AIMS:Methanogens are methane-producing archaea that are present in the human gut. Yet, their adaptation to diverse human lifestyles remains poorly understood. Here, we report the isolation of Methanobrevibacter intestini G0370_i3 from the stool of a healthy adult from Southern Gabon, Africa, where inhabitants maintain traditional subsistence lifestyles with diets distinct from industrialized populations. MATERIALS AND METHODS:M. intestini was enriched from human stool, phenotypically characterized, and sequenced. RESULTS:G0370_i3 growth relied on the presence of H2 and CO2 and could also grow on formate, in contrast to reports for the type strain. The genome encoded pathways for amino acid biosynthesis, cofactor metabolism, and secondary metabolite production. We identified 23 mobile genetic elements and five defense systems, indicating horizontal gene transfer and antiviral defense. No prophage regions were detected.The genome also encoded uridine diphosphate (UDP)-sugar metabolism pathways, indicating capacity for energy storage and cell wall adaptability. Genes encoding adhesin-like proteins suggest capabilities for host interaction. Phenotypically, G0370_i3 is a coccobacillus, grows optimally at 37°C, and tolerates antibiotics, salt, and oxygen stress. CONCLUSIONS:These findings highlight the stress resilience and selective metabolic capabilities of M. intestini and underscore the importance of representing African populations in microbiome research.

The immune response to malaria vaccines is generally stronger in malaria-naive individuals than in those with lifelong exposure. The immunological basis for this is unclear. IgM, total IgG, and IgG subclass (IgG1, IgG2, IgG3, and IgG4) antibody responses against 21 pre-erythrocytic and erythrocytic Plasmodium falciparum proteins before and after controlled human malaria infection (CHMI) via the direct venous inoculation of 3,200 P. falciparum sporozoites (PfSPZ) were compared in three groups of volunteers: 1) malaria-naïve (n = 22); 2) malaria-naïve immunized with a PfSPZ chemoattenuated vaccine (PfSPZ-CVac) (n = 27); and 3) lifelong malaria-exposed individuals from Africa (n = 20), including those with normal hemoglobin (n = 11) or sickle cell trait (n = 9). Before and after CHMI, PfSPZ-CVac-immunized individuals exhibited higher levels of IgM and IgG to CSP and SSP-2/TRAP than the other two groups. Malaria-experienced Africans exhibited more intense and broader antibody responses to blood-stage (BS) antigens than naïve and vaccinated individuals, longer pre-patent periods (PPPs), and fewer symptoms. Among confirmed malaria cases, cytophilic IgG1 and IgG3 antibodies to BS antigens were positively associated with longer PPPs, whereas IgG2, IgG4, and IgM were not. IgG2 and IgG4 (noncytophilic) P. falciparum-specific antibodies were higher in the semi-immune group, including elevated anti-CSP IgG4 (regulatory) levels. The IgM response in African volunteers post-CHMI was stronger than that in malaria-naïve and vaccinated individuals and had the hallmark of a secondary memory response. Cytophilic immunoglobulins controlled parasitaemia better than noncytophilic immunoglobulins. However, elevation of the latter in lifelong malaria-exposed individuals could be associated with regulatory responses and hamper vaccine efficacy.

Continuous exposure to Plasmodium falciparum (Pf) has been associated with alterations in B cells. We investigated the effect of controlled human malaria infection (CHMI) on B cell phenotypes in individuals with different Pf immunity status: malaria-naïve, immunized with PfSPZ-CVac and semi-immune (lifelong-exposed) volunteers. Compared to naïve, semi-immune but not vaccinated individuals, had increased baseline frequencies of immature B cells (CD19+CD10+), active naive (IgD+CD27−CD21−) B cells, active atypical (IgD−CD27−CD21−) memory B cells (MBCs), active classical (IgD−CD27+CD21−) MBCs and CD1c+-B cells but lower frequencies of some IgG+-B cells. The frequencies of CD1c+ active atypical MBCs correlated positively with anti-Pf antibodies and negatively with circulating eotaxin levels, while the opposite was observed for IgG+ resting atypical MBCs. During early blood-stage infection (day 11 after CHMI), there was an expansion of resting classical (IgD−CD27+CD21+) MBCs in all three groups. Vaccination, compared to placebo, altered the effect of CHMI on B cells, showing a positive association with resting classical MBCs (β = 0.190, 95

