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Summary: Background: Plasmodium blood-stage parasites balance asexual multiplication with gametocyte development. Few studies link these dynamics with parasite genetic markers in vivo; even fewer in longitudinally monitored infections. Environmental influences on gametocyte formation, such as mosquito exposure, may influence the parasite's investment in gametocyte production. Methods: We investigated gametocyte production and asexual multiplication in two Plasmodium falciparum infected populations; a controlled human malaria infection (CHMI) study and a 28-day observational study in naturally infected individuals in Burkina Faso with controlled mosquito exposure. We measured gene transcript levels previously related to gametocyte formation (ap2-g, surfin1.2, surfin13.1, gexp-2) or inhibition of asexual multiplication (sir2a) and compared transcript levels to ring-stage parasite and mature gametocyte densities. Findings: Three of the five markers (ap2-g, surfin1.2, surfin13.1) predicted peak gametocytaemia in the CHMI study. An increase in all five markers in natural infections was associated with an increase in mature gametocytes 14 days later; the effect of sir2a on future gametocytes was strongest (fold change = 1.65, IQR = 1.22–2.24, P = 0.004). Mosquito exposure was not associated with markers of gametocyte formation (ap2-g P = 0.277; sir2a P = 0.499) or carriage of mature gametocytes (P = 0.379). Interpretation: All five parasite genetic markers predicted gametocyte formation over a single cycle of gametocyte formation and maturation in vivo; sir2a and ap2-g were most closely associated with gametocyte growth dynamics. We observed no evidence to support the hypothesis that exposure to Anopheles mosquito bites stimulates gametocyte formation. Funding: This work was funded by the Bill & Melinda Gates Foundation (INDIE OPP1173572), the European Research Council fellowship (ERC-CoG 864180) and UKRI Medical Research Council (MR/T016272/1) and Wellcome Center (218676/Z/19/Z).

The ACE2 gene polymorphisms (rs143936283, rs146676783, and rs4646116) in infected and noninfected persons by SARS-CoV-2 in Burkina Faso. Our cross-sectional study population comprised 137 SARS-CoV-2 infected persons and 181 non-infected persons. Three ACE2 gene polymorphisms rs143936283, rs146676783, and rs4646116, were genotyped using the real-time PCR standard TaqMan allelic discrimination technique. The association between SARS-CoV-2 infection and the polymorphisms were evaluated by a binary logistic regression. There was no association between the polymorphisms rs143936283, rs4646116 haplotypes, and SARS-CoV-2 infection in our study population. However, in the female population, the heterozygous genotype CT of rs146676783 increased by two and half the risk (OR=2.58 95%CI (1.2-5.48), p= 0.014) of being infected by SARS-CoV-2. Additionally, carrying the homozygous minor allele (genotype TT) of rs146676783 increased by more than five and half the risk (OR=5.57 95%CI (1.64-18.78), p=0.006) of being infected by SARS-CoV-2 among females. This study showed that the ACE2 gene variant rs146676783 was associated with an increased risk of being infected by SARS-CoV-2 in females, suggesting a need for further investigation to contribute to a better understanding of the African COVID-19 enigma.

Introduction Seasonal malaria chemoprevention (SMC) involves repeated administrations of sulfadoxine-pyrimethamine plus amodiaquine to children below the age of 5 years during the peak transmission season in areas of seasonal malaria transmission. While highly impactful in reducing Plasmodium falciparum malaria burden in controlled research settings, the impact of SMC on infection prevalence is moderate in real-life settings. It remains unclear what drives this efficacy decay. Recently, the WHO widened the scope for SMC to target all vulnerable populations. The Ministry of Health (MoH) in Burkina Faso is considering extending SMC to children below 10 years old. We aim to assess the impact of SMC on clinical incidence and parasite prevalence and quantify the human infectious reservoir for malaria in this population.Methods and analysis We will perform a cluster randomised trial in Saponé Health District, Burkina Faso, with three study arms comprising 62 clusters of three compounds: arm 1 (control): SMC in under 5-year-old children, implemented by the MoH without directly observed treatment (DOT) for the full course of SMC; arm 2 (intervention): SMC in under 5-year-old children, with DOT for the full course of SMC; arm 3 (intervention): SMC in under 10-year-old children, with DOT for the full course of SMC. The primary endpoint is parasite prevalence at the end of the malaria transmission season. Secondary endpoints include the impact of SMC on clinical incidence. Factors affecting SMC uptake, treatment adherence, drug concentrations, parasite resistance markers and transmission of parasites will be determined.Ethics and dissemination The London School of Hygiene & Tropical Medicine’s Ethics Committee (29193) and the Burkina Faso National Medical Ethics Committee (Deliberation No 2023-05-104) approved this study. The findings will be presented to the community; disease occurrence data and study outcomes will also be shared with the Burkina Faso MoH. Findings will be published irrespective of their results.Trial registration number NCT05878366.

