High resolution melting: a useful field-deployable method to measure dhfr and dhps drug resistance in both highly and lowly endemic Plasmodium populations

Background: Emergence and spread of drug resistance to every anti-malarial used to date, creates an urgent need for development of sensitive, specifc and feld-deployable molecular tools for detection and surveillance of validated drug resistance markers. Such tools would allow early detection of mutations in resistance loci. The aim of this study was to compare common population signatures and drug resistance marker frequencies between two populations with diferent levels of malaria endemicity and history of anti-malarial drug use: Tanzania and Sénégal. This was accomplished by implementing a high resolution melting assay to study molecular markers of drug resistance as compared to polymerase chain reaction–restriction fragment length polymorphism (PCR/RFLP) methodology. Methods: Fifty blood samples were collected each from a lowly malaria endemic site (Sénégal), and a highly malaria endemic site (Tanzania) from patients presenting with uncomplicated Plasmodium falciparum malaria at clinic. Data representing the DHFR were derived using both PCR–RFLP and HRM assay; while genotyping data representing the DHPS were evaluated in Senegal and Tanzania using HRM. Msp genotyping analysis was used to characterize the multiplicity of infection in both countries. Results: A high prevalence of samples harbouring mutant DHFR alleles was observed in both population using both genotyping techniques. HRM was better able to detect mixed alleles compared to PCR/RFLP for DHFR codon 51 in Tanzania; and only HRM was able to detect mixed infections from Senegal. A high prevalence of mutant alleles in DHFR (codons 51, 59, 108) and DHPS (codon 437) were found among samples from Sénégal while no mutations were observed at DHPS codons 540 and 581, from both countries. Overall, the frequency of samples harbouring either a single DHFR mutation (S108N) or double mutation in DHFR (C59R/S108N) was greater in Sénégal compared to Tanzania Conclusion: Here the results demonstrate that HRM is a rapid, sensitive, and feld-deployable alternative technique to PCR–RFLP genotyping that is useful in populations harbouring more than one parasite genome (polygenomic infections). In this study, a high levels of resistance polymorphisms was observed in both dhfr and dhps, among samples from Tanzania and Sénégal. A routine monitoring by molecular markers can be a way to detect emergence of resistance involving a change in the treatment policy

West Africa International Centers of Excellence for Malaria Research: Drug Resistance Patterns to Artemether-Lumefantrine in Senegal, Mali, and The Gambia.

In 2006, artemether-lumefantrine (AL) became the first-line treatment of uncomplicated malaria in Senegal, Mali, and the Gambia. To monitor its efficacy, between August 2011 and November 2014, children with uncomplicated Plasmodium falciparum malaria were treated with AL and followed up for 42 days. A total of 463 subjects were enrolled in three sites (246 in Senegal, 97 in Mali, and 120 in Gambia). No early treatment failure was observed and malaria infection cleared in all patients by day 3. Polymerase chain reaction (PCR)-adjusted adequate clinical and parasitological response (ACPR) was 100% in Mali, and the Gambia, and 98.8% in Senegal. However, without PCR adjustment, ACPR was 89.4% overall; 91.5% in Mali, 98.8% in Senegal, and 64.3% in the Gambia (the lower value in the Gambia attributed to poor compliance of the full antimalarial course). However, pfmdr1 mutations were prevalent in Senegal and a decrease in parasite sensitivity to artesunate and lumefantrine (as measured by ex vivo drug assay) was observed at all sites. Recrudescent parasites did not show Kelch 13 (K13) mutations and AL remains highly efficacious in these west African sites

Completeness of information in electronic compared with paper-based patients’ records in a maternity setting in Dakar, Senegal

Background: Evaluate the consistency of information in paper-based records when registered in parallel with an electronic medical record.Methods: The study was performed at PMSHC in Dakar Senegal. From the end of year 2016, patients’ files were recorded on both paper-based and electronically. Additionally, previous records were electronically registered. To investigate the completeness of records before and after the electronic recording system has been implemented, information about some maternal and fetal/neonatal characteristics were assessed. When the variable was recorded, the system returned 1, unrecorded variables were coded as 0. We then calculated, for each variable, the unrecorded rate before 2017 and after that date. The study period extended from 2011 to June 2019, a nearly ten-year period. Data were extracted from E-perinatal to MS excel 2019 then SPSS 25 software. Frequencies of unrecorded variables were compared with chi-squared test at a level of significance of 5%.Results: A total of 48,270 unique patients’ records were identified during the eight-year period.  Among the study population, data for patients’ age, address and parity were available most of the time before and after 2017 (0.5% missing data versus 0.3% for age and 2.6% versus 1.3% for home address and from 0.3% to 0.0% for parity). However, phone number, maternal weight, maternal height, last menstrual period and blood group were found to be missing up to 96% before 2017. From 2017, these rates experienced a sudden decrease at a significant level: from 82.4% to 27.8% for phone number, from 96% to 56.3% for maternal weight and from 60.1% to 21.3% for blood group. Regarding newborns’ data, it was found that fetal height, head circumference and chest circumference were missing up to just under 25% before 2017. After that date, their completeness improved and flattened under 5%.Conclusions: Structured and computerized files reduce missing data. There is an urgent need the Ministry of health provides hospitals and health care providers with guidelines that describes the standardized information that should be gathered and shared in health and care records

