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Genetic polymorphism of merozoite surface protein-1 and merozoite surface protein-2 in Plasmodium falciparum isolates from Brazzaville, Republic of Congo
Pembe Mayengue, Mathieu Ndounga, Freddy Malonga, Michel Bitemo, Francine Ntoumi
Malaria Journal , 2011, DOI: 10.1186/1475-2875-10-276
Abstract: A total of 125 isolates from patients with uncomplicated malaria attending Terinkyo and Madibou health centres were collected between January and June 2005 while evaluating the therapeutic efficacy of amodiaquine-artesunate combination. DNA was extracted and msp-1 and msp-2 genes were genotyped using allele-specific nested-PCR.Out of 468 distinct fragments detected, 15 msp-1 and 20 msp-2 genotypes were identified. For the msp-1 gene, K1 family was the predominant allelic type carried alone or in association with RO33 and Mad20 types, whereas the 3D7 family was the most prevalent in the msp-2 gene. Overall, the mean multiplicity of infection was 2.2. Out of 125 samples, 104 (83%) harboured more than one parasite genotype. There was no statistical significant difference in the multiplicity of infection by either sex or age of patients. However, a statistically significant correlation was found between parasite densities and the number of genotypes.Polymorphism in P. falciparum clinical isolates from Brazzaville was high and mainly of multiple clones. The basis for the positive association between parasite densities and multiplicity of infection is discussed.Malaria, a disease mostly caused by Plasmodium falciparum, is a major public health problem. The global burden is estimated at 225 million malaria cases every year resulting into 781,000 deaths [1], sub-Saharan Africa being the most affected region. In the Republic of Congo, like in many other sub-Saharan African endemic countries, malaria vulnerable groups are children and pregnant women [2].Data collected from 2000 to 2003 by the University Teaching Hospital of Brazzaville during studies on chloroquine resistance showed that 22% of children deaths at pediatric health facilities were due to malaria [3]. Recent studies on anti-malarial drug resistance in Brazzaville have confirmed high level chloroquine resistance and the inefficacy of sulphadoxine-pyrimethamine and amodiaquine either singly or in combination for t
Artesunate-amodiaquine efficacy in Congolese children with acute uncomplicated falciparum malaria in Brazzaville
Ndounga Mathieu,Mayengue Pembe Issamou,Casimiro Prisca Nadine,Loumouamou Dieudonné
Malaria Journal , 2013, DOI: 10.1186/1475-2875-12-53
Abstract: Background Congo-Brazzaville adopted artemisinin-based combination therapy (ACT) in 2006. Artesunate-amodiaquine (AS + AQ) and artemether-lumefantrine are the first-line and second-line anti-malarial drugs to treat uncomplicated Plasmodium falciparum malaria, respectively. The baseline efficacy of AS + AQ was evaluated from February to August 2005 in patients living in Brazzaville, the capital city of the Republic of Congo. Methods One hundred and ninety-seven patients (96 ≤5 years old and 101 >5 years old, including adults) were recruited in a non-randomized study, treated under supervision with AS + AQ, and were followed up for 28 days in accordance with the 2003 World Health Organization protocol. Plasmodium falciparum recrudescent isolates from day 7 to day 28 were compared to pretreatment isolates by polymerase chain reaction (PCR) to distinguish between re-infection and recrudescence. Results The overall efficacy of AS + AQ after PCR correction on day 28 was 94.4%. An adequate clinical and parasitological response was observed in 94.3% and 94.4% of children aged ≤5 years old and those aged >5 years old (including adults), respectively. The main reported adverse events were dizziness, vomiting, diarrhoea, pruritus, headache, anorexia, and abdominal pain. Conclusion This study has shown the high efficacy of AS + AQ in Congolese patients of all ages with acute uncomplicated falciparum malaria and serves as the baseline efficacy and tolerance of this ACT in Brazzaville.