BackgroundBK-SE36/CpG is a recombinant blood-stage malaria vaccine candidate based on the N-terminal Plasmodium falciparum serine repeat antigen5 (SE36), adsorbed to aluminium hydroxide gel and reconstituted, prior to administration, with synthetic oligodeoxynucleotides bearing CpG motifs. In healthy Japanese adult males, BK-SE36/CpG was well tolerated. This study assessed its safety and immunogenicity in healthy malaria-exposed African adults and children.MethodsA double-blind, randomised, controlled, age de-escalating clinical trial was conducted in an urban area of Ouagadougou, Burkina Faso. Healthy participants (n=135) aged 21-45 years (Cohort 1), 5-10 years (Cohort 2) and 12-24 months (Cohort 3) were randomised to receive three vaccine doses (Day 0, 28 and 112) of BK-SE36/CpG or rabies vaccine by intramuscular injection.ResultsOne hundred thirty-four of 135 (99.2%) subjects received all three scheduled vaccine doses. Vaccinations were well tolerated with no related Grade 3 (severe) adverse events (AEs). Pain/limitation of limb movement, headache in adults and fever in younger children (all mild to moderate in intensity) were the most frequently observed local and systemic AEs. Eighty-three of BK-SE36/CpG (91%) recipients and 37 of control subjects (84%) had Grade 1/2 events within 28 days post vaccination. Events considered by the investigator to be vaccine related were experienced by 38% and 14% of subjects in BK-SE36/CpG and control arms, respectively. Throughout the trial, six Grade 3 events (in 4 subjects), not related to vaccination, were recorded in the BK-SE36/CpG arm: 5 events (in 3 subjects) within 28 days of vaccination. All serious adverse events (SAEs) (n=5) were due to severe malaria (52-226 days post vaccination) and not related to vaccination. In all cohorts, BK-SE36/CpG arm had higher antibody titres after Dose 3 than after Dose 2. Younger cohorts had stronger immune responses (12–24-month-old > 5-10 years-old > 21-45 years-old). Sera predominantly reacted to peptides that lie in intrinsically unstructured regions of SE36. In the control arm, there were no marked fold changes in antibody titres and participants’ sera reacted poorly to all peptides spanning SE36.ConclusionBK-SE36/CpG was well-tolerated and immunogenic. These results pave the way for further proof-of-concept studies to demonstrate vaccine efficacy.Clinical trial registrationhttps://pactr.samrc.ac.za/TrialDisplay.aspx?TrialID=1921, PACTR201701001921166.

The regulation of immune cell responses to infection is a complex process that involves various molecular mechanisms, including epigenetic regulation. DNA methylation has been shown to play central roles in regulating gene expression and modulating cell response during infection. However, the nature and extent to which DNA methylation is involved in the host immune response in human malaria remains largely unknown. Here, we present a longitudinal study investigating the temporal dynamics of genome-wide in vivo DNA methylation profiles using 189 MethylationEPIC 850 K profiles from 66 children in Burkina Faso, West Africa, sampled three times: before infection, during symptomatic parasitemia, and after malaria treatment. The results revealed major changes in the DNA methylation profiles of children in response to both Plasmodium falciparum infection and malaria treatment, with widespread hypomethylation of CpGs upon infection (82% of 6.8 K differentially methylated regions). We document a remarkable reversal of CpG methylation profiles upon treatment to pre-infection states. These changes implicate divergence in core immune processes, including the regulation of lymphocyte, neutrophil, and myeloid leukocyte function. Integrative DNA methylation-mRNA analysis of a top differentially methylated region overlapping the pro-inflammatory gene TNF implicates DNA methylation of TNF cis regulatory elements in the molecular mechanisms of TNF regulation in human malaria. Our results highlight a central role of epigenetic regulation in mounting the host immune response to P. falciparum infection and in response to malaria treatment.

Abstract Background In Burkina Faso, malaria remains the first cause of medical consultation and hospitalization in health centres. First-line case management of malaria in the country’s health facilities is based on the use of artemisinin-based combination therapy (ACT). To optimize the use of these anti-malarial drugs in the perspective of mitigating the emergence of artemisinin resistance, which is a serious threat to malaria control and elimination, a pilot programme using multiple first-line therapies (MFTs) [three artemisinin-based combinations—pyronaridine–artesunate, dihydroartemisinin–piperaquine and artemether-lumefantrine] has been designed for implementation. As the success of this MFT pilot programme depends on the perceptions of key stakeholders in the health system and community members, the study aimed to assess their perceptions on the implementation of this strategy. Methods Semi-structured interviews, including 27 individual in-depth interviews and 41 focus groups discussions, were conducted with key stakeholders including malaria control policymakers and implementers, health system managers, health workers and community members. Volunteers from targets stakeholder groups were randomly selected. All interviews were recorded, transcribed and translated. Content analysis was performed using the qualitative software programme QDA Miner. Results The interviews revealed a positive perception of stakeholders on the implementation of the planned MFT programme. They saw the strategy as an opportunity to strengthen the supply of anti-malarial drugs and improve the management of fever and malaria. However, due to lack of experience with the products, health workers and care givers expressed some reservations about the effectiveness and side-effect profiles of the two anti-malarial drugs included as first-line therapy in the MFT programme (pyronaridine–artesunate, dihydroartemisinin–piperaquine). Questions were raised about the appropriateness of segmenting the population into three groups and assigning a specific drug to each group. Conclusion The adherence of both populations and key stakeholders to the MFT implementation strategy will likely depend on the efficacy of the proposed drugs, the absence of, or low frequency of, side-effects, the cost of drugs and availability of the different combinations.

