Absence of Plasmodium falciparum artemisinin resistance gene mutations eleven years after the adoption of artemisinin-based combination therapy in Nigeria

Abstract

Background

The occurrence of artemisinin resistance (ART)-associated polymorphism of Plasmodium falciparum K13-propeller ( pfk13) gene before and after the introduction of artemisinin-based combination therapy (ACT) in two regions of Nigeria was investigated in this study. Regular surveillance is necessary to make a definite conclusion on the emergence and pattern of possible resistance to ART.

Methods

This cross-sectional study was carried out in the Southwestern and Southeastern geopolitical zones of Nigeria. A total of 150, 217, and 475 participants were enrolled for the study in the Southwest (2004_Group A), Southwest (2015_Group B), and southeast (2015_Group C), respectively. Blood samples were collected from the study participants for DNA extraction and a nested PCR for P. falciparum identification. Samples that were positive for P. falciparum were genotyped for the pfk13 gene using the Sanger sequencing method. The single nucleotide polymorphisms were analysed using the Bioedit software.

Results

A total of 116, 125, and 83 samples were positive for P. falciparum, respectively for the samples collected from the Southwest (2004 and 2015) and southeast (2015). Parasite DNA samples collected from febrile children in 2004 (Group A; n = 71) and 2015 (Group B; n = 73) in Osogbo Western Nigeria and 2015_Group C (n = 36) in southeast Nigeria were sequenced successfully. This study did not observe mutations associated with the in vitro resistance in southeast Asia, such as Y493H, R539T, I543T, and C580Y. Two new polymorphisms V520A and V581I were observed in two samples collected in Osogbo, Southwest Nigeria. These two mutations occurred in the year 2004 (Group A) before the introduction of ACT. Six mutations were identified in 17% of the samples collected in southeast Nigeria. One of these mutations (D547G) was non-synonymous, while the remaining (V510V, R515R, Q613Q, E688E, and N458N) were synonymous. Also, one (2%) heterozygote allele was identified at codon 458 in the 2015 (Group C) samples.

Conclusions

None of the mutations observed in this study were previously validated to be associated with ART resistance. These results, therefore, suggest that artemisinin is likely to remain highly effective in treating malaria in the study areas that are malarious zone.