Genetic polymorphism of merozoite surface protein 2 and prevalence of K76T pfcrt mutation in Plasmodium falciparum field isolates from Congolese children with asymptomatic infections
Felix Koukouikila-Koussounda, Vladimir Malonga, Pembe Mayengue, Mathieu Ndounga, Christevy Vouvoungui, Francine Ntoumi
Malaria Journal , 2012, DOI: 10.1186/1475-2875-11-105
Abstract: Between April and June 2010, 313 children below 10 years of age enrolled in the cohort for malaria surveillance were screened for P. falciparum infection using microscopy and polymerase chain reaction (PCR). The children were selected on the basis of being asymptomatic. Plasmodium falciparum msp2 gene was genotyped by allele-specific nested PCR and the pfcrt K76T mutation was detected using nested PCR followed by restriction endonuclease digestion.The prevalence of asymptomatic P. falciparum infections was 8.6% and 16% by microscopy and by PCR respectively. Allele typing of the msp2 gene detected 55% and 45% of 3D7 and FC27 allelic families respectively. The overall multiplicity of infections (MOI) was 1.3. A positive correlation between parasite density and multiplicity of infection was found. The prevalence of the mutant pfcrt allele (T76) in the isolates was 92%.This is the first molecular characterization of P. falciparum field isolates in Congolese children, four years after changing the malaria treatment policy from chloroquine (CQ) to artemisinin-based combination therapy (ACT). The low prevalence of asymptomatic infections and MOI is discussed in the light of similar studies conducted in Central Africa.Despite significant reduction in malaria-related morbidity and mortality in the recent past, malaria remains endemic in the tropics and sub-tropics including sub-Saharan Africa. About 225 million clinical cases and 781,000 deaths were reported worldwide in 2009, whereby almost 90% occurred in sub-Saharan Africa [1]. Current malaria control strategies include the use of insecticide-treated bed nets (ITNs), indoor residual spraying of insecticide, intermittent preventive treatment to young children and pregnant women, and early parasitological diagnosis and treatment of clinical cases using artemisinin-based combination therapy (ACT) [2-4]. Deployment of these strategies has had significant impact on malaria in many endemic areas.Although the impact of current m
Genetic evidence of regulatory gene variants of the STAT6, IL10R and FOXP3 locus as a susceptibility factor in uncomplicated malaria and parasitaemia in Congolese children
Koukouikila-Koussounda Felix,Ntoumi Francine,Ndounga Mathieu,Tong Hoang V
Malaria Journal , 2013, DOI: 10.1186/1475-2875-12-9
Abstract: Background Regulatory T cells (Tregs) are a subset of T cells that play an important role in modulating T effector responses during infectious challenges. The aim of this study was to evaluate possible associations between regulatory gene polymorphisms and the risk of uncomplicated malaria and the control of Plasmodium falciparum parasite density levels. Methods Twelve regulatory single nucleotide polymorphisms (SNPs) in the promoter regions of FOXP3 (ss270137548, rs11091253), IL10RA (rs56356146, rs7925112), IL10RB (rs8178433, rs8178435, rs999788), STAT6 (rs3024941, rs3024943, rs3024944) and TNFRSF18 (ss2080581728, rs3753344) were genotyped in a cohort of Congolese children. Studied subjects were followed up (passively) during one year. The children who experienced one or several clinical episodes were genotyped as “uncomplicated malaria” group (n=179) and those children who did not experience any episode were genotyped as “asymptomatic children” group (n=138). Results The prevalence of rs3024944CC genotype of STAT6 was significantly higher in the group of asymptomatic children compared to that of uncomplicated malaria (P=0.003). Similarly, the minor allele rs3024944C was more prevalent in the group of asymptomatic children (P=0.019). Two novel SNPs were observed including -163T/G (ss491228441) in IL10RA gene and -163C/T (ss491228440) in TNFRSF18 gene. The genotype ss491228441TT and the minor allele ss491228441G of the IL10RA were more frequent in the group of asymptomatic children (P=0.006 and P=0.007, respectively). The genotype rs11091253CT of the FOXP3 was associated with high parasite density levels. In addition, a new promoter IL10RA variant (ss491228441) contributes to shield against mild malaria. Conclusion The study indicated that the STAT6 promoter polymorphism rs3024944 was associated with uncomplicated malaria, whereas the FOXP3 promoter variant rs11091253 was associated with significant P. falciparum parasitaemia levels. These genetic data may contribute to the understanding of molecular mechanisms that regulate immune response to P. falciparum infections.
