin-based combination therapy (ACT) as the first-line treatment for uncomplicated malaria. Since the introduction of ACT, few reports are available on the level of resistance of Plasmodium falciparum (P. falciparum) to antimalarial drugs in Mali. Dihydroartermisinin is the active metabolite of artemisinin derivatives. Here, we conducted an ex-vivo drug sensitivity testing in a rural area of southern Mali, namely the Kéniéroba village from 2016 to 2017. Methods: Seventy-five (75) isolates of P. falciparum were successfully evaluated for ex-vivo sensitivity to key anti-malarial drugs, namely chloroquine (CQ), quinine (QN), amodiaquine (AQ), mefloquine (MQ), lumefantrine (LUM), dihydroartermisinin (DHA) , and piperaquine (PPQ). P. falciparum sensitivity to these drugs was assessed using the World Wide Antimalarial Resistance Network (WWARN) SYBR-GREEN method of inhibitory concentration of 50% (IC50) determination. Reduced sensitivity to antimalarial drugs was defined as IC50 less than the WWARN standard IC50. Results: The proportion of resistant P. falciparum isolates was 20.2% for CQ, 40.5% for QN, 6.8% for AQ, and 1.3% for MQ. All tested P. falciparum isolates were sensitive to LUM, DHA, and PPQ. A statistically significant correlation was found between QN and AQ IC50 values (r = 0.80; r2 = 0.64, P