Background: Understanding the human malaria infectious reservoir is important for elimination initiatives. Here, we implemented mosquito membrane feeding experiments to prepare for larger studies to quantify the transmission potential and relative contribution of the human infectious reservoir. Methods: Patients with clinical malaria attending four health facilities with at least 16 Plasmodium falciparum gametocytes per μL were recruited during the 2018 transmission season. Infectiousness to mosquitoes was assessed by direct membrane feeding assay (DMFA). We compared our results with a Bayesian predictive model to investigate the relationship between infectiousness and gametocyte density and explore the impact of fever on gametocyte infectivity. Results: A total of 3177 suspected malaria cases were screened; 43.3% (1376) had microscopically patent P. falciparum parasites and 3.6% (114) of them had gametocytes. Out of 68 DMFAs, 38 (55.9%) resulted in at least one infected mosquito, with a total of 15.4% (1178/7667) of mosquitoes infected with 1-475 oocysts per gut. The relationship between mosquito infection prevalence and gametocytaemia was similar to other African settings and negatively associated with fever (OR: 0.188, 95% CI 0.0603 to 0.585, p=0.0039). Conclusions: Among symptomatic malaria patients, fever is strongly associated with transmission failure. Future studies can use DMFA to better understand the human malaria reservoir in settings of low endemicity in The Gambia and inform malaria elimination initiatives.