Abstract Background Malaria case management relies on World Health Organization (WHO)-recommended artemisinin-based combination therapy (ACT), and a continuous understanding of local community knowledge, attitudes, and practices may be a great support for the success of malaria disease control efforts. In this context, this study aimed to identify potential facilitators or barriers at the community level to inform a health district-wide implementation of multiple first-line therapies (MFT) as a new strategy for uncomplicated malaria case management. Methods A community-based cross-sectional study using a mixed-method design was carried out from November 2018 to February 2019, in the health district (HD) of Kaya in Burkina Faso. Quantitative data were collected using a standardized questionnaire from 1394 individuals who had fever/malaria episodes four weeks prior to the survey. In addition, 23 focus group discussions (FGDs) were conducted targeting various segments of the community. Logistic regression models were used to assess the predictors of community care-seeking behaviours. Results Overall, 98% (1366/1394) of study participants sought advice or treatment, and 66.5% did so within 24 h of fever onset. 76.4% of participants preferred to seek treatment from health centres as the first recourse to care, 5.8% were treated at home with remaining drug stock, and 2.3% preferred traditional healers. Artemether-lumefantrine (AL) was by far the most used anti-malarial drug (98.2%); reported adherence to the 3-day treatment regimen was 84.3%. Multivariate analysis identified less than 5 km distance travelled for care (AOR = 2.7; 95% CI 2.1–3.7) and education/schooling (AOR = 1.8; 95% CI 1.3–2.5) as determinants of prompt care-seeking for fever. Geographical proximity (AOR = 1.5, 95% CI 1.2–2.1), having a child under five (AOR = 4.6, 95% CI 3.2–6.7), being pregnant (AOR = 6.5, 95% CI 1.9–22.5), and living in an urban area (AOR = 2.8, 95% CI 1.8–4.2) were significant predictors for visiting health centres. The FGDs showed that participants had good knowledge about malaria symptoms, prevention tools, and effective treatment. Behaviour change regarding malaria treatment and free medication for children under five were the main reasons for participants to seek care at health centres. Conclusions The study showed appropriate knowledge about malaria and positive community care-seeking behaviour at health centres for fever/malaria episodes. This could potentially facilitate the implementation of a MFT pilot programme in the district. ClinicalTrials.gov Identifier: NCT04265573.

Abstract Burkina Faso has one of the highest malaria burdens in sub-Saharan Africa despite the mass deployment of insecticide-treated nets (ITNs) and use of seasonal malaria chemoprevention (SMC) in children aged up to 5 years. Identification of risk factors for Plasmodium falciparum infection in rural Burkina Faso could help to identify and target malaria control measures. A cross-sectional survey of 1,199 children and adults was conducted during the peak malaria transmission season in the Cascades Region of south-west Burkina Faso in 2017. Logistic regression was used to identify risk factors for microscopically confirmed P. falciparum infection. A malaria transmission dynamic model was used to determine the impact on malaria cases averted of administering SMC to children aged 5–15 year old. P. falciparum prevalence was 32.8% in the study population. Children aged 5 to

Objectives: In 2017, the World Health Organisation (WHO) pre-qualified a single-dose typhoid conjugate vaccine (TCV) and identified TCV co-administration studies as a research priority. Accordingly, we tested co-administration of Typbar TCV® (Bharat Biotech International) with measles-rubella (MR) and yellow fever (YF) vaccines. Methods: We conducted a randomized, double-blind, and controlled, phase 2 trial in Ouagadougou, Burkina Faso. Healthy children aged 9–11 months were randomized 1:1 to receive TCV (Group 1) or control vaccine (inactivated polio vaccine (IPV), Group 2). Vaccines were administered intramuscularly with routine MR and YF vaccines. Safety was assessed by (1) local and systemic reactions on days 0, 3, and 7; (2) unsolicited adverse events within 28 days; and (3) serious adverse events (SAEs) within six months after immunization. Results: We enrolled, randomized, and vaccinated 100 eligible children (49 Group 1 and 51 Group 2). Safety outcomes occurred with similar frequency in both groups: local/solicited reactions (Group 1: 1/49, Group 2: 3/50), systemic/solicited reactions (Group 1: 4/49, Group 2: 9/50), unsolicited adverse events (Group 1: 26/49, Group 2: 33/51), and SAEs (Group 1: 2/49, Group 2: 3/51). TCV conferred robust immunogenicity without interference with MR or YF vaccines. Conclusion: TCV can be safely co-administered with MR and YF vaccines to children at the 9-month vaccination visit.

