Mwesigwa, AlexGolumbeanu, MonicaJones, SamCantoreggi, L. SaraMusinguzi, BensonNankabirwa, I. JoaniterBikaitwoha, Everd ManipleKalyango, N. JoanKaramagi, CharlesPlucinski, MateuszNsobya, L. SamuelNsanzabana, ChristianByakika-Kibwika, Pauline2025-02-062025-02-062025Mwesigwa, A., Golumbeanu, M., Jones, S., Cantoreggi, S. L., Musinguzi, B., Nankabirwa, J. I., Bikaitwoha, E. M., Kalyango, J. N., Karamagi, C., Plucinski, M., Nsobya, S. L., Nsanzabana, C., & Byakika-Kibwika, P. (2025). Assessment of different genotyping markers and algorithms for distinguishing Plasmodium falciparum recrudescence from reinfection in Uganda. Springer Nature, 15, 4375.ht t p s : / / d o i . o r g / 1 0 . 1 0 3 8 / s 4 1 5 9 8 - 0 2 5 - 8 8 8 9 2 - 7 .https://doi.org/10.1038/s41598-025-88892-7http://hdl.handle.net/20.500.12493/2874Antimalarial therapeutic efficacy studies are vital for monitoring drug efficacy in malaria-endemic regions. The WHO recommends genotyping polymorphic markers including msp-1, msp-2, and glurp for distinguishing recrudescences from reinfections. Recently, WHO proposed replacing glurp with microsatellites (Poly-α, PfPK2, TA1). However, suitable combinations with msp-1 and msp- 2, as well as the performance of different algorithms for classifying recrudescence, have not been systematically assessed. This study investigated various microsatellites alongside msp-1 and msp-2 for molecular correction and compared different genotyping algorithms across three sites in Uganda. Microsatellites 313, Poly-α, and 383 exhibited the highest diversity, while PfPK2 and Poly-α revealed elevated multiplicity of infection (MOI) across all sites. The 3/3 match-counting algorithm classified significantly fewer recrudescences than both the ≥ 2/3 and Bayesian algorithms at probability cutoffs of ≥ 0.7 and ≥ 0.8 (P < 0.05). The msp-1/msp-2/2490 combination identified more recrudescences using the ≥ 2/3 and 3/3 algorithms in the artemether-lumefantrine (AL) treatment arm, while msp-1/msp- 2/glurp combination classified more cases of recrudescence using the ≥ 2/3 in the dihydroartemisinin-piperaquine (DP) arm. Microsatellites PfPK2 and Poly-α, potentially sensitive to detecting minority clones, are promising replacements for glurp. Discrepancies in recrudescence classification between match-counting and Bayesian algorithms highlight the need for standardized PCR correction practicesenAttribution-NonCommercial-NoDerivs 3.0 United Stateshttp://creativecommons.org/licenses/by-nc-nd/3.0/us/Plasmodium falciparumRecrudescenceReinfectionmicrosatellitesmsp-1msp-2antimalarial drugArticle