Meta-analysis of the global distribution of clinically relevant CYP2C8 alleles and their inferred functional consequences.

BACKGROUND: CYP2C8 is responsible for the metabolism of 5% of clinically prescribed drugs, including antimalarials, anti-cancer and anti-inflammatory drugs. Genetic variability is an important factor that influences CYP2C8 activity and modulates the pharmacokinetics, efficacy and safety of its substrates. RESULTS: We profiled the genetic landscape of CYP2C8 variability using data from 96 original studies and data repositories that included a total of 33,185 unrelated participants across 44 countries and 43 ethnic groups. The reduced function allele CYP2C8*2 was most common in West and Central Africa with frequencies of 16-36.9%, whereas it was rare in Europe and Asia (< 2%). In contrast, CYP2C8*3 and CYP2C8*4 were common throughout Europe and the Americas (6.9-19.8% for *3 and 2.3-7.5% for *4), but rare in African and East Asian populations. Importantly, we observe pronounced differences (> 2.3-fold) between neighboring countries and even between geographically overlapping populations. Overall, we found that 20-60% of individuals in Africa and Europe carry at least one CYP2C8 allele associated with reduced metabolism and increased adverse event risk of the anti-malarial amodiaquine. Furthermore, up to 60% of individuals of West African ancestry harbored variants that reduced the clearance of pioglitazone, repaglinide, paclitaxel and ibuprofen. In contrast, reduced function alleles are only found in < 2% of East Asian and 8.3-12.8% of South and West Asian individuals. CONCLUSIONS: Combined, the presented analyses mapped the genetic and inferred functional variability of CYP2C8 with high ethnogeographic resolution. These results can serve as a valuable resource for CYP2C8 allele frequencies and distribution estimates of CYP2C8 phenotypes that could help identify populations at risk upon treatment with CYP2C8 substrates. The high variability between ethnic groups incentivizes high-resolution pharmacogenetic profiling to guide precision medicine and maximize its socioeconomic benefits, particularly for understudied populations with distinct genetic profiles.

Population-specific variations in KCNH2 predispose patients to delayed ventricular repolarization upon dihydroartemisinin-piperaquine therapy.

Dihydroartemisinin-piperaquine is efficacious for the treatment of uncomplicated malaria and its use is increasing globally. Despite the positive results in fighting malaria, inhibition of the Kv11.1 channel (hERG; encoded by the KCNH2 gene) by piperaquine has raised concerns about cardiac safety. Whether genetic factors could modulate the risk of piperaquine-mediated QT prolongations remained unclear. Here, we first profiled the genetic landscape of KCNH2 variability using data from 141,614 individuals. Overall, we found 1,007 exonic variants distributed over the entire gene body, 555 of which were missense. By optimizing the gene-specific parametrization of 16 partly orthogonal computational algorithms, we developed a KCNH2-specific ensemble classifier that identified a total of 116 putatively deleterious missense variations. To evaluate the clinical relevance of KCNH2 variability, we then sequenced 293 Malian patients with uncomplicated malaria and identified 13 variations within the voltage sensing and pore domains of Kv11.1 that directly interact with channel blockers. Cross-referencing of genetic and electrocardiographic data before and after piperaquine exposure revealed that carriers of two common variants, rs1805121 and rs41314375, experienced significantly higher QT prolongations (ΔQTc of 41.8 ms and 61 ms, respectively, vs 14.4 ms in controls) with more than 50% of carriers having increases in QTc >30 ms. Furthermore, we identified three carriers of rare population-specific variations who experienced clinically relevant delayed ventricular repolarization. Combined, our results map population-scale genetic variability of KCNH2 and identify genetic biomarkers for piperaquine-induced QT prolongation that could help to flag at-risk patients and optimize efficacy and adherence to antimalarial therapy.

Safety and efficacy of pyronaridine-artesunate paediatric granules in the treatment of uncomplicated malaria in children: insights from randomized clinical trials and a real-world study.