BackgroundA blood-stage vaccine targeting the erythrocytic-stages of the malaria parasite Plasmodium falciparum could play a role to protect against clinical disease. Antibodies against the P. falciparum serine repeat antigen 5 (SE47 and SE36 domains) correlate well with the absence of clinical symptoms in sero-epidemiological studies. A previous phase Ib trial of the recombinant SE36 antigen formulated with aluminum hydroxyl gel (BK-SE36) was promising. This is the first time the vaccine candidate was evaluated in young children below 5 years using two vaccination routes.MethodsSafety and immunogenicity of BK-SE36 was assessed in a double-blind, randomized, controlled, age de-escalating phase Ib trial. Fifty-four Burkinabe children in each age cohort, 25–60 or 12–24 months, were randomized in a 1:1:1 ratio to receive three doses of BK-SE36 either by intramuscular (BK IM) or subcutaneous (BK SC) route on Day 0, Week 4, and 26; or the control vaccine, Synflorix®via IM route on Day 0, Week 26 (and physiological saline on Week 4). Safety data and samples for immunogenicity analyses were collected at various time-points.ResultsOf 108 subjects, 104 subjects (96.3%) (Cohort 1: 94.4%; Cohort 2: 98.1%) received all three scheduled vaccine doses. Local reactions, mostly mild or of moderate severity, occurred in 99 subjects (91.7%). The proportion of subjects that received three doses without experiencing Grade 3 adverse events was similar across BK-SE36 vaccines and control arms (Cohort 1: 100%, 89%, and 89%; and Cohort 2: 83%, 82%, and 83% for BK IM, BK SC, and control, respectively). BK-SE36 vaccine was immunogenic, inducing more than 2-fold change in antibody titers from pre-vaccination, with no difference between the two vaccination routes. Titers waned before the third dose but in both cohorts titers were boosted 6 months after the first vaccination. The younger cohort had 2-fold and 4-fold higher geometric mean titers compared to the 25- to 60-month-old cohort after 2 and 3 doses of BK-SE36, respectively.ConclusionBK-SE36 was well tolerated and immunogenic using either intramuscular or subcutaneous routes, with higher immune response in the younger cohort.Clinical Trial Registrationhttps://pactr.samrc.ac.za/TrialDisplay.aspx?TrialID=934, identifier PACTR201411000934120.

Abstract Background For uncomplicated Plasmodium falciparum malaria, highly efficacious single-dose treatments are expected to increase compliance and improve treatment outcomes, and thereby may slow the development of resistance. The efficacy and safety of a single-dose combination of artefenomel (800 mg) plus ferroquine (400/600/900/1200 mg doses) for the treatment of uncomplicated P. falciparum malaria were evaluated in Africa (focusing on children ≤ 5 years) and Asia. Methods The study was a randomized, double-blind, single-dose, multi-arm clinical trial in patients aged > 6 months to 5 years and 20 Asian patients. None of the treatment arms met the target efficacy criterion for PCR-adjusted ACPR at Day 28 (lower limit of 95% confidence interval [CI] > 90%). PCR-adjusted ACPR at Day 28 [95% CI] in the PP Set ranged from 78.4% [64.7; 88.7%] to 91.7% [81.6; 97.2%] for the 400 mg to 1200 mg ferroquine dose. Efficacy rates were low in Vietnamese patients, ranging from 20 to 40%. A clear relationship was found between drug exposure (artefenomel and ferroquine concentrations at Day 7) and efficacy (primary endpoint), with higher concentrations of both drugs resulting in higher efficacy. Six distinct kelch-13 mutations were detected in parasite isolates from 10/272 African patients (with 2 mutations known to be associated with artemisinin resistance) and 18/20 Asian patients (all C580Y mutation). Vomiting within 6 h of initial artefenomel administration was common (24.6%) and associated with lower drug exposures. Conclusion The efficacy of artefenomel/ferroquine combination was suboptimal in African children aged ≤ 5 years, the population of interest, and vomiting most likely had a negative impact on efficacy. Trial registration ClinicalTrials.gov, NCT02497612. Registered 14 Jul 2015, https://clinicaltrials.gov/ct2/show/NCT02497612?term=NCT02497612&draw=2&rank=1

In this longitudinal study of an incident (new infections) and chronic (asymptomatic infections) cohort of Plasmodium falciparum infection in children in Burkina Faso, the authors show higher gametocyte production and mosquito infectivity in chronic infections.

Objectives: The World Health Organization pre-qualified single-dose typhoid conjugate vaccine (TCV) and requested data on co-administration with routine vaccines. The co-administration of Typbar TCV (Bharat Biotech International) with routine group A meningococcal conjugate vaccine (MCV-A) and measles–rubella (MR) vaccine was tested. Methods: This was a double-blind, randomized controlled trial performed in Ouagadougou, Burkina Faso. Children were recruited at the 15-month vaccination visit and were assigned randomly (1:1:1) to three groups. Group 1 children received TCV plus control vaccine (inactivated polio vaccine) and MCV-A 28 days later; group 2 children received TCV and MCV-A; group 3 children received MCV-A and control vaccine. Routine MR vaccine was administered to all participants. Safety was assessed at 0, 3, and 7 days after immunization, and unsolicited adverse events and serious adverse events were assessed for 28 days and 6 months after immunization, respectively. Results: A total of 150 children were recruited and vaccinated. Solicited symptoms were infrequent and similar for TCV and control recipients, as were adverse events (group 1, 61.2%; group 2, 64.0%; group 3, 68.6%) and serious adverse events (group 1, 2.0%; group 2, 8.0%; group 3, 5.9%). TCV generated robust immunity without interference with MCV-A vaccine. Conclusions: TCV can be safely co-administered at 15 months with MCV-A without interference. This novel study on the co-administration of TCV with MCV-A provides data to support large-scale uptake in sub-Saharan Africa.

