Screening for antifolate and artemisinin resistance in Plasmodium falciparum clinical isolates from three hospitals of Eritrea

TL;DR

This study analyzed genetic markers for antimalarial drug resistance in malaria parasites from Eritrea, finding specific mutations linked to resistance.

Contribution

First report of specific genetic mutations in Eritrea linked to antifolate and artemisinin resistance in Plasmodium falciparum.

Findings

Pfdhfr C59R and Pfdhps K540E mutations were identified in clinical isolates from Eritrea.

No PfK13 mutations were found, indicating no artemisinin resistance in the studied population.

Eight out of nineteen samples were successfully analyzed for genetic mutations.

Abstract

Background: Antimalarial drug resistance is a major challenge hampering malaria control and elimination.
Plasmodium falciparum, the leading causative parasite species, has developed resistance to basically all antimalarials. Continued surveillance of drug resistance using genetic markers provides important molecular data for treatment policies. This study sought to verify the genetic mechanism of resistance to sulfadoxine-pyrimethamine and assess the occurrence of point mutations associated with artemisinin resistance in
P. falciparum clinical isolates from Eritrea.

Methods: Nineteen dried blood spot samples were collected from patients visiting Adi Quala, Keren and Gash Barka Hospitals, Eritrea. The patients were followed up after receiving treatment with first line artesunate-amodiaquine. Nested polymerase chain reaction and Sanger sequencing techniques were employed to genotype point mutations in the
P. falciparum bifunctional dihydrofolate reductase-thymidylate synthase (
Pfdhfr, PF3D7_0417200), dihydropteorate synthase (
Pfdhps, PF3D7_0810800) and kelch 13 (
PfK13, PF3D7_1343700) genes.

Results: Eight of nineteen (42%) of the dried blood spot samples were successful for PCR-amplification. Data analyses of the PCR-positive isolates revealed the following point mutations:
Pfdhfr N51I in four isolates, C59R in one isolate, S108N in four isolates, a rare non-synonymous substitution V45A in four isolates and
Pfdhps K540E in four isolates. No
PfK13 point mutations were reported.

Conclusions:
Pfdhfr C59R and
Pfdhps K540E point mutations are reliable markers for the sulfadoxine-pyrimethamine quintuple mutant haplotype combination. These findings highlight first reports in Eritrea, which verify the underlying genetic mechanism of antifolate resistance. Continuous monitoring of the
PfK13 marker is recommended.

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