Gametocyte production and infectivity among Ugandan malaria patients infected with P. falciparum with partial resistance to artemisinins

BackgroundPartial resistance to artemisinins (ART-R) has emerged in East Africa, associated with mutations in the Plasmodium falciparum kelch13 gene. It is currently unclear whether ART-R has implications for gametocyte production or for onward transmission to mosquitoes. MethodsIn a cohort of uncomplicated malaria patients attending Kalongo Hospital in northern Uganda, we quantified carriage of PfKelch13 mutant parasites by conventional sequencing and droplet digital PCR (ddPCR) for the C469Y and A675V mutations. Prevalence and density of gametocytes and ring-stage parasites were assessed by microscopy and quantitative reverse-transcriptase PCR (qRT-PCR). Lumefantrine concentrations, indicative of prior malaria treatment, were determined by ultra-high performance liquid chromatography-tandem mass spectrometry. Transmission potential of wild-type and PfKelch13 mutant parasites was assessed by mosquito feeding assays and complemented with molecular characterization of parasites in wild-caught mosquitoes from household resting catches. FindingsWe enrolled 235 patients with symptomatic P. falciparum infection; PfKelch13 C469Y or A675V mutations were detected in 35.8% (78/218) of infections by sequencing and 59.1% (136/230) by ddPCR. Gametocyte carriage was 24.0% (56/233) by microscopy and 56.6% (133/235) by qRT-PCR and not associated with the abundance of PfKelch13 mutant parasites by ddPCR (p=0.603). Among a total of 227 mosquito feeds with patient whole blood, 1.4% (120/8745) of mosquitoes became infected. Mosquito infection rates were positively associated with gametocyte density ({beta} = 0.39, 95% CI = 0.23-0.59, p < 0.001) without an observed interaction with the abundance of PfKelch13 mutant parasites (p = 0.452). PfKelch13 C469Y or A675V mutations were detected in 40.1% (21/52) of malaria-infected bloodmeals of field-caught mosquitoes and in 28.0% (7/25) of sporozoite-positive mosquitoes. InterpretationWe conclude that pfkelch13 mutations are very common in patients in northern Uganda with uncomplicated malaria, mostly in multiclonal infections. We observed no evidence that ART-R affected gametocyte production or transmission to mosquitoes. FundingDutch Research Council (NWO) Research in contextO_ST_ABSEvidence before this studyC_ST_ABSPartial resistance to artemisinins (ART-R) might enhance or inhibit gametocyte formation or transmission of malaria parasites to Anopheles mosquitoes. However, few studies directly assessed P. falciparum transmission potential in relation to sensitivity to ART-R or associated mutations. We searched PubMed on January 14th 2026, with no restrictions on publication date or language, for studies assessing gametocyte carriage or transmissibility to mosquitoes in uncomplicated malaria cases in relation to artemisinin resistance using the search terms ("uncomplicated malaria" OR "patient") AND ("gametocyte" OR "anopheles") AND (("artemisinin" AND "resistance") OR "pfkelch13" OR "kelch13"). From the 101 identified articles, the majority did not report on parasite resistance or resistance markers. Nineteen articles reported original patient data, with four additional studies examining parasite isolates for gametocyte production and transmissibility in vitro. In vitro studies all reported gametocyte formation in parasite isolates with pfkelch13 mutations and, for three studies where transmission was directly determined, evidence for successful mosquito infections from ART-R parasite isolates. None of these studies demonstrated consistent differences in gametocyte production or transmissibility between ART-R and sensitive parasite isolates. One clinical study from Thailand observed higher levels of gametocyte carriage during follow-up for infections with slow asexual blood-stage parasite clearance following artemisinin-combination therapy (ACT) and an increase in gametocyte carriage at clinical presentation over a period when the prevalence of ART-R was rising. A study with patient data from 7 Asian and 3 African countries reported higher proportions of pretreatment and post-treatment gametocytemia in patients with slow parasite clearance. Another study from Cambodia reported a higher prevalence of gametocytes at enrolment in areas affected by ART-R, but no association between gametocyte carriage and either individual-level parasite clearance or treatment failure. In contrast, a meta-analysis of ACT treatment efficacy in pregnant women observed no association between gametocyte carriage at baseline and treatment failure. Only one study, directly examined gametocyte carriage at clinical presentation with pfkelch13 mutations and found no association between gametocyte carriage and molecular markers of ART-R or ex vivo drug sensitivity in Cambodia. Added value of this studyWe performed a direct assessment of gametocyte production, carriage of mature gametocytes and transmission to Anopheles gambiae s.s. mosquitoes in relation to validated ART-R markers in patients presenting with uncomplicated P. falciparum malaria in an area affected by ART-R in Uganda. Our work demonstrates that parasites with the PfKelch13 C469Y and A675V mutations were very common, mostly in multiclonal infections. Gametocyte carriage and gametocyte commitment were similar between patients presenting with pure or predominantly wild-type infections and those presenting with PfKelch13 mutant infections. In feeding studies, mosquito infection rates were similar for patients with wild-type or PfKelch13 mutant infections; wild-caught mosquitoes confirmed ongoing transmission of parasites with molecular signatures of ART-R. Implications of all the available evidenceTaken together, there are no indications for altered gametocyte production or transmissibility to mosquitoes of infections with PfKelch13 C469Y and A675V. Future studies may examine impacts of other mutations in pfkelch13, study transmission potential in low endemic settings where monoclonal infections dominate and aim to understand how gametocyte clearance and post-treatment transmission potential may differ between ART-R and wild-type infections upon treatment.