(1) Background: Effective malaria case management relies on World Health Organization (WHO) recommended artemisinin-based combination therapies (ACTs), but partial resistance to artemisinin has emerged and is spreading, threatening malaria control and elimination efforts. The strategy of deploying multiple first-line therapies (MFT) may help mitigate this threat and extend the therapeutic life of current ACTs. (2) Methods: A district-wide pilot quasi-experimental study was conducted, deploying three different ACTs at the public health facility (PHF) level for uncomplicated malaria treatment from December 2019 to December 2020 in the health district (HD) of Kaya, Burkina Faso. Mixed methods, including household and health facility-based quantitative and qualitative surveys, were used to evaluate the pilot programme. (3) Results: A total of 2008 suspected malaria patients were surveyed at PHFs, of which 79.1% were tested by rapid diagnostic test (RDT) with 65.5% positivity rate. In total, 86.1% of the confirmed cases received the appropriate ACT according to the MFT strategy. The adherence level did not differ by study segment (p = 0.19). Overall, the compliance level of health workers (HWs) with MFT strategy was 72.7% (95% CI: 69.7–75.5). The odds of using PHF as the first source of care increased after the intervention (aOR = 1.6; 95% CI, 1.3–1.9), and the reported adherence to the 3-day treatment regimen was 82.1%; (95% CI: 79.6–84.3). Qualitative results showed a high acceptance of the MFT strategy with positive opinions from all stakeholders. (4) Conclusions: Implementing an MFT strategy is operationally feasible and acceptable by stakeholders in the health systems in Burkina Faso. This study provides evidence to support the simultaneous use of multiple first-line artemisinin combination therapies in malaria-endemic countries such as Burkina Faso.

Here, the authors identify signatures of miRNA expression differentiation associated with Plasmodium falciparum infection and parasitemia in a longitudinal pediatric cohort in Burkina Faso. In particular, expression of several miRNAs known to promote lymphocyte cell death is affected during infection.

Abstract Background The emergence and spread of multidrug-resistant gram-negative bacteria (MDR) has become a major public health concern worldwide. This resistance is caused by enzymes-mediated genes (i.e., extended spectrum beta-lactamases) that are common in certain Enterobacterioceae species. However, the distribution of these genes is poorly documented in Burkina Faso. This study aims to determine the prevalence and distribution of the resistant genes coding for broad spectrum beta-lactamases and quinolones in rural Burkina Faso. Methods Multiplex PCR assays were carried out to detect ESBL-encoding genes, including bla OXA, bla TEM, bla CTX-M, bla SHV. The assays also assessed the presence of quinolone resistance gene namely qnrA, qnrB and qnrS in the quinolone-resistance DEC and Salmonella strains. Results The Extended-Spectrum Beta-Lactamases (ESBL) resistance phenotype was reported in all the E. coli isolates (5/5). Cross-resistance phenotype to quinolones (CRQ) was shown by one Salmonella strain (1/9) and three E. coli (3/5). Cross-resistance phenotypes to fluoroquinolones (CRFQ) were harboured by one Salmonella (1/9) and carbapenemase phenotypes were detected in two E. coli strains (2/5). Whilst the bla OXA genes were detected in 100% (5/5) of E. coli isolates and in 33.33% (3/9) Salmonella isolates. One strain of E. coli (1/5) harbored the bla CTX−M gene and the qnrB gene simultaneously. Conclusions This study identified β-lactam (bla) and quinolone resistance (qnr) genes in multidrug-resistant E. coli and Salmonella spp. in rural Burkina Faso. Our finding which highlighted the enterobacteriaceae strains resistance to β-lactams and quinolones are of high interest for adequate management of antimicrobial resistant genes outbreak in Burkina Faso.

Abstract While significant advances have been made in understanding Plasmodium falciparum gametocyte biology and its relationship with malaria parasite transmission, the gametocyte sex ratio contribution to this process still remains a relevant research question. The present review discusses the biology of sex determination in P. falciparum, the underlying host and parasite factors, the sex specific susceptibility to drugs, the effect of sex ratio dynamics on malaria parasite transmission and the development of gametocyte sex specific diagnosis tools. Despite the inherent differences across several studies and approaches, the emerging picture highlights a potentially relevant contribution of the P. falciparum gametocyte sex ratio in the modulation of malaria parasite transmission. The increasing availability of molecular methods to measure gametocyte sex ratio will enable evaluation of important parameters, such as the impact of drug treatment on gametocyte sex ratio in vitro and in vivo as well as the changes of gametocyte sex ratios in natural infections, key steps towards elucidating how these parameters affect parasite infectiousness to the mosquito vectors.

