Prevalence of the Pfdhfr and Pfdhps mutations among asymptomatic pregnant women in Southeast Nigeria
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Sulfadoxine-pyrimethamine (SP) is the recommended drug for intermittent preventive treatment of malaria in pregnancy in most of sub-Saharan Africa. Resistance to SP is related to mutations in the dhfr and dhps gene of Plasmodium falciparum. This study determined the prevalence of Pfdhfr and Pfdhps polymorphisms found in asymptomatic pregnant women attending antenatal care in Calabar, Nigeria. From October 2013 to November 2014, asymptomatic pregnant women attending antenatal care clinics were enrolled after obtaining informed consent. Malaria diagnosis testing was done using thick and thin smears. Dried blood spot filter papers were collected. Parasite DNA was extracted from the filter papers using a chelex extraction. Extraction was followed by nested PCR and restriction enzyme digestion. P. falciparum infection was detected by microscopy in 7% (32/459) participants. Twenty-eight P. falciparum isolates were successfully genotyped. In the Pfdhfr gene, the triple mutation was almost fixed; S108N mutation was (100%), N51I (93%) and C59R mutations (93%), whereas the I164L mutation was absent. The prevalence of Pfdhps S436A, A437G, A581G and A613S mutations was 82.1% (23/28), 96.4% (27/28), 71.4% (20/28) and 71.4% (20/28) respectively. The K540E mutation was absent. The prevalence of the Pfdhfr triple mutation IRNI was 92.9% (26/28). The efficacy of SP as IPTp in Southeast Nigeria may be severely threatened. The continuous monitoring of SP molecular markers of resistance is required to assess thresholds. The evaluation of alternative preventive treatment strategies and drug options for preventing malaria in pregnancy may be necessary.
KeywordsMolecular markers SP resistance Malaria Antenatal clinic
dried blot spot
dihydrofolate reductase enzyme
dihydrofolate reductase gene
dihydropteroate synthetase enzyme
dihydropteroate synthetase gene
intermittent preventive treatment in pregnancy
insecticide-treated bed net
polymerase chain reaction-restriction fragment length polymorphism
World Health Organization
We thank the study participants, the staff of the antenatal clinic and management of the General Hospital Calabar where this study took place. Special thanks to Dr. Olabisi Oduwole, Ms.Obiamaka Okafo and Mr. David Agamse at the Calabar Institute of Tropical Diseases Research and Prevention, University of Calabar Teaching Hospital, Calabar, Nigeria, for their invaluable help. We acknowledge the assistance of Prof Dr. Frank Mockenhaupt at the Institute of Tropical Medicine and International Health, Charité-Universitätsmedizin Berlin, with genotyping of molecular markers. We are also grateful to the Center for International Health (CIH), Ludwig-Maximilians University of Munich, DAAD, BMZ and Exceed for supporting the first author for his PhD program.
EE, MM, NB, MP and TL designed the study. CT and PG were responsible for the laboratory work. EE wrote the first draft of the paper. All authors contributed to the interpretation of the data and the revision of the manuscript.
Compliance with ethical standards
Consent for publication
Availability of data and materials
The datasets generated during the current study are available from the corresponding author on reasonable request.
Conflict of interest
The authors declare that they have no competing interests.
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