Comparative effect of artemether-lumefantrine and artesunate-amodiaquine on gametocyte clearance in children with uncomplicated Plasmodium falciparum malaria in Madagascar.

BACKGROUND: Gametocytes are the sexual stages ensuring continuity of the development cycle of the parasite, as well as its transmission to humans. The efficacy of artemisinin-based anti-malarials against asexual stages of Plasmodium has been reported in Madagascar, but their effects on gametocytes are not well documented. The present study aims to determine the emergence of gametocyte and gametocyte clearance after artesunate-amodiaquine (ASAQ) or artemether-lumefantrine (AL) treatment in children with uncomplicated Plasmodium falciparum malaria in 5 regions of Madagascar. METHODS: 558 children with uncomplicated P. falciparum malaria, aged between 1 and 15 years, were assigned randomly to AL or ASAQ treatment. They come from 5 regions of Madagascar with different epidemiological facies related to malaria: Ankilivalo, Benenitra, Ampanihy, Ankazomborona and Matanga. Gametocytes were identified by microscopy, from t blood smears at day 1, day 2, day 3, day 7, day 14, day 21 and day 28 after treatment. RESULTS: At baseline, 9.7% (54/558) children [95% CI: 7.4-12.5%] had detectable gametocyte by microscopy. Among the 54 enrolled children, gametocytes emergence rate was high during the first days of treatment in both treatment arms (AL and ASAQ), especially on day 1. Gametocytes were undetectable from day 14 for AL arm while for ASAQ arm, gametocyte carriage was gradually decreased but persisted until day 21. CONCLUSION: This study demonstrates that AL has a more rapid effect on gametocyte clearance compared to ASAQ in children with uncomplicated Plasmodium falciparum malaria.

[Pcr-rflp genotyping of pfcrt and pfmdr1 in plasmodium falciparum isolates from children in Vatomandry, Madagascar]. / Génotypage par pcr-rflp de pfcrt et pfmdr1 sur des isolats de plasmodium falciparum collectés chez des enfants de Vatomandry, Madagascar.

Background: Malaria is a parasitic disease caused by a hematozoan of the genus Plasmodium. Early diagnosis followed by effective treatment is one of the keys to control this disease. In Madagascar, after more than 60 years of use for the treatment of uncomplicated malaria, chloroquine (CQ) was abandoned in favor of artesunate + amodiaquine (ASAQ) combination because of high prevalence of CQ treatment failure. Surveillance based on the assessment of therapeutic efficacy and genetic markers of resistance to antimalarials is therefore essential in order to detect the emergence of potentially resistant parasites as early as possible. In this context, our study aimed to genotype the Plasmodium falciparum chloroquine resistance transporter gene or Pfcrt and Plasmodium falciparum multidrug resistance gene 1 or Pfmdr1 in isolates collected from children in the district of Vatomandry. Methods: A total of 142 P. falciparum isolates collected during active case detection of malaria in children under 15 years old, between February and March of 2016 and 2017 in Vatomandry district, were analyzed. Pfcrt (K76T codon) and Pfmdr1 (N86Y codon) genotyping was carried out by polymerase chain reaction followed by enzymatic digestion (restriction fragment length polymorphism) or PCR-RFLP. Results: The successful rates of amplification of Pfcrt and Pfmdr1 genes were low, around 27% and 39% respectively. The prevalence of isolates carrying the mutant Pfcrt K76T codon and the mutant Pfmdr1 N86Y codon was 2.6% [95% confidence interval (95% CI): 0.1 - 15.0%] and 36% [95% CI: 23.7 - 49.7%] respectively. Conclusion: Despite the limited number of samples analyzed, our study highlighted the circulation of isolates carrying both the mutant Pfcrt K76T and Pfmdr1 N86Y alleles. Although the prevalence of mutations in Pfcrt and Pfmdr1 genes that we observed was low, other studies should be carried out in order to follow the evolution of these markers in time and space. The use of more sensitive methods will better characterize P. falciparum strains circulating in Madagascar. Artesunate-amodiaquine is used as a first-line treatment for uncomplicated malaria in the country; it is also crucial to monitor the other codons, i.e. 184 and 1246 of the Pfmdr1 gene, implicated in the resistance of P. falciparum to amodiaquine in Africa.

DNA recovery from used malaria RDT to detect Plasmodium species and to assess Plasmodium falciparum genetic diversity: a pilot study in Madagascar.

BACKGROUND: Rapid diagnostic tests (RDT) are widely used for malaria diagnosis in Madagascar, where Plasmodium falciparum is the predominant species. Molecular diagnosis is essential for malaria surveillance, but requires additional blood samples for DNA extraction. Used RDTs is an attractive alternative that can be used as a source of DNA. Plasmodium falciparum genetic diversity and multiplicity of infection, usually determined by the genotyping of polymorphic regions of merozoite surface proteins 1 and 2 genes (msp1, msp2), and the repeated region RII of the glutamate-rich protein gene (glurp) have been associated with malaria transmission levels and subsequently with the impact of the deployed control strategies. Thus, the study aims to use RDT as DNA source to detect Plasmodium species, to characterize Plasmodium falciparum genetic diversity and determine the multiplicity of infection. METHODS: A pilot study was conducted in two sites with different epidemiological patterns: Ankazomborona (low transmission area) and Matanga (high transmission area). On May 2018, used RDT (SD BIOLINE Malaria Ag P.f/Pan, 05FK63) were collected as DNA source. Plasmodium DNA was extracted by simple elution with nuclease free water. Nested-PCR were performed to confirm Plasmodium species and to analyse P. falciparum msp1, msp2 and glurp genes polymorphisms. RESULTS: Amongst the 170 obtained samples (N = 74 from Ankazomborona and N = 96 from Matanga), Plasmodium positivity rate was 23.5% (40/170) [95% CI 17.5-30.8%] by nested-PCR with 92.2% (37/40) positive to P. falciparum, 5% (2/40) to Plasmodium vivax and 2.5% (1/40) to P. falciparum/P. vivax mixed infection. Results showed high polymorphisms in P. falciparum msp1, msp2 and glurp genes. Multiple infection rate was 28.6% [95% CI 12.2-52.3%]. The mean of MOI was 1.79 ± 0.74. CONCLUSION: This pilot study highlighted that malaria diagnosis and molecular analysis are possible by using used malaria RDT. A large-scale study needs to be conducted to assess more comprehensively malaria parasites transmission levels and provide new data for guiding the implementation of local strategies for malaria control and elimination. Trial registration Retrospectively registered.

Monitoring parasite diversity for malaria elimination in sub-Saharan Africa.

The African continent continues to bear the greatest burden of malaria and the greatest diversity of parasites, mosquito vectors, and human victims. The evolutionary plasticity of malaria parasites and their vectors is a major obstacle to eliminating the disease. Of current concern is the recently reported emergence of resistance to the front-line drug, artemisinin, in South-East Asia in Plasmodium falciparum, which calls for preemptive surveillance of the African parasite population for genetic markers of emerging drug resistance. Here we describe the Plasmodium Diversity Network Africa (PDNA), which has been established across 11 countries in sub-Saharan Africa to ensure that African scientists are enabled to work together and to play a key role in the global effort for tracking and responding to this public health threat.