Major genome region underlying artemisinin resistance in malaria parasite isolates from Kisumu County, Western Kenya.

Abstract

Clinical resistance to artemisinin and its derivatives is now well established in Southeast Asian (SEA). Studies from SEA have identified single nucleotide polymorphisms (SNPs) which are significantly associated with delayed parasite clearance rates. In this study we screened SNPs and established the SNP profile in Kenyan isolates. DNA was extracted from 129 Plasmodium falciparum (P. falciparum) positive patients collected in Kisumu from 2013-2014 and 1995-2010. 25 SNPs were genotyped by Sequenome MassARRAY. P. falciparum multi-drug resistance gene 1 (pfmdr1) copy number variation was quantified using real time PCR. The in vitro artemether susceptibility of the isolates was analyzed by malaria SYBR Green I assay. Microsatellite analysis of MSP1/MSP2 was used to establish recrudescent verses new infections in 20 follow up samples obtained from the efficacy study. Data showed that pre-ACT samples contained 60% of the SNPs analyzed while the Post-Act samples had 24% of the SNPs. One Pre-ACT Sample (1.4%) contained multiple pfmdr1 copy number and 7 of post ACT sample (9.9%) contained >1 pfmdr1 copy number. There was a significant increase in Pfmdr1 copy number between pre-ACT and post-ACT samples (P<0.001). The average clearance half-life was 2.6 hours. All parasites were sensitive to artemisinin with significance difference in clearance half-life within subject (p= 0.0001). There was no significant association between clearance half-life and genotypes (p> 0.05). In vitro susceptibility to piperaquine reduced for isolates with multiple pfmdr1 copy number to piperaquine and amodiaquine (P< 0.04). In conclusion there is a possibility of selective sweep of SNP after elimination of previous drug pressure and introduction of ACT. Pfmdr1 may putatively play a role in ACT resistance.