BACKGROUND: Children are particularly at risk of malaria. This analysis consolidates the clinical data for pyronaridine-artesunate (PA) paediatric granules in children from three randomized clinical trials and a real-world study (CANTAM). METHODS: An integrated safety analysis of individual patient data from three randomized clinical trials included patients with microscopically-confirmed Plasmodium falciparum, body weight ≥ 5 kg to < 20 kg, who received at least one dose of study drug (paediatric safety population). PA was administered once daily for 3 days; two trials included the comparator artemether-lumefantrine (AL). PCR-adjusted day 28 adequate clinical and parasitological response (ACPR) was evaluated. Real-world PA granules safety and effectiveness was also considered. RESULTS: In the integrated safety analysis, 63.9% (95% CI 60.2, 67.4; 426/667) of patients had adverse events following PA and 62.0% (95% CI 56.9, 66.9; 222/358) with AL. Vomiting was more common with PA (7.8% [95% CI 6.0, 10.1; 52/667]) than AL (3.4% [95% CI 1.9, 5.8; 12/358]), relative risk 2.3 (95% CI 1.3, 4.3; P = 0.004), occurring mainly following the first PA dose (6.7%, 45/667), without affecting re-dosing or adherence. Prolonged QT interval occurred less frequently with PA (3.1% [95% CI 2.1, 4.8; 21/667]) than AL (8.1% [95% CI 5.7, 11.4; 29/358]), relative risk 0.39 (95% CI 0.22, 0.67; P = 0.0007). In CANTAM, adverse events were reported for 17.7% (95% CI 16.3, 19.2; 460/2599) of patients, most commonly vomiting (5.4% [95% CI 4.6, 6.4; 141/2599]), mainly following the first dose, (4.5% [117/2599]), with all patients successfully re-dosed, and pyrexia (5.4% [95% CI 4.6, 6.3; 140/2599]). In the two comparative clinical trials, Day 28 ACPR in the per-protocol population for PA was 97.1% (95% CI 94.6, 98.6; 329/339) and 100% (95% CI 99.3, 100; 514/514) versus 98.8% (95% CI 95.7, 99.9; 165/167) and 98.4% (95% CI 95.5, 99.7; 188/191) for AL, respectively. In CANTAM, PA clinical effectiveness was 98.0% (95% CI 97.3, 98.5; 2273/2320). CONCLUSIONS: Anti-malarial treatment with PA paediatric granules administered once daily for 3 days was well tolerated in children and displayed good clinical efficacy in clinical trials, with effectiveness confirmed in a real-world study. Trial registration Clinicaltrials.gov: SP-C-003-05: identifier NCT00331136; SP-C-007-07: identifier NCT0541385; SP-C-021-15: identifier NCT03201770. Pan African Clinical Trials Registry: SP-C-013-11: identifier PACTR201105000286876.

High frequency of antimicrobial resistance in Salmonella and Escherichia coli causing diarrheal diseases at the Yirimadio community health facility, Mali.

BACKGROUND: Diarrhoea is a public health problem, especially in developing countries where it is the second leading cause of child mortality. In Low Income Countries like in Mali, self-medication and inappropriate use of antibiotics due to the scarcity of complementary diagnostic systems can lead to the development of multidrug-resistant bacteria causing diarrhoea. The objective of this work was to determine the microorganisms responsible for diarrhoea in children under 15 years of age and to characterize their sensitivity to a panel of antibiotics used in a peri-urban community in Mali. The study involved outpatient children visiting the Yirimadio Community Health Centre and diagnosed with diarrhoea. Stool samples from those patients were collected and analysed by conventional stools culture and the susceptibility to antibiotics of detected bacteria was determined by the disc diffusion method in an agar medium. RESULT: Overall, 554 patients were included. Children under the age of 3 years accounted for 88.8% (492 of 554) of our study population. Two bacterial species were isolated in this study, Escherichia coli 31.8% (176 of 554) and Salmonella 2.9% (16 of 554). In the 176, E. coli strains resistance to amoxicillin and to cotrimoxazole was seen in 93.8% (165 of 176) and 92.6% ( 163 of 176), respectively. The ESBL resistance phenotype accounted for 39,8% (70 of 176) of E. coli. Sixteen (16) strains of Salmonella were found, of which one strain (6.3%) was resistant to amoxicillin and to amoxicillin + clavulanic acid. Another one was resistant to chloramphenicol (6.3%). Two strains of Salmonella were resistant to cotrimoxazole (12.5%) and two others were resistant to cefoxitin (12.5%). CONCLUSIONS: The data suggest that E. coli is frequently involved in diarrhoea in children under 3 years of age in this peri-urban setting of Bamako, Mali, with a high rate of resistance to amoxicillin and cotrimoxazole, the most widely used antibiotics in the management of diarrhoea in this setting.