Clinical Efficacy of Artemether-Lumefantrine in Congolese Children with Acute Uncomplicated Falciparum Malaria in Brazzaville
Mathieu Ndounga,Rachida Tahar,Prisca N. Casimiro,Dieudonné Loumouamou
Malaria Research and Treatment , 2012, DOI: 10.1155/2012/749479
Abstract:
Reduction of multiplicity of infections but no change in msp2 genetic diversity in Plasmodium falciparum isolates from Congolese children after introduction of artemisinin-combination therapy
Ibara-Okabande Rod,Koukouikila-Koussounda Felix,Ndounga Mathieu,Vouvoungui Jeannhey
Malaria Journal , 2012, DOI: 10.1186/1475-2875-11-410
Abstract: Background In this first study conducted after the introduction of artemisinin-combination therapy (ACT), the major objective was to evaluate Plasmodium falciparum genetic diversity and multiplicity of infection in isolates from Congolese children between one and nine years of age enrolled and followed up for one year. The secondary objective was to characterize the msp2 profiles of P. falciparum isolates collected from successive malaria episodes in ten children who had four or more clinical episodes during the follow up. Methods Three-hundred and thirteen children residing in southern part of Brazzaville participated in this study. Blood samples were obtained from all children at enrollment and checked for P. falciparum infection. Based on the one year follow-up data, two clinical groups were considered according to the number of malaria episodes presented over the follow up period: “protected”(children who did not experience any episode) and “unprotected” (those who experienced more that two episodes). Therefore, the msp2 genetic diversity of P. falciparum isolates collected at enrollment in the two groups was characterized by allele-specific nested PCR and compared. The msp2 profiles of P. falciparum isolates collected from successive malaria episodes was also characterized by allele-specific nested PCR. Results Forty-three percent of FC27 and fifty-seven percent of 3D7 in protected vs fifty-six percent of FC27 and forty-four percent of 3D7 in isolates from unprotected children were detected. Seven and two alleles belonging to the FC27, and six and three alleles belonging to 3D7 families were distinguished in isolates from protected and unprotected children respectively. The mean multiplicity of infection (MOI) values at inclusion for the msp2 locus was 1.29 and 1.43 for protected and unprotected children respectively. 43 isolates were obtained from the ten children who had four or more clinical episodes during the follow up. A total of 63 alleles or fragments corresponding to 57% (36/63) FC27 and 43% (27/63) 3D7 were detected. The variant 400bp of FC27 was the most prevalent. 46% (20/43), 42% (18/43), 2% (1/43) and 2% (1/43) of isolates were found to have 1, 2, 3 and 4 parasite genotypes respectively and the mean MOI was 1.78. Conclusion This study shows that the introduction of ACT in the Republic of Congo has reduced the MOI but not the genetic diversity of P. falciparum isolates from children living in Southern districts of Brazzaville.
