Assessment of plasmodium species prevalence, antimalarial drug resistance genes and gametocytes in malaria infectionsbetween 2018 and 2021 in Kombewa sub-county, Kenya

Abstract

In Western Kenya, malaria is a major cause of morbidity and mortality with more than 70% of the population at risk.Its decline over the last 25 years indicates a shift to older children majority of who are asymptomatic. Kenya uses artemisinin combination therapy (ACT) specifically artemether-lumefantrine (AL) as first line for treatment of malaria infections and sulfadoxine-pyrimethamine (SP) as prophylaxis in pregnant women. However, delayed parasite clearance has been observed in Kenya. Resistance to antimalarials specifically ACT‟s and SP is associated with various polymorphisms in P. falciparum multidrug resistance gene 1 (Pfmdr1), multidrug resistance-associated protein 1 (Pfmrp1), dihydrofolate-reductase (Pfdhfr), dihydropteroate-synthase (Pfdhps), and chloroquine resistance transporter (Pfcrt) genes. While symptomatic malaria infections are recognized and treated, recent reports have revealed a largeproportion of asymptomaticinfections.A recent study in Kombewa indicated that some asymptomatic participants did not clear parasites or disrupt transmission in a large proportion of study population despite adequate treatment. Given the limited data on drug resistance in asymptomatic infections, molecular epidemiological studies of drug resistance are required to assess these infections. The first objective was to determine Plasmodium species prevalence (P.falciparum, P.malarie, P.ovale wallikeri, P.ovale curtisi- Pf, Pm, Pow, Poc), second objective was to determine frequency of anti-malarial resistance gene polymorphisms and third objective was toinvestigate gametocyte variability in symptomatic and asymptomatic infections. In a retrospective cross-sectional study molecular techniqueswere used toanalyze 230 archived whole blood samples collected between 2018 and 2021 in Kombewa under malaria epidemiology surveillance and malaria transmission study representing symptomatic and asymptomatic infection. The species composition(Pf, Pm, Pow, Poc) and gametocyte carriage (Pfs16, Pfs25) were determined using real-time polymerase chainreaction and analyzed using excel.Genotyping of Pfmdr1 86, 184 & 1246; Pfmrp1 437, 876 &1390; Pfdhfr 16, 22, 59 & 164; Pfdhps 436, 437 & 581, and Pfcrt 72, 76, 271, 326, 356 single nucleotide polymorphisms (SNPs)were assayed using Mass ARRAY platform and analyzed against the reference 3D7 genome. Data and statistical analysis wasdone using excel and Chi square tests in STATA.Of the 230 samples analyzed, Plasmodium species prevalence was; Pf 64.35% (148/230), Pm 26.52%, (61/230), Pow 9.57% (22/230), Poc 6.09% (14/230).The symptomatic Pf comprised 70.59% (24/34), Pm 17.65% (6/34), Poc 11.76% (4/34), and Pow 8.82% (3/34) while for asymptomatic Pf 63.27% (63/196), Pm 28.06% (55/196), Poc 5.1% (10/196), and Pow 9.69% (19/196). Co-infections were higherfor Pf/Pm; symptomatic 11.76% (4/34), asymptomatic 19.39% (38/196) compared to all the other species combinations (≤6%).The Pfmdr1_184 harbored symptomatic 68.75% (11/16) and asymptomatic 52% (26/50) mutations, while Pfmdr1_1246 had 6% mutants in both symptomatic (1/16) and asymptomatic (2/30).For Pfmrp1 gene codon 437 had no mutations in symptomatic while asymptomatic had only one mutation (1/30). Pfmrp1 codon 876, symptomatic reported 47.05% (8/17) & asymptomatic 37.93% (11/29). Pfmrp1 1390 symptomatic had 6.67% (1/15) and asymptomatic 6.9% (2/29) mutations respectively. Pfdhfr codons 16 and 22 had no mutations for symptomatic and asymptomatic. Pfdhfr 59 & 164 had 88.24% (15/17) and 90.91% (30/33) mutants for symptomatic and asymptomatic respectively. For both symptomatic and asymptomatic Pfdhps codons 436, 437 and 581 did not reveal any mutants.The Pfcrt gene, codons72, 76, & 356 did not have any mutations for either symptomatic or asymptomatic. Pfcrt 326 & 371 had 3.23% (1/31) and 11.11% (4/36) mutations in asymptomatic only.Overall gametocyte carriage was 65.65% (151/230), symptomatic cases positives werePf16 85.29 (29/34); Pf25 79.41% (27/34); Pf16Pf25 93.1% (29/29) while asymptomatic had Pf16 68.88% (135/196); Pf25 67.86% (133/196) &Pf16Pf25 86.1% (124/144). Even though proportional comparisons did not reveal statistical significance, this study was critical in revealing variations of Plasmodium species prevalence, frequency of drug resistance markers, and gametocyte variability in symptomatic and asymptomatic infections. Findings highlight the need for heightened molecular surveillance and management of malaria infectionsfor timely and informed interventions in all infections.