Background: Individuals living in malaria-endemic regions develop immunity against severe malaria, but it is unclear whether immunity against pre-erythrocytic stages that blocks initiation of blood-stage infection after parasite inoculation develops following continuous natural exposure. Methods: We cleared schoolchildren living in an area (health district of Saponé, Burkina Faso) with highly endemic seasonal malaria of possible sub-patent infections and examined them weekly for incident infections by nested PCR. Plasma samples collected at enrolment were used to quantify antibodies to the pre-eryhrocytic-stage antigens circumsporozoite protein (CSP) and Liver stage antigen 1 (LSA-1). In vitro sporozoite gliding inhibition and hepatocyte invasion inhibition by naturally acquired antibodies were assessed using Plasmodium falciparum NF54 sporozoites. Associations between antibody responses, functional pre-erythrocytic immunity phenotypes and time to infection detected by 18S quantitative PCR were studied. Results: A total of 51 children were monitored. Anti-CSP antibody titres showed a positive association with sporozoite gliding motility inhibition (P

Background: Individuals living in malaria-endemic regions develop naturally acquired immunity against severe malarial disease, but it is unclear whether immunity that affects the establishment of infections develops following continuous natural exposure. Methods: We cleared schoolchildren in Burkina Faso of possible sub-patent infections and examined them weekly for incident infections by PCR. Plasma samples collected at enrolment were used to quantify antibodies to the pre-eryhrocytic-stage antigens circumsporozoite protein (CSP) and liver stage antigen. Sporozoite gliding inhibition by naturally acquired antibodies was assessed using Plasmodium falciparum NF54 sporozoites; hepatocyte invasion was assessed using the human HC-04 hepatoma cell line and NF54 sporozoites. The associations between these functional pre-erythrocytic immunity phenotypes and time to PCR-detected infection were studied. Results: A total of 51 children were monitored; the median time to first detection of infection by PCR or development of clinical symptoms was 28 days. Anti-CSP antibody titres showed a strong positive association with sporozoite gliding motility inhibition (P

Variation in biting frequency by Anopheles mosquitoes can explain some of the heterogeneity in malaria transmission in endemic areas. In this study in Burkina Faso, we assessed natural exposure to mosquitoes by matching the genotype of blood meals from 1066 mosquitoes with blood from residents of local households. We observed that the distribution of mosquito bites exceeded the Pareto rule (20/80) in two of the three surveys performed (20/85, 76, and 96) and, at its most pronounced, is estimated to have profound epidemiological consequences, inflating the basic reproduction number of malaria by 8-fold. The distribution of bites from sporozoite-positive mosquitoes followed a similar pattern, with a small number of individuals within households receiving multiple potentially infectious bites over the period of a few days. Together, our findings indicate that heterogeneity in mosquito exposure contributes considerably to heterogeneity in infection risk and suggest significant variation in malaria transmission potential.

During the season of high malaria transmission, most children are infected by Plasmodium, which targets red blood cells (RBCs), affecting haematological parameters. To describe these variations, we examined the haematological profiles of two groups of children living in a malaria-endemic area. A cross-sectional survey was conducted at the peak of the malaria transmission season in a rural area of Burkina Faso. After informed consent and clinical examination, blood samples were obtained from the participants for malaria diagnosis and a full blood count. Of the 414 children included in the analysis, 192 were not infected with Plasmodium, whereas 222 were asymptomatic carriers of Plasmodium infection. The mean age of the infected children was 41.8 months (range of 26.4-57.2) compared to 38.8 months (range of 22.4-55.2) for the control group (p = 0.06). The asymptomatic infected children tended to have a significantly lower mean haemoglobin level (10.8 g/dL vs. 10.4 g/dL; p < 0.001), mean lymphocyte count (4592/µL vs. 5141/µL; p = 0.004), mean platelet count (266 x 103/µL vs. 385 x 103/µL; p < 0.001) and mean RBC count (4.388 x 106/µL vs. 4.158 x 106/µL; p < 0.001) and a higher mean monocyte count (1403/µL vs. 1192/µL; p < 0.001) compared to the control group. Special attention should be applied when interpreting haematological parameters and evaluating immune responses in asymptomatic infected children living in malaria-endemic areas and enrolled in vaccine trials.

The association between P. falciparum eba-175, ama-1, and msp-3 polymorphism in the pathogenicity of malaria disease was investigated. We therefore compared the prevalence of different alleles between symptomatic and asymptomatic malarial children under five years of age living in Burkina Faso. Blood filter papers were collected during the 2008 malaria transmission season from 228 symptomatic and 199 asymptomatic children under five years of age. All patients were living in the rural area of Saponé at about 50 km from Ouagadougou, the capital city of Burkina Faso. P. falciparum parasite DNA was extracted using QIAGEN kits and the alleles diversity was assessed by a nested PCR. PCR products were then digested by restriction enzymes based on already described polymorphic regions of the eba-175, ama-1, and msp-3 genes. The individual alleles eba-175_FCR3 and msp-3_K1 frequencies were statistically higher (p0.05). The comparative analysis of P. falciparum genotypes indicated that the polymorphism in eba-175 and msp-3 genotypes varied between asymptomatic and symptomatic clinical groups and may contribute to the pathogenesis of malaria.