BACKGROUND:Central nervous system (CNS) infections pose a significant public health challenge in resource-limited settings. Traditional culture-based and targeted molecular diagnostic methods have limitations in sensitivity and speed. This study retrospectively analyzed the data and cerebrospinal fluid (CSF) samples from our previous study to assess the diagnostic efficacy of untargeted 16S Oxford Nanopore Technology (ONT) sequencing compared to conventional CSF culture methods, with the goal of improving diagnostic accuracy, reducing time to treatment, and enhancing patient outcomes. METHODS:A total of 329 patients from 4 hospitals were enrolled in the study. CSF samples were collected and processed for both CSF culture and 16S ONT sequencing. DNA were extracted from CSF and amplified for 16S rRNA sequencing using the MinION platform. Descriptive analyses were conducted to assess pathogen detection rates and the potential impact of sequencing on antimicrobial stewardship. RESULTS:Of the 329 samples, 40 (12%) were positive for bacterial or fungal pathogens. 16S ONT detected pathogens in 28 samples (9%), while CSF culture identified pathogens in 23 samples (7%). 16S ONT sequencing identified 17 pathogens not detected by CSF culture, including Streptococcus suis and Acinetobacter baumannii. Based on 16S ONT findings, 61% of patients were found to have received inappropriate empirical antibiotic therapy and could have benefited from improved antimicrobial management, including de-escalation in 11, escalation in 5, and adjustments in 2 cases. CONCLUSIONS:16S ONT sequencing showed higher sensitivity and diagnostic yield than CSF culture, providing clinical insights for managing CNS infections through targeted antibiotic use and enhanced antimicrobial stewardship in resource-limited settings.

OBJECTIVES:This study investigated the impact of causative pathogens on the outcomes of central nervous system (CNS) infections and assessed clinical parameters to identify patients at risk for unfavourable outcomes. METHODS:Patients with suspected CNS infections underwent blood and cerebrospinal fluid (CSF) culture and advanced molecular testing, including real-time PCR assays for bacterial and viral pathogens. Patients were classified into clinical categories and their outcomes assessed using the Glasgow Outcome Scale. RESULTS:Pathogens were identified in 24% (80/330) of patients, with Mycobacterium tuberculosis, Klebsiella pneumoniae, and Acinetobacter baumannii being the most common. Mortality was 10%, with fungal meningitis and dual infections having the highest rates. Unfavourable outcomes were observed in 57% of patients. The most common pathogens associated with unfavourable outcomes were M. tuberculosis followed by K. pneumoniae, A. baumannii, and HSV-1. Multivariate analysis identified community-onset infection as a protective factor, while a longer duration of illness before admission (≥5 days) and altered mental status on admission were significant predictors for unfavourable outcomes. Furthermore, the timely administration of appropriate empirical therapy was significantly associated with a reduced risk of mortality. CONCLUSIONS:CNS infections in northern Vietnam have diverse causes and overlapping clinical features, complicating diagnosis and management.

Plasmodium falciparum strains lacking P. falciparum histidine-rich protein 2 (PfHRP2) and PfHRP3 threaten malaria rapid test reliability. We show that pfhrp2/pfhrp3-deleted parasites circulated in Ethiopia as early as 2009, before widespread PfHRP2-based rapid test use, and had high pfhrp3 deletion prevalence. Monitoring of pfhrp2 and of pfhrp3 deletions is needed.