Seasonal vaccination with RTS,S/AS01E vaccine with or without seasonal malaria chemoprevention in children up to the age of 5 years in Burkina Faso and Mali: a double-blind, randomised, controlled, phase 3 trial.

BACKGROUND: Seasonal vaccination with the RTS,S/AS01E vaccine combined with seasonal malaria chemoprevention (SMC) prevented malaria in young children more effectively than either intervention given alone over a 3 year period. The objective of this study was to establish whether the added protection provided by the combination could be sustained for a further 2 years. METHODS: This was a double-blind, individually randomised, controlled, non-inferiority and superiority, phase 3 trial done at two sites: the Bougouni district and neighbouring areas in Mali and Houndé district, Burkina Faso. Children who had been enrolled in the initial 3-year trial when aged 5-17 months were initially randomly assigned individually to receive SMC with sulphadoxine-pyrimethamine and amodiaquine plus control vaccines, RTS,S/AS01E plus placebo SMC, or SMC plus RTS,S/AS01E. They continued to receive the same interventions until the age of 5 years. The primary trial endpoint was the incidence of clinical malaria over the 5-year trial period in both the modified intention-to-treat and per-protocol populations. Over the 5-year period, non-inferiority was defined as a 20% increase in clinical malaria in the RTS,S/AS01E-alone group compared with the SMC alone group. Superiority was defined as a 12% difference in the incidence of clinical malaria between the combined and single intervention groups. The study is registered with ClinicalTrials.gov, NCT04319380, and is complete. FINDINGS: In April, 2020, of 6861 children originally recruited, 5098 (94%) of the 5433 children who completed the initial 3-year follow-up were re-enrolled in the extension study. Over 5 years, the incidence of clinical malaria per 1000 person-years at risk was 313 in the SMC alone group, 320 in the RTS,S/AS01E-alone group, and 133 in the combined group. The combination of RTS,S/AS01E and SMC was superior to SMC (protective efficacy 57·7%, 95% CI 53·3 to 61·7) and to RTS,S/AS01E (protective efficacy 59·0%, 54·7 to 62·8) in preventing clinical malaria. RTS,S/AS01E was non-inferior to SMC (hazard ratio 1·03 [95% CI 0·95 to 1·12]). The protective efficacy of the combination versus SMC over the 5-year period of the study was very similar to that seen in the first 3 years with the protective efficacy of the combination versus SMC being 57·7% (53·3 to 61·7) and versus RTS/AS01E-alone being 59·0% (54·7 to 62·8). The comparable figures for the first 3 years of the study were 62·8% (58·4 to 66·8) and 59·6% (54·7 to 64·0%), respectively. Hospital admissions for WHO-defined severe malaria were reduced by 66·8% (95% CI 40·3 to 81·5), for malarial anaemia by 65·9% (34·1 to 82·4), for blood transfusion by 68·1% (32·6 to 84·9), for all-cause deaths by 44·5% (2·8 to 68·3), for deaths excluding external causes or surgery by 41·1% (-9·2 to 68·3), and for deaths from malaria by 66·8% (-2·7 to 89·3) in the combined group compared with the SMC alone group. No safety signals were detected. INTERPRETATION: Substantial protection against malaria was sustained over 5 years by combining seasonal malaria vaccination with seasonal chemoprevention, offering a potential new approach to malaria control in areas with seasonal malaria transmission. FUNDING: UK Joint Global Health Trials and PATH's Malaria Vaccine Initiative (through a grant from the Bill & Melinda Gates Foundation). TRANSLATION: For the French translation of the abstract see Supplementary Materials section.

Imidazolopiperazine (IPZ)-Induced Differential Transcriptomic Responses on Plasmodium falciparum Wild-Type and IPZ-Resistant Mutant Parasites.

