Molecular Epidemiology of Plasmodium falciparum kelch13 Mutations in Senegal Determined by Using Targeted Amplicon Deep Sequencing

Abstract

The emergence of Plasmodium falciparum resistance to artemisinin in Southeast Asia threatens malaria control and elimination activities worldwide. Multiple polymorphisms in the P. falciparum kelch gene found in chromosome 13 (Pfk13) have been associated with artemisinin resistance. Surveillance of potential drug resistance loci within a population that may emerge under increasing drug pressure is an important public health activity. In this context, P. falciparum infections from an observational surveillance study in Senegal were genotyped using targeted amplicon deep sequencing (TADS) for Pfk13 polymorphisms. The results were compared to previously reported Pfk13 polymorphisms from around the world. A total of 22 Pfk13 propeller domain polymorphisms were identified in this study, of which 12 have previously not been reported. Interestingly, of the 10 polymorphisms identified in the present study that were also previously reported, all had a different amino acid substitution at these codon positions. Most of the polymorphisms were present at low frequencies and were confined to single isolates, suggesting they are likely transient polymorphisms that are part of naturally evolving parasite populations. The results of this study underscore the need to identify potential drug resistance loci existing within a population, which may emerge under increasing drug pressure.

Keywords: Plasmodium falciparum; artemisinin resistance; kelch13; molecular epidemiology.

FIG 1

Plasmodium falciparum kelch13 propeller domain showing polymorphisms identified in Southeast Asia, Senegal, and other African countries. The 6-blade tertiary structure and 4 β strands (highlighted by different colors: β strand 1, yellow; β strand 2, pink; β strand 3, green; and β strand 4, blue) are shown for the 442- to 726-amino-acid propeller domain region. Polymorphisms observed in SEA and Africa are shown by blue and red circles, respectively. Polymorphisms observed in this study (Senegal) are indicated by green circles. Major alleles are represented by circles and minor alleles by squares. Polymorphisms confirmed to be associated with artemisinin resistance are indicated by asterisks. This figure is a recreation of a figure previously reported by the MalariaGen community project (19).

FIG 2

Plasmodium falciparum kelch13 propeller domain polymorphisms identified in Senegal during the malaria transmission seasons in 2011 and 2014. Pfk13 propeller domain polymorphisms identified in this population are shown along with their respective sequence read frequencies (>2%). Each bar corresponds to an individual patient sample. Polymorphisms not previously reported in Africa (shown in blue), along with previously identified polymorphisms, are shown. Codon positions that have been confirmed to confer artemisinin resistance are indicated by #.