Genetic Polymorphisms in<i> Plasmodium falciparum </i>Chloroquine Resistance Gene<i>, pfcrt </i>in Massakory (Chad)

Abstract

<i>Background and Objective</i>: In 2005, Chad, like several other WHO countries, withdrew chloroquine as a first-line treatment for <i>Plasmodium falciparum</i> malaria in response to WHO recommendations related to the reason for the increase in treatment failures and the global spread of chloroquine resistance. Artemisinin-based combination therapy (ACTs), Artemether-lumefantrine, has replaced chloroquine as the first-choice treatment for malaria. The present study assessed <i>pfcrt</i> polymorphism in <i>Plasmodium falciparum</i> isolates in Massakory. <i>Methodology and Results</i>: Blood samples for PCR analysis were collected on Whatman 3MM filter paper in Massakory during a therapeutic efficacy study (TES) conducted from December 14, 2019 to March 14, 2020. Genomic DNA was extracted from 113 dried blood spots with the QIAamp DNA Micro Kit (Qiagen, Valencia, CA) as per manufacturer’s protocol and amplified by nested-PCR with <i>pfcrt</i> specific primer. The amplification products were revealed by electrophoresis on 2% agarose gel and then sequenced according to Sanger method. A total of 71 sequences were readable. The <i>pfcrt</i> analysis showed that of the 71 readable sequences, high mutation prevalence: 66 (92.96%) <I>IET</I>, 2 (4.22%) <I>IDT</I> and 3 (4.22%) <I>MNK</I> wild <i>pfcrt</i> isolates. <i>Conclusion</i>: These results challenge the highest health authorities in the country. The government, through the Ministry of Public Health and National Solidarity and the National Malaria Control Program, must raise awareness for the effective withdrawal of chloroquine. This action will promote on the one hand the re-emergence of parasites sensitive to chloroquine, and on the other hand make possible the reintroduction of chloroquine in the treatment of simple malaria after the suppression of drug pressure.