Flaviviruses such as dengue virus (DENV), Zika virus (ZIKV), and Japanese encephalitis virus (JEV) pose a major health burden in Vietnam, where overlapping clinical features and serological cross-reactivity complicate accurate diagnosis and outbreak control. This study aimed to investigate circulating DENV serotypes and assess serological cross-reactivity with other flaviviruses during the 2016 dengue outbreak in central Vietnam. Aretrospective study was conducted on 146 hospitalized dengue patients during the 2016 outbreak in Binh Dinh province. Laboratory diagnosis included NS1antigen testing, ELISA (IgM/IgG), and real-time RT-PCR for DENV serotyping. IgM and IgG cross-reactivity with five flaviviruses, including DENV, ZIKV, JEV, West Nile virus (WNV), and tick-borne encephalitis virus (TBEV), and onealphavirus, chikungunya virus (CHIKV), was evaluated using ELISA. DENV-1 positive samples were further analysed by sequencing the capsid-premembrane (CprM) gene. DENV-1 was the predominant serotype (86%), with all sequenced strains clustering within genotype I. Secondary infections were more frequent (64%) than primary infections (36%) and were associated with a significantly higher median age (p = 0.003) and elevated hs-CRP levels (p = 0.029). Strong IgG cross-reactivity was observed among flaviviruses, particularly DENV, JEV, WNV, and TBEV (r > 0.85), while ZIKV and CHIKV showed low seropositivity. Incontrast, IgM responses demonstrated greater virus specificity. Ten PCR-negative cases showed broad serological reactivity, suggesting possible misdiagnosis or late-stage infection. Our findings reveal that DENV-1 genotype I was the predominant serotype during the 2016 outbreak in central Vietnam. Extensive IgG cross-reactivity among flaviviruses hinders serological diagnosis, highlighting the need for integrated molecular surveillance to ensure accurate outbreak response.

Malaria remains a preventable and treatable disease; however, recent efforts to reduce mortality have plateaued. Although artemisinin-based combination therapy demonstrates high efficacy in controlled clinical settings, its real-world effectiveness is often compromised by suboptimal patient adherence. Specifically, the artemether–lumefantrine regimen, administered twice daily over 3 days, has been associated with reduced compliance due to its complexity. Simplified therapeutic regimens that enhance adherence could, therefore, play a critical role in reinvigorating progress toward malaria elimination. Over the past decade, substantial progress has been made in the discovery and development of new chemical entities for malaria treatment, although the most advanced candidate still requires a 3-day dosing regimen. Treatment shortening most likely requires multiple drug combinations. Multi-drug regimens, such as artemether–lumefantrine–amodiaquine appear to be well tolerated, but these are under development to address emerging resistance to lumefantrine and will be unlikely to improve compliance. Sulfadoxine–pyrimethamine was originally developed as a single-dose curative treatment for malaria, and although use was curtailed early due to rapid selection for resistance, it continues to be deployed as a single therapy or in combination with other medicines, in treatment and in prevention. Combining with artemisinin-based combinations would be an option for potential treatment shortening. Of the registered antimalarial treatments, only a few of the artemisinin-based combinations are suitable. Mefloquine is excluded for tolerability concerns, amodiaquine because of its use in seasonal malaria chemoprevention, and lumefantrine and piperaquine due to concerns of emerging resistance. Pyronaridine–artesunate emerges as a promising candidate for association with sulfadoxine–pyrimethamine. A four-drug, single-dose antimalarial regimen would transform compliance, and play a major role in disease elimination. However, to ensure its success it will be important to assess the safety and tolerability of the novel association and understand its efficacy in regions with evolving resistance to sulfadoxine–pyrimethamine. Clinical studies need to assess the risk for selection of strains with novel resistance mechanisms against artesunate or pyronaridine. Importantly, a comprehensive clinical evaluation will generate valuable real-world insights into community acceptance and operational feasibility. This information will be an important foundation for future design of single dose malaria therapies involving new chemical entities.