Mise en place d’un Village Intelligent face au Climat pour la réduction des risques climatiques et de l’insécurité alimentaire à Daga-Birame, Sénégal. Guide de visite de terrain pour la Réunion du Comité Scientifique Indépendant du CCAFS

À Daga Birame, au Sénégal, CCAFS et ses partenaires ont mis en place un village climato-intelligent (CSV) dans lequel plusieurs activités sont menées. Sur la base de la vision du village et de son avenir, un ensemble d'actions ont été identifiées par la communauté afin d'atteindre les changements souhaités dans la productivité agricole et la sécurité alimentaire tels que les activités génératrices de revenus, l'amélioration de la résilience et la gestion durable des ressources naturelles du village. Ces actions ont été structurées autour de quatre composantes: Les services d'information climatologique (SIC); le développement des technologies / pratiques agricoles adaptées au climat; le renforcement de capacités des villageois et celle des connaissances et des institutions locales

Case study of Daga-Birame CSV for CCAFS ISP11/6.1.2 – Senegal

Senegal, with 196,712 km2 land area, is located at the extreme west of the African continent (Longitudes 11°21W - 17°32N and Latitudes 12°8N - 16°41N). The country’s soils are in general of low fertility, fragile and very susceptible to wind and water erosion. The climate is of Sudano-Sahelian type characterized by alternating dry season (November to May) and rainy season (June to October). The 700 km coastline brings climatic differences between coastal areas and inland zones. Rainfall amount follows a latitudinal variation going from 300 mm in the north semi-desertic areas to 1200 mm in the south. Senegal is divided into 7 agro-ecological zones for management perspectives: River Valley, Niayes, Groundnut Basin (North and South), Silvo-Pastoral zone, Eastern Senegal and Upper Casamance, Lower Casamance (CIAT-BFS/USAID, 2016). The country’s economy is mainly driven by crop and livestock production contributing 17% of the GDP and employing about 70% of the population (NAPA, Republic of Senegal 2006). Like other sub-Saharan African countries, Senegal faces food insecurity as a consequence of climate variability and change combined with other global changes (Zougmoré et al., 2015)

Selection of N86F184D1246 haplotype of Pfmrd1 gene by artemether–lumefantrine drug pressure on Plasmodium falciparum populations in Senegal

Background: The use of artemisinin as a monotherapy resulted in the emergence of artemisinin resistance in 2005 in Southeast Asia. Monitoring of artemisinin combination therapy (ACT) is critical in order to detect and prevent the spread of resistance in endemic areas. Ex vivo studies and genotyping of molecular markers of resistance can be used as part of this routine monitoring strategy. One gene that has been associated in some ACT partner drug resistance is the Plasmodium falciparum multidrug resistance protein 1 (pfmdr1) gene. The purpose of this study was to assess the drug susceptibility of P. falciparum populations from Thiès, Senegal by ex vivo assay and typing molecular markers of resistance to drug components of ACT currently used for treatment. Methods: The ex vivo susceptibility of 170 P. falciparum isolates to chloroquine, amodiaquine, lumefantrine, artesunate, and artemether was determined using the DAPI ex vivo assay. The high resolution melting technique was used to genotype the pfmdr1 gene at codons 86, 184 and 1246. Results: A significant decrease in IC50 values was observed between 2012 and 2013: from 13.84 to 6.484 for amodiaquine, 173.4 to 113.2 for lumefantrine, and 39.72 to 18.29 for chloroquine, respectively. Increase of the wild haplotype NYD and the decrease of the mutant haplotype NFD (79 and 62.26 %) was also observed. A correlation was observed between the wild type allele Y184 in pfmdr1 and higher IC50 for all drugs, except amodiaquine. Conclusion: This study has shown an increase in sensitivity over the span of two transmission seasons, marked by an increase in the WT alleles at pfmdr1. Continuous the monitoring of the ACT used for treatment of uncomplicated malaria will be helpful