BACKGROUND:Children below six months are reported to be less susceptible to clinical malaria. Maternally derived antibodies and foetal haemoglobin are important putative protective factors. We examined antibodies to Plasmodium falciparum merozoite surface protein 3 (MSP3) and glutamate-rich protein (GLURP), in children in their first two years of life in Burkina Faso and their risk of malaria. METHODS:A cohort of 140 infants aged between four and six weeks was recruited in a stable transmission area of south-western Burkina Faso and monitored for 24 months by active and passive surveillance. Malaria infections were detected by examining blood smears using light microscopy. Enzyme-linked immunosorbent assay was used to quantify total Immunoglobulin G to Plasmodium falciparum antigens MSP3 and two regions of GLURP (R0 and R2) on blood samples collected at baseline, three, six, nine, 12, 18 and 24 months. Foetal haemoglobin and variant haemoglobin fractions were measured at the baseline visit using high pressure liquid chromatography. RESULTS:A total of 79.6% of children experienced one or more episodes of febrile malaria during monitoring. Antibody titres to MSP3 were prospectively associated with an increased risk of malaria while antibody responses to GLURP (R0 and R2) did not alter the risk. Antibody titres to MSP3 were higher among children in areas of high malaria risk. Foetal haemoglobin was associated with delayed first episode of febrile malaria and haemoglobin CC type was associated with reduced incidence of febrile malaria. CONCLUSIONS:We did not find any evidence of association between titres of antibodies to MSP3, GLURP-R0 or GLURP-R2 as measured by enzyme-linked immunosorbent assay and early protection against malaria, although anti-MSP3 antibody titres may reflect increased exposure to malaria and therefore greater risk. Foetal haemoglobin was associated with protection against febrile malaria despite the study limitations and its role is therefore worthy further investigation.

Ad35.CS.01 is a pre-erythrocytic malaria candidate vaccine. It is a codon optimized nucleotide sequence representing the P. falciparum circumsporozoite (CS) surface antigen inserted in a replication deficient Adenovirus 35 backbone. A Phase 1a trial has been conducted in the USA in naïve adults and showed that the vaccine was safe. The aim of this study is to assess the safety and immunogenicity of ascending dosages in sub Saharan Africa.A double blind, randomized, controlled, dose escalation, phase Ib trial was conducted in a rural area of Balonghin, the Saponé health district (Burkina Faso). Forty-eight healthy adults aged 18-45 years were randomized into 4 cohorts of 12 to receive three vaccine doses (day 0, 28 and 84) of 10(9), 10(10), 5X10(10), 10(11) vp of Ad35.CS.01 or normal saline by intra muscular injection. Subjects were monitored carefully during the 14 days following each vaccination for non serious adverse events. Severe and serious adverse events were collected throughout the participant study duration (12 months from the first vaccination). Humoral and cellular immune responses were measured on study days 0, 28, 56, 84, 112 and 140.Of the forty-eight subjects enrolled, forty-four (91.7%) received all three scheduled vaccine doses. Local reactions, all of mild severity, occurred in thirteen (27.1%) subjects. Severe (grade 3) laboratory abnormalities occurred in five (10.4%) subjects. One serious adverse event was reported and attributed to infection judged unrelated to vaccine. The vaccine induced both antibody titers and CD8 T cells producing IFNγ and TNFα with specificity to CS while eliciting modest neutralizing antibody responses against Ad35.Study vaccine Ad35.CS.01 at four different dose levels was well-tolerated and modestly immunogenic in this population. These results suggest that Ad35.CS.01 should be further investigated for preliminary efficacy in human challenge models and as part of heterologous prime-boost vaccination strategies.ClinicalTrials.gov NCT01018459 http://clinicaltrials.gov/ct2/show/NCT01018459.

Malariometric parameters are often primary endpoints of efficacy trials of malaria vaccine candidates. This study aims to describe the epidemiology of malaria prior to the conduct of a series of drug and vaccine trials in a rural area of Burkina Faso.Malaria incidence was prospectively evaluated over one year follow-up among two cohorts of children aged 0-5 years living in the Saponé health district. The parents of 1089 children comprising a passive case detection cohort were encouraged to seek care from the local health clinic at any time their child felt sick. Among this cohort, 555 children were randomly selected for inclusion in an active surveillance sub-cohort evaluated for clinical malaria during twice weekly home visits. Malaria prevalence was evaluated by cross-sectional survey during the low and high transmission seasons.Number of episodes per child ranged from 0 to 6 per year. Cumulative incidence was 67.4% in the passive and 86.2% in the active cohort and was highest among children 0-1 years. Clinical malaria prevalence was 9.8% in the low and 13.0% in the high season (p>0.05). Median days to first malaria episode ranged from 187 (95% CI 180-193) among children 0-1 years to 228 (95% CI 212, 242) among children 4-5 years. The alternative parasite thresholds for the malaria case definition that achieved optimal sensitivity and specificity (70-80%) were 3150 parasites/µl in the high and 1350 parasites/µl in the low season.Clinical malaria burden was highest among the youngest age group children, who may represent the most appropriate target population for malaria vaccine candidate development. The pyrogenic threshold of parasitaemia varied markedly by season, suggesting a value for alternative parasitaemia levels in the malaria case defintion. Regional epidemiology of malaria described, Sapone area field centers are positioned for future conduct of malaria vaccine trials.

