High frequency of artemisinin partial resistance mutations in the great lake region revealed through rapid pooled deep sequencing

Abstract

In Africa, the firstPlasmodium falciparumKelch13 (K13) artemisinin partial resistance mutation 561H was first detected and validated in Rwanda. Surveillance to better define the extent of the emergence in Rwanda and neighboring countries as other mutations arise in East Africa is critical. We employ a novel scheme of liquid blood drop preservation combined with pooled sequencing to provide a cost-effective rapid assessment of resistance mutation frequencies at multiple collection sites across Rwanda and neighboring countries. Malaria-positive samples (n=5,465) were collected from 39 health facilities in Rwanda, Uganda, Tanzania, and the Democratic Republic of the Congo (DRC) between May 2022 and March 2023 and sequenced in 199 pools. In Rwanda, K13 561H and 675V were detected in 90% and 65% of sites with an average frequency of 19.0% (0-54.5%) and 5.0% (0-35.5%), respectively. In Tanzania, 561H had high frequency in multiple sites while it was absent from the DRC although 675V was seen at low frequency. Conceringly candidate mutations were observed: 441L, 449A, and 469F co-occurred with validated mutations suggesting they are arising under the same pressures. Other resistance markers associated with artemether-lumefantrine are common:P. falciparummultidrug resistance protein 1 N86 at 98.0% and 184F at 47.0% (0-94.3%) andP. falciparumchloroquine resistance transporter 76T at 14.7% (0-58.6%). Additionally, sulfadoxine-pyrimethamine-associated mutations show high frequencies. Overall, K13 mutations are rapidly expanding in the region further endangering control efforts with the potential of engendering partner drug resistance.

Temporal genomic analysis ofPlasmodium falciparumreveals increased prevalence of mutations associated with delayed clearance following treatment with artemisinin-lumefantrine in Choma District, Southern Province, Zambia

“…We also identified the common K189 T polymorphism in Africa that is not associated with resistance in 13.2% (95% CI, 9.5–17.4) of samples. The Kelch 13 mutations R561 H , A675 V , and C469 Y recently reported in Rwanda, Tanzania, and Uganda 31 were not identified. None of the samples carried mutations that were associated with the genetic backbone for ART-R in south-east Asia such as apicoplast ribosomal protein s10 (ARPS10 V127 M ), ferredoxin (FD D193 Y ), P. falciparum multidrug resistance 2 transporter (MDR2 T484 I ), putative phosphoinositide-binding protein (PIB7 C1484 F ), P. falciparum protein phosphatase (PPH V1157 L ), and P. falciparum chloroquine resistance transporter (CRT N326 S and CRT I356 T ).…”

National genomic profiling ofPlasmodium falciparumantimalarial resistance in Zambian children participating in the 2018 Malaria Indicator Survey

“…51 Considering the historical use and importance of ART to malaria control in Zambia, increased polymorphisms in the kelch13 gene and the identification three WHO-validated mutations associated with partial artemisinin resistance suggest an early signal of partial artemisinin resistance in Zambia. Close monitoring of local emerging or spreading kelch13 mutations (R561H, A675V and C469Y) [52][53][54] that were recently reported from East Africa and confer partial artemisinin resistance is warranted. While not unexpected, it was also encouraging to note that no mutations (Pfcrt I356T, Pffd D193Y, Pfmdr2 T484I, Pfap2mu S160N and Pfubp1 E1528D) associated with ART resistance in Southeast Asia 55 were identified.…”