Document Type : Original Article

Author

Gombe State University, Gombe

10.22034/ijabbr.2022.550156.1386

Abstract

Malaria Chemoprevention depend on synthetic drugs, but the parasite is continuously developing resistance to the antimalarial armament, hence a consequential need for surveillance studies on the sensitivity of the drugs. Therefore, the aim of this paper was to determine the presence of biomarkers associated with drug sensitivity in DHFR and DHPS gene of Plasmodium falciparum.200 blood samples were collected using vein puncture technique and analysed using Microscopy, RDT and PCR. DNA was extracted using Quick-DNA™ Miniprep extraction kit, Purity and Concentration of the DNA were determined using Nanodrop Spectrophotometer. 57 samples were selected for molecular analysis. Nested PCR was used to amplify PFDHFR and PFDHPS genes, all PCR reactions were carried out in 25µl reaction mixture (5µlDNA template, 1µlPrimer, 6.5µldistilled water and 12.5µlMaster mix). For the primary PCR of DHFR gene the cyclic conditions were; Initial denaturation 940C,90s, denaturation 940C,30s, annealing 590C,30s, extension 670C,45s and final extension 680C,5minutes, while for the secondary PCR were 940C,90seconds, 940C,30secods, 550C,30secods, 680C 45seconds and 680C,5minutes. For the primary PCR of DHPS gene, the cyclic conditions were; 940C,90s, 940C,30s, 570C,30secods, 680C,45s and 680C,5minutes, while for Secondary PCR were 940C,90seconds, 940C,30s, 570C,30s, 680C,45s and 680C,5minutes. The PCR products were subjected to electrophoresis using 2% agarose gel. The amplicons were purified, sequenced and subjected to BLAST software. Mutations were recorded from A16V 05(8.77%), N51I 18(31.58%), C59R 03(5.26%), I164L 12(21.05) variants of DHFR gene, while in DHPS gene mutation were recorded from K540E 6(10.52%) variant. Basic Biomarkers of resistance in DHFR and DHPS gene were recorded from Gombe.

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