Field Tour Guide for the CCAFS Independant Science Panel Meeting

CSA initiatives aim to sustainably increase productivity, enhance resilience, and reduce/remove greenhouse gases (GHGs), and require planning to address trade-offs and synergies between these three pillars: productivity, adaptation, and mitigation (CIAT; BFS/USAID. 2016). The CSV concept reflects an ambition to improve the integration of agriculture development and climate responsiveness. It is an approach that requires site-spe- cific monitoring and assessments of the social, economic and environmental conditions to identify appropriate agricultural production technologies and practices. In the Daga-Birame village, an initial diagnosis using the TOP-SECAC toolkit (Somda et al., 2011) was conducted starting by an analysis of vulnerability and adaptation capacity to climate change. It entailed participatory drawing of a map on livelihood resources and climate hazards in order to locate them in the territory under the community management plan. Then a vulnerability matrix was developed to analyze the perception of village commu- nity members about the level of influence of climatic hazards on main livelihood resources. Finally, a matrix of adaptation strategies was used to help identify and analyze (current and future) adaptation strategies. It included the level of importance of main livelihood resources in the implementation of the defined strategies. This identification of local constraints and opportunities was key to designing the CSV of Daga-Birame. Based on the village vision of its future, a set of actions have been identified by the community in order to achieve the desired changes in agricultural productivity and food security, income genera- tion, resilience improvement and sustainable management of the environment in the village. These actions were structured around four components: Climate Information Services (CIS); development of climate-smart agriculture technologies/practices; village development planning and local knowledge and institutions

Investigating the etiologies of non-malarial febrile illness in Senegal using metagenomic sequencing

Abstract The worldwide decline in malaria incidence is revealing the extensive burden of non-malarial febrile illness (NMFI), which remains poorly understood and difficult to diagnose. To characterize NMFI in Senegal, we collected venous blood and clinical metadata in a cross-sectional study of febrile patients and healthy controls in a low malaria burden area. Using 16S and untargeted sequencing, we detected viral, bacterial, or eukaryotic pathogens in 23% (38/163) of NMFI cases. Bacteria were the most common, with relapsing fever Borrelia and spotted fever Rickettsia found in 15.5% and 3.8% of cases, respectively. Four viral pathogens were found in a total of 7 febrile cases (3.5%). Sequencing also detected undiagnosed Plasmodium, including one putative P. ovale infection. We developed a logistic regression model that can distinguish Borrelia from NMFIs with similar presentation based on symptoms and vital signs (F1 score: 0.823). These results highlight the challenge and importance of improved diagnostics, especially for Borrelia, to support diagnosis and surveillance

Quality assessment of malaria microscopic diagnosis at the Aristide Le Dantec University Hospital of Dakar, Senegal, in 2020

Abstract Background Following WHO guidelines, microscopy is the gold standard for malaria diagnosis in endemic countries. The Parasitology-Mycology laboratory (LPM) is the National Reference Laboratory and is currently undergoing ISO 15189 accreditation. In this context, we assessed the performance of the laboratory by confirming the reliability and the accuracy of results obtained in accordance with the requirements of the ISO 15189 standards. This study aimed to verify the method of microscopic diagnosis of malaria at the LPM, in the Aristide Le Dantec hospital (HALD) in Dakar, Senegal. Methods This is a validation/verification study conducted from June to August 2020. Twenty (20) microscopic slides of thick/thin blood smear with known parasite densities (PD) selected from the Cheick Anta Diop University malaria slide bank in Dakar were used for this assessment. Six (6) were used to assess microscopists’ ability to determine PD and fourteen (14) slides were used for detection (positive vs negative) and identification of parasites. Four (4) LPM-HALD microscopists read and recorded their results on prepared sheets. Data analysis was done with Microsoft Excel 2010 software. Results A minimum threshold of 50% concordance was used for comparison. Of the twenty (20) slides read, 100% concordance was obtained on eight (8) detection (positive vs negative) slides. Four (4) out of the six (6) parasite density evaluation slides obtained a concordance of less than 50%. Thirteen (13) out of the fourteen (14) identification slides obtained a concordance greater than 50%. Only one (1) identification slide obtained zero agreement from the microscopists. For species identification a concordance greater than 80% was noted and the microscopists obtained scores between 0.20 and 0.4 on a scale of 0 to 1 for parasite density reading. The microscopists obtained 100% precision, sensitivity, specificity and both negative and positive predictive values. Conclusion This work demonstrated that the microscopic method of malaria diagnosis used in the LPM/HALD is in accordance with the requirements of WHO and ISO 15189. Further training of microscopists may be needed to maintain competency