Temporal Prevalence of Plasmodium Falciparum Drug Resistance Markers, Pfcrt (K76T), PFMDR1 (N86Y), PFDHFR (C59R) and PFDHPS (K540E) in Kanyawegi Sub-Location, Western Kenya

Abstract

Malaria remains one of the most deadly infectious disease in sub-Saharan Africa accounting for high rates of mortality and morbidity especially in children less than 5 years. Plasmodium falciparum (Pf) parasite causes the most virulent form of malaria partially due to development of high levels of resistance against most anti-malaria drugs used. In malaria holoendemic areas of western Kenya, P. falciparum drug resistance has been noted since 1980s. Various genetic mutations have been identified and associated with P. falciparum resistance to drugs, such as chloroquine (CQ), sulphadoxinepyrimethamine (SP) and recently, Artemisinin-based combination therapy (ACT). Even though previous studies in Kanyawegi in western Kenya have addressed the presence of the genes associated with antimalarial resistance at one time point, the genes associated with ACT resistance in this malaria endemic population remains unknown. As such the temporal stability in the prevalence of pfcrt (K76T), pfmdr1 (N86Y), pfdhfr (C59R) and pfdhps (K540E), and the association within and between the mutations that confer resistance against anti-malarials and those considered predictive of ACT treatment failure was investigated over three time points (July 2004, August 2004 and July 2009). A total of 95 paired retrospective blood samples confirmed positive for P. falciparum in a previous study at Chulaimbo Health Centre in western Kenya were used. Using Polymerase Chain Reaction (PCR) and Nested PCR, the genes pfcrt K76T, pfmdr1 N86Y, pfdhfr (C59R) and pfdhps (K540E) were amplified and presence of mutations determined by gel electrophoresis after Restriction Fragment Length Polymorphism (RFLP). The prevalence of Pfdhps wild type 540K increased from 14.7% (n =14/95) in July 2004 to 53.7% (n=51/95) in August 2004 (p=0.0004) and subsequently to 94.1% (16/17) in July 2009 (p=0.015). For Pfdhfr wild type C59, the prevalence in July 2004 was 0.0% (n =0/95) and 1.1% (n=1/95) (p=0.144) in August 2004 and 0.0% (n = 0/17) in July 2009. The prevalence of Pfmdr1 wild type 86N was insignificant (p=0.223) from 0% (n=0/95) in July 2004 to 2.1% (n =2/95) in August 2004 and reduced to 0% (n = 0/17) in July 2009 (p = 0.759). Prevalence of Pfcrt wild type 76T remained constant in July and August 2004 at 2.1% (n=2/95) and rose to 5.9% (n=1/17) in 2009 (p=0.138). There was a negative correlation between the two SP resistant markers (r=-0.162, p=0.025) with the prevalence of pfdhps (K540E) increasing us pfdhfr (C59R) decreasing. Findings presented here suggest that resistant markers against CQ and SP have not faded and as such not recommended antimalarials in this P.falciparum holoendemic region. This study will complement existing data on anti-malarial drug resistance monitoring and enhance future prediction of resistance levels that would be critical in informing ant-malarial drug policy aimed at reducing malaria-related morbidity and mortality.