Assessment of Artemisinin Tolerance in Plasmodium Falciparum Field Isolates in Children with Uncomplicated Malaria in a Ghanaian Population

Background Artemisinin-based combination therapies (ACTs) are the first-line treatments for uncomplicated malaria in Ghana. Artemisinin (ART) tolerance in Plasmodium falciparum has arisen in Southeast Asia (SEA) and recently, in parts of East Africa. This is ascribed to the survival of ring-stage parasites post treatment. We sought to assess and characterize correlates of potential ART tolerance based on post-treatment parasite clearance, ex vivo and in vitro drug vulnerability, and molecular markers of drug resistance in P. falciparum isolates from children with uncomplicated malaria in Ghana. Methods Six months to fourteen years old children having uncomplicated malaria (n = 115) were enrolled from two hospitals and a health center in Ghana’s Greater Accra region and treated with artemether-lumefantrine (AL) according to body weight. Pre- and post-treatment parasitemia (day 0 and day 3) was confirmed by microscopy. The ex vivo ring-stage survival assay (RSA) was used to detect percent ring survival while the 72 hr SYBR Green 1 assay was used to measure the 50% inhibition concentration (IC50s) of ART and its derivatives and partner drugs. Genetic markers of drug tolerance /resistance were evaluated using selective whole genome sequencing. Results Of the total of 115 participants, 85 were successfully followed up on day 3 post-treatment and had 2/85 (2.4%) parasitemia. The IC50 values of ART, artesunate (AS), artemether (AM), dihydroartemisinin (DHA), amodiaquine (AQ), and lumefantrine (LUM) were not indicative of drug tolerance. However, 7/90 (7.8%) pre-treatment isolates had > 10% ring survival rates against DHA. Of the four isolates (2 RSA positive and 2 RSA negative) with high genomic coverage, P. falciparum kelch 13 K188* and Pfcoronin V424I mutations were only present in the two RSA positive isolates with > 10% ring survival rates. Conclusions The observed low proportion of participants with post-treatment parasitemia suggests AL remains efficacious. However, the increased rates of survival observed in the ex vivo RSA against DHA, maybe a pointer of an early start of ART tolerance. Furthermore, the role of two novel mutations in PfK13 and Pfcoronin genes, harbored by the two RSA positive isolates that had high ring survival in our study, remains to be elucidated.