Imidazolopiperazine (IPZ), KAF156, a close analogue of GNF179, is a promising antimalarial candidate. IPZ is effective against Plasmodium falciparum and Plasmodium vivax clinical malaria in human with transmission blocking property in animal models and effective against liver stage parasites. Despite these excellent drug efficacy properties, in vitro parasites have shown resistance to IPZ. However, the mechanism of action and resistance of IPZ remained not fully understood. Here, we used transcriptomic analysis to elucidate mode of action of IPZs. We report, in wild-type parasites GNF179 treatment down regulated lipase enzymes, two metabolic pathways: the hydrolysis of Phosphoinositol 4,5-bipohosphate (PIP2) that produce diacyglycerol (DAG) and the cytosolic calcium Ca2+ homeostasis which are known to be essential for P. falciparum survival and proliferation, as well for membrane permeability and protein trafficking. Furthermore, in wild-type parasites, GNF179 repressed expression of Acyl CoA Synthetase, export lipase 1 and esterase enzymes. Thus, in wild-type parasites only, GNF179 treatment affected enzymes leading lipid metabolism, transport, and synthesis. Lastly, our data revealed that IPZs did not perturb known IPZ resistance genes markers pfcarl, pfact, and pfugt regulations, which are all instead possibly involved in the drug resistance that disturb membrane transport targeted by IPZ.

Variation in neutrophil levels and artemisinin-based combination therapy efficacy in West-Africa.

INTRODUCTION: Polymorphonuclear neutrophils (PMN) are involved in pathogen clearance by phagocytosis. However, the role of PMNs in the efficacy of artemisinin-based combination therapy (ACT) is poorly understood. METHODOLOGY: In a prospective longitudinal in vivo study, neutrophil rates were compared with malaria carriage after treatment with different ACTs: Artemether - lumefantrine (AL), Artesunate - amodiaquine (ASAQ), Dihydroartemisinin - piperaquine (DP) or Pyronaridine artesunate (PA). The study cases were classified as having neutropenia, normal neutrophil levels or neutrophilia depending on the level of neutrophils in the blood. This study included 3148 patients and was analyzed using R. RESULTS: On day 7, only four patients in the neutropenia group and treated with AL had a malaria positive blood smear based on microscopy. On day 28, the rate of recurrent parasitemia in the AL arm was significantly higher in neutropenia patients (50.9%) than in patients with normal rates of neutrophils (43.1%) or in those with neutrophilia (6.0%) (p < 0.001). In ASAQ arm, the rate of recurrent Plasmodium falciparum parasitemia was 58.8% in the neutropenia group versus 29.4% in patients with normal rates of neutrophils and 11.8% in patients with neutrophilia (p < 0.001). No patient treated with DP with normal neutrophil counts or neutrophilia was carrying malaria parasites on day 28. Among the 15 patients with parasitemia on day 28 in the PA arm, 11 (73.33%) had neutropenia while 4 (26.67%) had a normal neutrophil count (p < 0.001). CONCLUSIONS: Patients with neutropenia had higher rates of recurrent P. falciparum parasitemia after ACT.

The Prevalence of Human Plasmodium Species during Peak Transmission Seasons from 2016 to 2021 in the Rural Commune of Ntjiba, Mali.

Up-to-date knowledge of key epidemiological aspects of each Plasmodium species is necessary for making informed decisions on targeted interventions and control strategies to eliminate each of them. This study aims to describe the epidemiology of plasmodial species in Mali, where malaria is hyperendemic and seasonal. Data reports collected during high-transmission season over six consecutive years were analyzed to summarize malaria epidemiology. Malaria species and density were from blood smear microscopy. Data from 6870 symptomatic and 1740 asymptomatic participants were analyzed. The median age of participants was 12 years, and the sex ratio (male/female) was 0.81. Malaria prevalence from all Plasmodium species was 65.20% (95% CI: 60.10-69.89%) and 22.41% (CI: 16.60-28.79%) for passive and active screening, respectively. P. falciparum was the most prevalent species encountered in active and passive screening (59.33%, 19.31%). This prevalence was followed by P. malariae (1.50%, 1.15%) and P. ovale (0.32%, 0.06%). Regarding frequency, P. falciparum was more frequent in symptomatic individuals (96.77% vs. 93.24%, p = 0.014). In contrast, P. malariae was more frequent in asymptomatic individuals (5.64% vs. 2.45%, p < 0.001). P. ovale remained the least frequent species (less than 1%), and no P. vivax was detected. The most frequent coinfections were P. falciparum and P. malariae (0.56%). Children aged 5-9 presented the highest frequency of P. falciparum infections (41.91%). Non-falciparum species were primarily detected in adolescents (10-14 years) with frequencies above 50%. Only P. falciparum infections had parasitemias greater than 100,000 parasites per µL of blood. P. falciparum gametocytes were found with variable prevalence across age groups. Our data highlight that P. falciparum represented the first burden, but other non-falciparum species were also important. Increasing attention to P. malariae and P. ovale is essential if malaria elimination is to be achieved.