Therapeutic efficacy of sulfadoxine-pyrimethamine, amodiaquine and the sulfadoxine-pyrimethamine-amodiaquine combination against uncomplicated Plasmodium falciparum malaria in young children in Cameroon
Basco Leonardo K.,Same-Ekobo Albert,Ngane Vincent Foumane,Ndounga Mathieu
Bulletin of the World Health Organization , 2002,
Abstract: OBJECTIVE: To evaluate the therapeutic efficacy of sulfadoxine-pyrimethamine, amodiaquine, and the sulfadoxine-pyrimethamine-amodiaquine combination for the treatment of uncomplicated Plasmodium falciparum malaria in young children in Cameroon. METHODS: In a randomized study we evaluated the effectiveness and tolerance of (i) sulfadoxine-pyrimethamine (SP) (25 mg/kg body weight of sulfadoxine and 1.25 mg/kg of pyrimethamine in a single oral dose), (ii) amodiaquine (AQ) (30 mg/kg body weight in three divided daily doses), and (iii) the sulfadoxine-pyrimethamine-amodiaquine combination (SP+AQ) (same doses as in the other two treatment groups, given simultaneously on day 0) in young children in southern Cameroon. The parasitological and clinical responses were studied until day 28 in accordance with the modified 1996 WHO protocol for the evaluation of the therapeutic efficacy of antimalarial drugs. FINDINGS: Of 191 enrolled patients, 6 and 8 were excluded or lost to follow-up before day 14 and between day 14 and day 28, respectively. For the AQ-treated patients, parasitological and clinical evaluation on day 14 showed late treatment failure in 2 of 61 (3.3%) and adequate clinical response with parasitological failure in one (1.6%). There was an adequate clinical response in all patients treated with SP or SP+AQ. Therapeutic failure rates on day 28 were 13.6%, 10.2% and 0% in the SP, AQ, and SP+AQ groups, respectively. Anaemia improved in all three regimens. AQ produced faster fever clearance but was associated with more transient minor side-effects than SP. SP+AQ reduced the risk of recrudescence between day 14 and day 28 but increased the incidence of minor side-effects. CONCLUSION: SP+AQ can be recommended as a temporary means of slowing the spread of multidrug resistance in Plasmodium falciparum in Africa while the introduction of other combinations, including artemisinin derivatives, is awaited.
Clinical Efficacy of Artemether-Lumefantrine in Congolese Children with Acute Uncomplicated Falciparum Malaria in Brazzaville
Mathieu Ndounga,Rachida Tahar,Prisca N. Casimiro,Dieudonné Loumouamou,Leonardo K. Basco
Malaria Research and Treatment , 2012, DOI: 10.1155/2012/749479
Abstract: The Republic of the Congo adopted artemisinin-based combination therapies (ACTs) in 2006: artesunate-amodiaquine and artemether-lumefantrine as the first-line and second-line drugs, respectively. The baseline efficacy of artemether-lumefantrine was evaluated between March and July 2006 in Brazzaville, the capital city of Congo. Seventy-seven children aged between 6 months and 10 years were enrolled in a nonrandomized study. The children were treated under supervision with 6 doses of artemether-lumefantrine and followed up for 28 days in accordance with the 2003 World Health Organization guideline. Pretreatment (i.e., day 0) and recrudescent Plasmodium falciparum isolates between day 14 and day 28 were compared by the polymerase chain reaction to distinguish between true recrudescence and reinfection. The overall cure rate on day 28 was 96.9% after PCR correction. Reported adverse effects included pruritus and dizziness. Artemether-lumefantrine was highly efficacious in Brazzaville. Approximately 30% of the Congolese population reside in Brazzaville, the capital city. The epidemiology of malaria in the city of Brazzaville is heterogeneous [1]. Depending on the district, malaria transmission is low or intense. In general, malaria is meso- to hypoendemic in the city centre and hyperendemic in the periphery [2]. In terms of malaria burden, there are twice as many malaria-infected patients consulting health centres in the periphery, as compared with health centres in the city centre [3]. Surveys conducted in the main hospital in Brazzaville have shown that malaria is the first cause of admission in the department of paediatrics, mostly in children aged less than 4 years old [4, 5]. Due to the high levels of clinical resistance to chloroquine, amodiaquine, and sulphadoxine-pyrimethamine [6, 7], the Congolese Ministry of Public Health changed the national antimalarial drug policy in 2006. Two artemisinin-based combination therapies (ACTs) were adopted: artesunate-amodiaquine and artemether-lumefantrine for the first-line and second-line treatment of uncomplicated malaria, respectively. Before the drug policy change, only a single clinical study on the efficacy of artesunate-amodiaquine and artemether-lumefantrine had been conducted in a rural area in Congo [8]. The present nonrandomized study was conducted between March and July 2006 to provide the baseline data of artemether-lumefantrine efficacy in an urban area where the majority of the Congolese population reside. The study was conducted in Tenrikyo health centre located in Makélékélé district, which is in
