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Background  Drug resistance is one of the greatest challenges of malaria control programme in Mali. Recent advances in next-generation sequencing (NGS) technologies provide new and effective ways of tracking drug-resistant malaria parasites in Africa. The diversity and the prevalence of  Plasmodium falciparum  drug-resistance molecular markers were assessed in Dangassa and Nioro-du-Sahel in Mali, two sites with distinct malaria transmission patterns. Dangassa has an intense seasonal malaria transmission, whereas Nioro-du-Sahel has an unstable and short seasonal malaria transmission.    Methods  Up to 270 dried blood spot samples (214 in Dangassa and 56 in Nioro-du-Sahel) were collected from  P. falciparum  positive patients in 2016. Samples were analysed on the Agena MassARRAY ®  iPLEX platform. Specific codons were targeted in  Pfcrt ,  Pfmdr1 ,  Pfdhfr , and  Pfdhps, Pfarps10, Pfferredoxin, Pfexonuclease  and  Pfmdr2  genes. The Sanger’s 101-SNPs-barcode method was used to assess the genetic diversity of  P. falciparum  and to determine the parasite species.    Results  The  Pfcrt _76 T  chloroquine-resistance genotype was found at a rate of 64.4% in Dangassa and 45.2% in Nioro-du-Sahel ( p  = 0.025). The  Pfdhfr_51I - 59R - 108N  pyrimethamine-resistance genotype was 14.1% and 19.6%, respectively in Dangassa and Nioro-du-Sahel. Mutations in the  Pfdhps  _S436 - A437 - K540 - A581 - 613A  sulfadoxine-resistance gene was significantly more prevalent in Dangassa as compared to Nioro-du-Sahel ( p  = 0.035). Up to 17.8% of the isolates from Dangassa  vs  7% from Nioro-du-Sahel harboured at least two codon substitutions in this haplotype. The amodiaquine-resistance  Pfmdr1 _N86Y mutation was identified in only three samples (two in Dangassa and one in Nioro-du-Sahel). The lumefantrine-reduced susceptibility  Pfmdr1_Y184F  mutation was found in 39.9% and 48.2% of samples in Dangassa and Nioro-du-Sahel, respectively. One piperaquine-resistance  Exo  _E415G  mutation was found in Dangassa, while no artemisinin resistance genetic-background were identified. A high  P. falciparum  diversity was observed, but no clear genetic aggregation was found at either study sites. Higher multiplicity of infection was observed in Dangassa with both COIL ( p  = 0.04) and Real McCOIL ( p  = 0.02) methods relative to Nioro-du-Sahel.    Conclusions  This study reveals high prevalence of chloroquine and pyrimethamine-resistance markers as well as high codon substitution rate in the sulfadoxine-resistance gene. High genetic diversity of  P. falciparum  was observed. These observations suggest that the use of artemisinins is relevant in both Dangassa and Nioro-du-Sahel.

A comprehensive analysis of drug resistance molecular markers and Plasmodium falciparum genetic diversity in two malaria endemic sites in Mali