Little Polymorphism at the K13 Propeller Locus in Worldwide Plasmodium falciparum Populations Prior to the Introduction of Artemisinin Combination Therapies

Abstract

The emergence and spread of artemisinin-resistant Plasmodium falciparum is of huge concern for the global effort toward malaria control and elimination. Artemisinin resistance, defined as a delayed time to parasite clearance following administration of artemisinin, is associated with mutations in the Pfkelch13 gene of resistant parasites. To date, as many as 60 nonsynonymous mutations have been identified in this gene, but whether these mutations have been selected by artemisinin usage or merely reflect natural polymorphism independent of selection is currently unknown. To clarify this, we sequenced the Pfkelch13 propeller domain in 581 isolates collected before (420 isolates) and after (161 isolates) the implementation of artemisinin combination therapies (ACTs), from various regions of endemicity worldwide. Nonsynonymous mutations were observed in 1% of parasites isolated prior to the introduction of ACTs. Frequencies of mutant isolates, nucleotide diversity, and haplotype diversity were significantly higher in the parasites isolated from populations exposed to artemisinin than in those from populations that had not been exposed to the drug. In the artemisinin-exposed population, a significant excess of dN compared to dS was observed, suggesting the presence of positive selection. In contrast, pairwise comparison of dN and dS and the McDonald and Kreitman test indicate that purifying selection acts on the Pfkelch13 propeller domain in populations not exposed to ACTs. These population genetic analyses reveal a low baseline of Pfkelch13 polymorphism, probably due to purifying selection in the absence of artemisinin selection. In contrast, various Pfkelch13 mutations have been selected under artemisinin pressure.

Limited genetic diversity in the PvK12 Kelch protein in Plasmodium vivax isolates from Southeast Asia

“…Therefore, without clinical resistance of P. vivax parasites to ART drugs, the point mutations observed in the world P. vivax populations could well represent background mutations. This finding is consistent with the low-level baseline mutations identified by Mita et al in P. falciparum populations before the deployment of ACT [59]. …”

Malaria epidemiological research in the Republic of Congo

“…A total of eight articles have been found reporting data on molecular markers of P. falciparum resistance to anti-malarials with samples collected from 1999 to 2015 [11, 15, 23–25, 31, 32, 36]. Overall, six molecular markers have been studied: the P. falciparum chloroquine resistance transporter ( pfcrt ) gene, associated with chloroquine resistance, the dihydrofolate reductase ( dhfr ) and the dihydropteroate synthase ( dhps ) genes, which are linked with pyrimethamine resistance and sulfadoxine resistance, respectively, the P. falciparum Klech-13 (K13) propeller gene, which has recently been linked with artemisinin resistance, and MAL10-688956 and MAL13-1718319 single nucleotide polymorphisms (SNPs) which have also recently been proposed as molecular markers of artemisinin resistance which is defined as a delayed clearance of P. falciparum parasites following ACT.…”

“…The K13 propeller gene was characterized by Mita et al [32], and MAL10-688956 and MAL13-1718319 SNPs have been characterized by Murai et al [31]. In both studies they used the same set of archived P. falciparum isolates collected in 2005–2006 in Brazzaville, Pointe-Noire and Gamboma, and none of the mutations associated with artemisinin resistance was found.…”

“…Overall in vivo efficacy studies conducted so far provided evidence of good efficacy of currently recommended ACT in RoC. It is observed that only one study analysed mutations on P. falciparum K13 propeller gene in isolates from RoC [32], and it would be important that local scientists screen regularly isolates from different parts of the country. Chloroquine and SP were banned for the treatment of uncomplicated malaria in the RoC since 2006 due to high level of parasite resistance.…”

Polymorphisms in Plasmodium falciparum Kelch 13 and P. vivax Kelch 12 Genes in Parasites Collected from Three South Pacific Countries Prior to Extensive Exposure to Artemisinin Combination Therapies

“…The third study examined Kelch 13 mutations in 581 isolates from 14 countries, mostly before the introduction of ACTs (19). The number and frequency of mutations observed was low in pre-ACT compared to post-ACT samples, although post-ACT samples came mainly from three Southeast Asian countries where artemisinin resistance has been reported.…”