Therapeutic efficacy of sulfadoxine-pyrimethamine, amodiaquine and the sulfadoxine-pyrimethamine-amodiaquine combination against uncomplicated Plasmodium falciparum malaria in young children in Cameroon
Basco,Leonardo K.; Same-Ekobo,Albert; Ngane,Vincent Foumane; Ndounga,Mathieu; Metoh,Theresia; Ringwald,Pascal; Soula,Georges;
Bulletin of the World Health Organization , 2002, DOI: 10.1590/S0042-96862002000700005
Abstract: objective: to evaluate the therapeutic efficacy of sulfadoxine-pyrimethamine, amodiaquine, and the sulfadoxine-pyrimethamine-amodiaquine combination for the treatment of uncomplicated plasmodium falciparum malaria in young children in cameroon. methods: in a randomized study we evaluated the effectiveness and tolerance of (i) sulfadoxine-pyrimethamine (sp) (25 mg/kg body weight of sulfadoxine and 1.25 mg/kg of pyrimethamine in a single oral dose), (ii) amodiaquine (aq) (30 mg/kg body weight in three divided daily doses), and (iii) the sulfadoxine-pyrimethamine-amodiaquine combination (sp+aq) (same doses as in the other two treatment groups, given simultaneously on day 0) in young children in southern cameroon. the parasitological and clinical responses were studied until day 28 in accordance with the modified 1996 who protocol for the evaluation of the therapeutic efficacy of antimalarial drugs. findings: of 191 enrolled patients, 6 and 8 were excluded or lost to follow-up before day 14 and between day 14 and day 28, respectively. for the aq-treated patients, parasitological and clinical evaluation on day 14 showed late treatment failure in 2 of 61 (3.3%) and adequate clinical response with parasitological failure in one (1.6%). there was an adequate clinical response in all patients treated with sp or sp+aq. therapeutic failure rates on day 28 were 13.6%, 10.2% and 0% in the sp, aq, and sp+aq groups, respectively. anaemia improved in all three regimens. aq produced faster fever clearance but was associated with more transient minor side-effects than sp. sp+aq reduced the risk of recrudescence between day 14 and day 28 but increased the incidence of minor side-effects. conclusion: sp+aq can be recommended as a temporary means of slowing the spread of multidrug resistance in plasmodium falciparum in africa while the introduction of other combinations, including artemisinin derivatives, is awaited.
Plasmodium falciparum: Differential Selection of Drug Resistance Alleles in Contiguous Urban and Peri-Urban Areas of Brazzaville, Republic of Congo
Yoko Tsumori, Mathieu Ndounga, Toshihiko Sunahara, Nozomi Hayashida, Megumi Inoue, Shusuke Nakazawa, Prisca Casimiro, Rie Isozumi, Haruki Uemura, Kazuyuki Tanabe, Osamu Kaneko, Richard Culleton
PLOS ONE , 2011, DOI: 10.1371/journal.pone.0023430
Abstract: The African continent is currently experiencing rapid population growth, with rising urbanization increasing the percentage of the population living in large towns and cities. We studied the impact of the degree of urbanization on the population genetics of Plasmodium falciparum in urban and peri-urban areas in and around the city of Brazzaville, Republic of Congo. This field setting, which incorporates local health centers situated in areas of varying urbanization, is of interest as it allows the characterization of malaria parasites from areas where the human, parasite, and mosquito populations are shared, but where differences in the degree of urbanization (leading to dramatic differences in transmission intensity) cause the pattern of malaria transmission to differ greatly. We have investigated how these differences in transmission intensity affect parasite genetic diversity, including the amount of genetic polymorphism in each area, the degree of linkage disequilibrium within the populations, and the prevalence and frequency of drug resistance markers. To determine parasite population structure, heterozygosity and linkage disequilibrium, we typed eight microsatellite markers and performed haplotype analysis of the msp1 gene by PCR. Mutations known to be associated with resistance to the antimalarial drugs chloroquine and pyrimethamine were determined by sequencing the relevant portions of the crt and dhfr genes, respectively. We found that parasite genetic diversity was comparable between the two sites, with high levels of polymorphism being maintained in both areas despite dramatic differences in transmission intensity. Crucially, we found that the frequencies of genetic markers of drug resistance against pyrimethamine and chloroquine differed significantly between the sites, indicative of differing selection pressures in the two areas.
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