Artemisinin Partial Resistance in Ethiopian Plasmodium Falciparum: A Multisite Clinical, Molecular and In Vitro Study

Didier Menard277 enrolled

Overview

Artemisinin-based combination therapies (ACTs) are the main treatment for falciparum malaria in Africa. Artemisinin partial resistance (ART-R), characterized by delayed parasite clearance after treatment, has been confirmed in four sub-Saharan African countries. In Ethiopia, molecular surveys have detected the Pfkelch13 R622I mutation associated with ART-R at multiple sites, but no study has yet combined clinical, molecular, and in vitro evidence to confirm ART-R per WHO criteria. This multisite study conducted across five sentinel sites in Ethiopia (2024-2025) assessed day-3 parasite positivity after artemether-lumefantrine treatment, Pfkelch13 genotyping, and ring-stage survival assay on culture-adapted field isolates, to determine whether ART-R is confirmed in Ethiopian Plasmodium falciparum populations.

This study integrated three WHO-required lines of evidence to confirm artemisinin partial resistance (ART-R) in Ethiopia: (1) clinical evidence through day-3 parasite positivity assessment after artemether-lumefantrine treatment in therapeutic efficacy studies; (2) molecular evidence through Pfkelch13 propeller domain genotyping; and (3) phenotypic in vitro evidence through ring-stage survival assay (RSA0-3h) on culture-adapted field isolates. Five sentinel sites were selected across malaria-endemic regions of Ethiopia. Blood samples were collected at enrollment (day 0) and day 3. Isolates were cryopreserved and shipped to France (Strasbourg) for RSA.

Study Type

INTERVENTIONAL

Allocation

Purpose

DIAGNOSTIC

Masking

NONE

Enrollment

277

Conditions

Malaria (Plasmodium Falciparum)

Eligibility

Sex: ALLMin age: 6 Months

Medical Language ↔ Plain English

Inclusion Criteria: * Age ≥ 6 months * Microscopically confirmed uncomplicated Plasmodium falciparum monoinfection * Axillary temperature ≥ 37.5°C or history of fever in the past 24 hours * Ability to take oral medication * Written informed consent from patient or parent/guardian * Residence in the study area with intention to remain during follow-up Exclusion Criteria: * Severe or complicated malaria (WHO criteria) * Mixed Plasmodium infection * Pregnancy or breastfeeding * Known hypersensitivity to artemether-lumefantrine * Antimalarial treatment in the 4 weeks prior to enrollment * Severe malnutrition * Concomitant febrile illness other than malaria requiring systemic treatment

Locations (5)

Mehoni Health Center

Mersa, Tigray, Ethiopia

Bako Health Center

Bako, Ethiopia

Workamba Health Center

Kechemo, Ethiopia

Metahara Health Center

Metehara, Ethiopia

Rama Health Center

Rama, Ethiopia

Outcomes

Primary Outcomes

Day-3 Parasite Positivity Rate

Proportion of patients with microscopically detectable Plasmodium falciparum parasitaemia on day 3 (72 ± 2 hours) after initiation of artemether-lumefantrine treatment, assessed by Giemsa-stained thick blood smear examination.

Time frame: Day 3 (72 hours after treatment initiation)

Secondary Outcomes

Prevalence of Pfkelch13 R622I Mutation

Proportion of day-0 samples carrying the validated Pfkelch13 R622I mutation, assessed by targeted amplicon sequencing.

Time frame: Day 0 (enrollment)

Ring-Stage Survival Rate

In vitro survival rate of culture-adapted field isolates after 6-hour exposure to 700 nM dihydroartemisinin, assessed by ring-stage survival assay (RSA0-3h). Resistance threshold: survival rate \>1%.

Time frame: Assessed on culture-adapted isolates collected at Day 0