Propensity of selecting mutant parasites for the antimalarial drug cabamiquine.

We report an analysis of the propensity of the antimalarial agent cabamiquine, a Plasmodium-specific eukaryotic elongation factor 2 inhibitor, to select for resistant Plasmodium falciparum parasites. Through in vitro studies of laboratory strains and clinical isolates, a humanized mouse model, and volunteer infection studies, we identified resistance-associated mutations at 11 amino acid positions. Of these, six (55%) were present in more than one infection model, indicating translatability across models. Mathematical modelling suggested that resistant mutants were likely pre-existent at the time of drug exposure across studies. Here, we estimated a wide range of frequencies of resistant mutants across the different infection models, much of which can be attributed to stochastic differences resulting from experimental design choices. Structural modelling implicates binding of cabamiquine to a shallow mRNA binding site adjacent to two of the most frequently identified resistance mutations.

Plasmodium falciparum Development from Gametocyte to Oocyst: Insight from Functional Studies.

Malaria elimination may never succeed without the implementation of transmission-blocking strategies. The transmission of Plasmodium spp. parasites from the human host to the mosquito vector depends on circulating gametocytes in the peripheral blood of the vertebrate host. Once ingested by the mosquito during blood meals, these sexual forms undergo a series of radical morphological and metabolic changes to survive and progress from the gut to the salivary glands, where they will be waiting to be injected into the vertebrate host. The design of effective transmission-blocking strategies requires a thorough understanding of all the mechanisms that drive the development of gametocytes, gametes, sexual reproduction, and subsequent differentiation within the mosquito. The drastic changes in Plasmodium falciparum shape and function throughout its life cycle rely on the tight regulation of stage-specific gene expression. This review outlines the mechanisms involved in Plasmodium falciparum sexual stage development in both the human and mosquito vector, and zygote to oocyst differentiation. Functional studies unravel mechanisms employed by P. falciparum to orchestrate the expression of stage-specific functional products required to succeed in its complex life cycle, thus providing us with potential targets for developing new therapeutics. These mechanisms are based on studies conducted with various Plasmodium species, including predominantly P. falciparum and the rodent malaria parasites P. berghei. However, the great potential of epigenetics, genomics, transcriptomics, proteomics, and functional genetic studies to improve the understanding of malaria as a disease remains partly untapped because of limitations in studies using human malaria parasites and field isolates.

Complexity of Plasmodium falciparum infection and genetic variations associated with differences in parasite clearance time in two Malian villages.

Background: Effective approaches to fight against malaria include disease prevention, an early diagnosis of malaria cases, and rapid management of confirmed cases by treatment with effective antimalarials. Artemisinin-based combination therapies are first-line treatments for uncomplicated malaria in endemic areas. However, cases of resistance to artemisinin have already been described in South-East Asia resulting in prolonged parasite clearance time after treatment. In Mali, though mutations in the K13 gene associated with delayed clearance in Asia are absent, a significant difference in parasite clearance time following treatment with artesunate was observed between two malaria endemic sites, Bougoula-Hameau and Faladje. Hypothetically, differences in complexity of Plasmodium falciparum infections may be accounted for this difference. Hence, the aims of this study were to assess the complexity of infection (COI) and genetic diversity of P. falciparum parasites during malaria treatment in Bougoula-Hameau and Faladje in Mali. Methods: Thirty (30) patients per village were randomly selected from 221 patients enrolled in a prospective artesunate monotherapy study conducted in Faladje and Bougoula-Hameau in 2016. All parasitemic blood samples of patients from enrollment to last positive slide were retained to assess malaria parasite COI and polymorphisms. DNA were extracted with a Qiagen kit and Pfcsp and Pfama1 encoding gene were amplified by nested PCR and sequenced using the Illumina platform. The parasite clearance time (PCT) was determined using the parasite clearance estimator of Worldwide Antimarial Resistance Network (WWARN). Data were analyzed with R®. Results: The median number of genetically distinct parasite clones was similar at enrollment, 7 (IQR of 5-9) in Faladje and 6 (IQR of 4-10) in Bougoula-Hameau (p-value = 0.1). On the first day after treatment initiation, the COI was higher in Faladje (6; CI:4-8) than in Bougoula-Hameau (4; CI:4-6) with a p-value =0. 02. Overall, COI was high with higher PCT. Finally, there was a low genetic diversity between Faladje and Bougoula-Hameau. Conclusion: This study demonstrated that the difference in PCT observed between the two villages could be due to differences in the complexity of infection of these two villages.

