Molecular investigation of malaria-infected patients in Djibouti city (2018-2021).

BACKGROUND: The Republic of Djibouti is a malaria endemic country that was in pre-elimination phase in 2006-2012. From 2013, however, malaria has re-emerged in the country, and its prevalence has been increasing every year. Given the co-circulation of several infectious agents in the country, the assessment of malaria infection based on microscopy or histidine-rich protein 2 (HRP2)-based rapid diagnostic tests (RDT) has shown its limitations. This study, therefore, aimed to assess the prevalence of malaria among febrile patients in Djibouti city using more robust molecular tools. METHODS: All suspected malaria cases reported to be microscopy-positive were randomly sampled (n = 1113) and included in four health structures in Djibouti city over a 4-year period (2018-2021), mainly during the malaria transmission season (January-May). Socio-demographic information was collected, and RDT was performed in most of the included patients. The diagnosis was confirmed by species-specific nested polymerase chain reaction (PCR). Data were analysed using Fisher's exact test and kappa statistics. RESULTS: In total, 1113 patients with suspected malaria and available blood samples were included. PCR confirmed that 788/1113 (70.8%) were positive for malaria. Among PCR-positive samples, 656 (83.2%) were due to Plasmodium falciparum, 88 (11.2%) Plasmodium vivax, and 44 (5.6%) P. falciparum/P. vivax mixed infections. In 2020, P. falciparum infections were confirmed by PCR in 50% (144/288) of negative RDTs. After the change of RDT in 2021, this percentage decreased to 17%. False negative RDT results were found more frequently (P < 0.05) in four districts of Djibouti city (Balbala, Quartier 7, Quartier 6, and Arhiba). Malaria occurred less frequently in regular bed net users than in non-users (odds ratio [OR]: 0.62, 95% confidence interval [CI]: 0.42-0.92). CONCLUSIONS: The present study confirmed the high prevalence of falciparum malaria and, to a lesser extent, vivax malaria. Nevertheless, 29% of suspected malaria cases were misdiagnosed by microscopy and/or RDT. There is a need to strengthen the capacity for diagnosis by microscopy and to evaluate the possible role of P. falciparum hrp2 gene deletion, which leads to false negative cases of P. falciparum.

Role of Anopheles stephensi Mosquitoes in Malaria Outbreak, Djibouti, 2019.

Anopheles stephensi mosquitoes share urban breeding sites with Aedes aegypti and Culex quinquefasciatus mosquitoes in the Republic of Djibouti. We present evidence that A. stephensi mosquitoes might be responsible for an increase in malaria incidence in this country. We also document resistance of Plasmodium falciparum to dihydroartemisinin/piperaquine.

Population genetics analysis during the elimination process of Plasmodium falciparum in Djibouti.

BACKGROUND: Case management of imported malaria within the context of malaria pre-elimination is increasingly considered to be relevant because of the risk of resurgence. The assessment of malaria importation would provide key data i) to select countries with propitious conditions for pre-elimination phase and ii) to predict its feasibility. Recently, a sero-prevalence study in Djibouti indicated low malaria prevalence, which is propitious for the implementation of pre-elimination, but data on the extent of malaria importation remain unknown. METHODS: Djiboutian plasmodial populations were analysed over an eleven-year period (1998, 1999, 2002 and 2009). The risk of malaria importation was indirectly assessed by using plasmodial population parameters. Based on 5 microsatellite markers, expected heterozygosity (H.e.), multiplicity of infection, pairwise Fst index, multiple correspondence analysis and individual genetic relationship were determined. The prevalence of single nucleotide polymorphisms associated with pyrimethamine resistance was also determined. RESULTS: Data indicated a significant decline in genetic diversity (0.51, 0.59, 0.51 and 0 in 1998, 1999, 2002 and 2009, respectively) over the study period, which is inconsistent with the level of malaria importation described in a previous study. This suggested that Djiboutian malaria situation may have benefited from the decline of malaria prevalence that occurred in neighbouring countries, in particular in Ethiopia. The high Fst indices derived from plasmodial populations from one study period to another (0.12 between 1999 and 2002, and 0.43 between 2002 and 2009) suggested a random sampling of parasites, probably imported from neighbouring countries, leading to oligo-clonal expansion of few different strains during each transmission season. Nevertheless, similar genotypes observed during the study period suggested recurrent migrations and imported malaria. CONCLUSION: In the present study, the extent of genetic diversity was used to assess the risk of malaria importation in the low malaria transmission setting of Djibouti. The molecular approach highlights i) the evolution of Djiboutian plasmodial population profiles that are consistent and compatible with Djiboutian pre-elimination goals and ii) the necessity to implement the monitoring of plasmodial populations and interventions at the regional scale in the Horn of Africa to ensure higher efficiency of malaria control and elimination.

Plasmodium vivax and Plasmodium falciparum infections in the Republic of Djibouti: evaluation of their prevalence and potential determinants.

BACKGROUND: Formerly known as a hypoendemic malaria country, the Republic of Djibouti declared the goal of pre-eliminating malaria in 2006. The aim of the present study was to evaluate the prevalence of Plasmodium falciparum, Plasmodium vivax and mixed infections in the Djiboutian population by using serological tools and to identify potential determinants of the disease and hotspots of malaria transmission within the country. METHODS: The prevalence of P. falciparum and P. vivax within the districts of the capital city and the rest of the Republic of Djibouti were assessed using 13 and 2 serological markers, respectively. The relationship between the immune humeral response to P. falciparum and P. vivax and variables such as age, gender, wealth status, urbanism, educational level, distance to rivers/lakes, living area, having fever in the last month, and staying in a malaria-endemic country more than one year was estimated and analysed by questionnaires administered to 1910 Djiboutians. Multivariate ordinal logistic regression models of the immune humeral response were obtained for P. falciparum and P. vivax. RESULTS: The P. falciparum and P. vivax seroprevalence rates were 31.5%, CI95% [29.4-33.7] and 17.5%, CI95% [15.8-19.3], respectively. Protective effects against P. falciparum and P. vivax were female gender, educational level, and never having visited a malaria-endemic area for more than one year. For P. falciparum only, a protective effect was observed for not having a fever in the last month, living more than 1.5 km away from lakes and rivers, and younger ages. CONCLUSIONS: This is the first study that assessed the seroprevalence of P. vivax in the Republic of Djibouti. It is necessary to improve knowledge of this pathogen in order to create an effective elimination programme. As supported by recent observations on the subject, the Republic of Djibouti has probably demonstrated a real decrease in the transmission of P. falciparum in the past seven years, which should encourage authorities to improve efforts toward elimination.