Intermittent preventive treatment of malaria in children (IPTc) is a promising new approach to the control of malaria in areas of seasonal malaria transmission but it is not known if IPTc adds to the protection provided by an insecticide-treated net (ITN).An individually randomised, double-blind, placebo-controlled trial of seasonal IPTc was conducted in Burkina Faso in children aged 3 to 59 months who were provided with a long-lasting insecticide-treated bednet (LLIN). Three rounds of treatment with sulphadoxine pyrimethamine plus amodiaquine or placebos were given at monthly intervals during the malaria transmission season. Passive surveillance for malaria episodes was established, a cross-sectional survey was conducted at the end of the malaria transmission season, and use of ITNs was monitored during the intervention period. Incidence rates of malaria were compared using a Cox regression model and generalized linear models were fitted to examine the effect of IPTc on the prevalence of malaria infection, anaemia, and on anthropometric indicators. 3,052 children were screened and 3,014 were enrolled in the trial; 1,505 in the control arm and 1,509 in the intervention arm. Similar proportions of children in the two treatment arms were reported to sleep under an LLIN during the intervention period (93%). The incidence of malaria, defined as fever or history of fever with parasitaemia ≥ 5,000/µl, was 2.88 (95% confidence interval [CI] 2.70-3.06) per child during the intervention period in the control arm versus 0.87 (95% CI 0.78-0.97) in the intervention arm, a protective efficacy (PE) of 70% (95% CI 66%-74%) (p

Interventions that reduce exposure to malaria infection may lead to delayed malaria morbidity and mortality. We investigated whether intermittent preventive treatment of malaria in children (IPTc) was associated with an increase in the incidence of malaria after cessation of the intervention.An individually randomised, trial of IPTc, comparing three courses of sulphadoxine pyrimethamine (SP) plus amodiaquine (AQ) with placebos was implemented in children aged 3-59 months during the 2008 malaria transmission season in Burkina Faso. All children in the trial were given a long lasting insecticide treated net; 1509 children received SP+AQ and 1505 received placebos. Passive surveillance for malaria was maintained until the end of the subsequent malaria transmission season in 2009, and active surveillance for malaria infection, anaemia and malnutrition was conducted.On thousand, four hundred and sixteen children (93.8%) and 1399 children (93.0%) initially enrolled in the intervention and control arms of the trial respectively were followed during the 2009 malaria transmission season. During the period July 2009 to November 2009, incidence rates of clinical malaria were 3.84 (95%CI; 3.67-4.02) and 3.45 (95%CI; 3.29-3.62) episodes per child during the follow up period in children who had previously received IPT or placebos, indicating a small increase in risk for children in the former intervention arm (IRR = 1.12; 95%CI 1.04-1.20) (P = 0.003). Children who had received SP+AQ had a lower prevalence of malaria infection (adjusted PR: 0.88 95%CI: 0.79-0.98) (P = 0.04) but they had a higher parasite density (P = 0.001) if they were infected. There was no evidence that the risks of moderately severe anaemia (Hb

BACKGROUND:A Phase Ia trial in European volunteers of the candidate vaccine merozoite surface protein 3 (MSP3), a Plasmodium falciparum blood stage membrane, showed that it induces biologically active antibodies able to achieve parasite killing in vitro, while a phase Ib trial in semi-immune adult volunteers in Burkina Faso confirmed that the vaccine was safe. The aim of this study was to assess the safety and immunogenicity of this vaccine candidate in children aged 12-24 months living in malaria endemic area of Burkina Faso. METHODS:The study was a double-blind, randomized, controlled, dose escalation phase Ib trial, designed to assess the safety, reactogenicity and immunogenicity of three doses of either 15 or 30 microg of MSP3-LSP adsorbed on aluminum hydroxide in 45 children 12 to 24 months of age randomized into three equal groups. Each group received 3 vaccine doses (on days 0, 28 and 56) of either 15 microg of MSP3-LSP, 30 microg of MSP3-LSP or of the Engerix B hepatitis B vaccine. Children were visited at home daily for the 6 days following each vaccination to solicit symptoms which might be related to vaccination. Serious adverse events occurring during the study period (1 year) were recorded. Antibody responses to MSP3-LSP were measured on days 0, 28, 56 and 84. RESULTS:All 45 enrolled children received three MSP3 vaccine doses. No serious adverse events were reported. Most of the adverse events reported were mild to moderate in severity. The only reported local symptoms with grade 3 severity were swelling and induration, with an apparently dose related response. All grade 3 adverse events resolved without any sequelae. Both MSP3 doses regimens were able to elicit high levels of anti-MSP3 specific IgG1 and IgG3 antibodies in the volunteers with very little or no increase in IgG2, IgG4 and IgM classes: i.e. vaccination induced predominantly the isotypes involved in the monocyte-dependent mechanism of P. falciparum parasite-killing. CONCLUSION:Our results support the promise of MSP3-LSP as a malaria vaccine candidate, both in terms of tolerability and of immunogenicity. Further assessment of the efficacy of this vaccine is recommended. TRIAL REGISTRATION:ClinicalTrials.gov NCT00452088.