Ethiopian Plasmodium vivax hypnozoites formation dynamics and their susceptibility to reference antimalarial drugs.

One of the key obstacles to malaria elimination is largely attributed to Plasmodium vivax's ability to form resilient hypnozoites in the host liver that cause relapsing infections. As a result, interruption of P. vivax transmission is difficult. P. vivax transmission occurs in Duffy-positive individuals and have been mainly thought to be absent in Africa. However, increasing studies using molecular tools detected P. vivax among Duffy-negative individuals in various African countries. Studies on the African P. vivax has been severely limited because most of malaria control program focus mainly on falciparum malaria. In addition, there is a scarcity of laboratory infrastructures to overcome the biological obstacles posed by P. vivax. Herein, we established field transmission of Ethiopian P. vivax for routine sporozoite supply followed by liver stage infection in Mali. Furthermore, we evaluated local P. vivax hypnozoites and schizonts susceptibilities to reference antimalarial drugs. The study enabled the assessment of local African P. vivax hypnozoite production dynamics. Our data displayed the ability of the African P. vivax to produce hypnozoite forms ex-vivo at different rates per field isolate. We report that while tafenoquine (1µM) potently inhibited both hypnozoites and schizont forms; atovaquone (0.25µM) and the phosphatidylinositol-4-OH kinase (PI4K)-specific inhibitor KDU691 (0.5µM) showed no activity against hypnozoites forms. Unlike hypnozoites forms, P. vivax schizont stages were fully susceptible to both atovaquone (0.25µM) and the (PI4K)-specific inhibitor KDU691 (0.5µM). Together, the data revealed the importance of the local platform for further biological investigation and implementation of drug discovery program on the African P. vivax clinical isolates.

SARS-CoV-2 seroprevalence and living conditions in Bamako (Mali): a cross-sectional multistage household survey after the first epidemic wave, 2020.

OBJECTIVES: In low-income settings with limited access to diagnosis, COVID-19 information is scarce. In September 2020, after the first COVID-19 wave, Mali reported 3086 confirmed cases and 130 deaths. Most reports originated from Bamako, with 1532 cases and 81 deaths (2.42 million inhabitants). This observed prevalence of 0.06% appeared very low. Our objective was to estimate SARS-CoV-2 infection among inhabitants of Bamako, after the first epidemic wave. We assessed demographic, social and living conditions, health behaviours and knowledges associated with SARS-CoV-2 seropositivity. SETTINGS: We conducted a cross-sectional multistage household survey during September 2020, in three neighbourhoods of the commune VI (Bamako), where 30% of the cases were reported. PARTICIPANTS: We recruited 1526 inhabitants in 3 areas, that is, 306 households, and 1327 serological results (≥1 years), 220 household questionnaires and collected answers for 962 participants (≥12 years). PRIMARY AND SECONDARY OUTCOME MEASURES: We measured serological status, detecting SARS-CoV-2 spike protein antibodies in blood sampled. We documented housing conditions and individual health behaviours through questionnaires among participants. We estimated the number of SARS-CoV-2 infections and deaths in the population of Bamako using the age and sex distributions. RESULTS: The prevalence of SARS-CoV-2 seropositivity was 16.4% (95% CI 15.1% to 19.1%) after adjusting on the population structure. This suggested that ~400 000 cases and ~2000 deaths could have occurred of which only 0.4% of cases and 5% of deaths were officially reported. Questionnaires analyses suggested strong agreement with washing hands but lower acceptability of movement restrictions (lockdown/curfew), and mask wearing. CONCLUSIONS: The first wave of SARS-CoV-2 spread broadly in Bamako. Expected fatalities remained limited largely due to the population age structure and the low prevalence of comorbidities. Improving diagnostic capacities to encourage testing and preventive behaviours, and avoiding the spread of false information remain key pillars, regardless of the developed or developing setting. ETHICS: This study was registered in the registry of the ethics committee of the Faculty of Medicine and Odonto-Stomatology and the Faculty of Pharmacy, Bamako, Mali, under the number: 2020/162/CA/FMOS/FAPH.

Comparative Susceptibility of Plasmodium ovale and Plasmodium falciparum Field Isolates to Reference and Lead Candidate Antimalarial Drugs in Ghana.

Malaria treatments resulted in the decline of the deadliest Plasmodium falciparum globally while species, such as P. ovale, infections have been increasingly detected across sub-Saharan Africa. Currently, no experimental drug sensitivity data are available to guide effective treatment and management of P. ovale infections, which is necessary for effective malaria elimination. We conducted a prospective study to evaluate P. ovale epidemiology over 1 year and determined ex vivo susceptibility of the field isolates to existing and lead advanced discovery antimalarial drugs. We report that while P. falciparum dominated both symptomatic and asymptomatic malaria cases, P. ovale in mono or co-infections caused 7.16% of symptomatic malaria. Frontline antimalarials artesunate and lumefantrine inhibited P. ovale as potently as P. falciparum. Chloroquine, which has been withdrawn in Ghana, was also highly inhibitory against both P. ovale and P. falciparum. In addition, P. ovale and P. falciparum displayed high susceptibility to quinine, comparable to levels observed with chloroquine. Pyrimethamine, which is a major drug for disease massive prevention, also showed great inhibition of P. ovale, comparable to effects on P. falciparum. Furthermore, we identified strong inhibition of P. ovale using GNF179, a close analogue of KAF156 imidazolopiperazines, which is a novel class of antimalarial drugs currently in clinical phase II testing. We further demonstrated that the Plasmodium phosphatidylinositol-4-OH kinase (PI4K)-specific inhibitor, KDU691, is highly inhibitory against P. ovale and P. falciparum field isolates. Our data indicated that existing and lead advanced discovery antimalarial drugs are suitable for the treatment of P. ovale infections in Ghana. IMPORTANCE Current malaria control and elimination tools such as drug treatments are not specifically targeting P.ovale. P. ovale can form hypnozoite and cause relapsing malaria. P. ovale is the third most dominant species in Africa and requires radical cure treatment given that it can form liver dormant forms called hypnozoites that escape all safe treatments. The inappropriate treatment of P. ovale would sustain its transmission in Africa where the medical need is the greatest. This is a hurdle for successful malaria control and elimination. Here, we provided experiment data that were lacking to guide P. ovale treatment and disease control policy makers using reference antimalarial drugs. We also provided key experimental data for 2 clinical candidate drugs that can be used for prioritization selection of lead candidate's identification for clinical development.

Pf7: an open dataset of Plasmodium falciparum genome variation in 20,000 worldwide samples.

We describe the MalariaGEN Pf7 data resource, the seventh release of Plasmodium falciparum genome variation data from the MalariaGEN network.  It comprises over 20,000 samples from 82 partner studies in 33 countries, including several malaria endemic regions that were previously underrepresented.  For the first time we include dried blood spot samples that were sequenced after selective whole genome amplification, necessitating new methods to genotype copy number variations.  We identify a large number of newly emerging crt mutations in parts of Southeast Asia, and show examples of heterogeneities in patterns of drug resistance within Africa and within the Indian subcontinent.  We describe the profile of variations in the C-terminal of the csp gene and relate this to the sequence used in the RTS,S and R21 malaria vaccines.  Pf7 provides high-quality data on genotype calls for 6 million SNPs and short indels, analysis of large deletions that cause failure of rapid diagnostic tests, and systematic characterisation of six major drug resistance loci, all of which can be freely downloaded from the MalariaGEN website.

IFNγ, TNFα polymorphisms and IFNγ serum levels are associated with the clearance of drug-resistant P. falciparum in Malian children.

Host immunity has been suggested to clear drug-resistant parasites in malaria-endemic settings. However, the immunogenetic mechanisms involved in parasite clearance are poorly understood. Characterizing the host's immunity and genes involved in controlling the parasitic infection can inform the development of blood-stage malaria vaccines. This study investigates host regulatory cytokines and immunogenomic factors associated with the clearance of Plasmodium falciparum carrying a chloroquine resistance genotype. Biological samples from participants of previous drug efficacy trials conducted in two Malian localities were retrieved. The P. falciparum chloroquine resistance transporter (Pfcrt) gene was genotyped using parasite DNA. Children carrying parasites with the mutant allele (Pfcrt-76T) were classified based on their ability to clear their parasites. The levels of the different cytokines were measured in serum. The polymorphisms of specific human genes involved in malaria susceptibility were genotyped using human DNA. The prevalence of the Pfcrt-76T was significantly higher in Kolle than in Bandiagara (81.6 % vs 38.6 %, p < 10-6). The prevalence of children who cleared their mutant parasites was significantly higher in Bandiagara than in Kolle (82.2 % vs 67.4 %, p < 0.05). The genotyping of host genes revealed that IFN-γ -874 T and TNF-α -308A alleles were positively associated with parasite clearance. Cytokine profiling revealed that IFN-γ level was positively associated with parasite clearance (p = 0.04). This study highlights the role of host's immunity and immunogenetic factors to clear resistant parasites, suggesting further characterization of these polymorphisms may help to develop novel approaches to antiparasitic treatment strategies.

The estimated burden of fungal diseases in Mali.

Mali is a developing country facing several health challenges with a high rate of tuberculosis (TB) and a moderate HIV infection burden. Little is known or done about fungal diseases, yet they represent a significant public health problem in certain populations. The aim of this study was to estimate the national burden of fungal disease, and summarize data, diagnostic and treatment gaps. We used national demographics and PubMed searches to retrieve articles on published data on these infections and at-risk populations (pulmonary TB, HIV/AIDS patients, patients receiving critical care etc.) in Mali. The estimated Malian population was 21,251,000 in 2020 (UN), of which 45% were children <14 years. Among HIV patients, we estimate an annual incidence of 611 cryptococcosis, 1393 Pneumocystis pneumonia, 180 histoplasmosis and >5,700 esophageal candidiasis and some microsporidiosis cases. Our prevalence estimates for tinea capitis are 2.3 million, for recurrent vulvovaginal candidiasis 272,460, ∼60,000 fungal asthma and 7,290 cases of chronic pulmonary aspergillosis (often mistaken for TB). Less common acute fungal infections are probably invasive aspergillosis (n=1230), fungal keratitis (n=2820), candidaemia (>1,060) and mucormycosis (n=43). Histoplasmin was found in 6% in general population. A few cases of mycetoma are described in Mali. Many WHO Essential medicines and Diagnostics are not available in Mali. This shows a marked disparity in documented and estimated cases of fungal diseases in Mali. These infections are underestimated due to the lack of accurate diagnosis tools and lack of support for fungal diseases diagnosis and management.

Plasmodium malariae structure and genetic diversity in sub-Saharan Africa determined from microsatellite variants and linked SNPs in orthologues of antimalarial resistance genes.

Plasmodium malariae, a neglected human malaria parasite, contributes up to 10% of malaria infections in sub-Saharan Africa (sSA). Though P. malariae infection is considered clinically benign, it presents mostly as coinfections with the dominant P. falciparum. Completion of its reference genome has paved the way to further understand its biology and interactions with the human host, including responses to antimalarial interventions. We characterized 75 P. malariae isolates from seven endemic countries in sSA using highly divergent microsatellites. The P. malariae infections were highly diverse and five subpopulations from three ancestries (independent of origin of isolates) were determined. Sequences of 11 orthologous antimalarial resistance genes, identified low frequency single nucleotide polymorphisms (SNPs), strong linkage disequilibrium between loci that may be due to antimalarial drug selection. At least three sub-populations were detectable from a subset of denoised SNP data from mostly the mitochondrial cytochrome b coding region. This evidence of diversity and selection calls for including P. malariae in malaria genomic surveillance towards improved tools and strategies for malaria elimination.