Plants used in traditional medicine represent an important source of new compounds. Hallea ledermannii (H. ledermannii) (Krause) Verdc. (Rubiaceae), Gossypium barbadense (G. barbadense) (Malvaceae), Pycnanthus angolensis (P. angolensis) (Myristicaceae), Drypetes gossweileri (D. gossweileri) S. Moore (Euphorbiaceae) and Scyphocephalium ochocoa (S. ochocoa) Warb (Myristicaceae) are five plants widely used in traditional Gabonese medicine as antimicrobials. However, little is known about the active compounds associated with their biological activities. Based on botanical studies and the claims of traditional healers regarding the antimicrobial effects of these plants, a study to evaluate the antimicrobial activity and phyto-chemical profile of aqueous extracts of three plants (bark of H. ledermannii, G. barbadense root bark and P. angolensis bark) and methanolic extracts of two plants (S. ochocoa stem bark and D. gossweileri root bark). Under the guidance of LC-MS detection, identified twenty seven (27) potentially active compounds. Eight (8) of these belong to the quinovic acid-type triterpenoid sap-onins identified in the aqueous extract of H. ledermannii, eleven (11) are dibenzofurans, chroman and stigmasterol detected in the methanolic extract of S. ochocoa and eight (8) compounds in the methanolic extract of D. gossweileri are friedelin, drypemolundein B and gossweilone, to name but a few. In parallel, the five extracts were tested on reference bacterial strains: Staphylococcus aureus ATCC 25923, Pseudomonas aeruginosa ATCC 278533, Salmonella thyphi ATCC 13311, Klebsiella pneumoniae ATCC 700603, Shigella flexneri ATCC 24570. Three of these extracts (aqueous extract of H. ledermannii and two methanolic extracts of S. ochocoa stem bark and D. gossweileri root bark) showed moderate activity against Staphylococcus aureus ATCC 25923, with inhibition zones of 12.3 ± 0.5 mm, 10.1 ± 0.5 mm and 7,6 ± 0 mm respectively. In addition, we assessed the toxicity of the three extracts that showed antimicrobial activity using an invertebrate model, Galleria mellonella (GM). We found that the LD varied according to the concentration of the plant material. The aqueous extract of H. ledermannii and the methanolic extracts of S. ochocoa and D. gossweileri were not toxic to G. mellonella. The LD50s (mg/mL) obtained were 93.2 mg/mL [717.2 g/kg body weight (bw)] and 100 mg/mL (762.3 g/kg bw), 95.4 mg/mL [721.1 g/kg body weight (bw)].

The prevailing focus of lifespan health research has predominantly centered on "healthy aging". This oversight may hinder the understanding of health across the lifespan, as disorders in earlier stages can substantially impact overall health and longevity. Aging, conceptually, begins at gestation. The trajectory of an individual's health is influenced from the earliest stages of life, where adverse conditions can set a foundation for lifelong health challenges. For example, suboptimal conditions during gestation leading to premature birth can predispose individuals to various health issues later in life. Additionally, precocious puberty defined as the onset of sexual maturity before eight years of age or early menopause-occurring before 50 years of age requires medical intervention and is indicative of atypical aging processes. To address these critical gaps in lifespan health research, the expansion of medical lexicons and research categorizations is advocated to include "healthy gestation," "healthy development," and "healthy reproduction" alongside "healthy aging." This broader terminology will enable a more comprehensive investigation of disorders at all life stages. An integrative approach underscores the interconnectedness of all life stages and the continuous nature of aging, advocating for a seamless continuum in health research and interventions from gestation through late adulthood.

Background Central nervous system (CNS) infections pose a significant public health challenge in resource-limited settings. Traditional culture-based and targeted molecular diagnostic methods have limitations in sensitivity and speed. This study retrospectively analyzed the data and cerebrospinal fluid (CSF) samples from our previous study to assess the diagnostic efficacy of untargeted 16S Oxford Nanopore Technology (ONT) sequencing compared to conventional CSF culture methods, with the goal of improving diagnostic accuracy, reducing time to treatment, and enhancing patient outcomes. Methods A total of 329 patients from 4 hospitals were enrolled in the study. CSF samples were collected and processed for both CSF culture and 16S ONT sequencing. DNA were extracted from CSF and amplified for 16S rRNA sequencing using the MinION platform. Descriptive analyses were conducted to assess pathogen detection rates and the potential impact of sequencing on antimicrobial stewardship. Results Of the 329 samples, 40 (12%) were positive for bacterial or fungal pathogens. 16S ONT detected pathogens in 28 samples (9%), while CSF culture identified pathogens in 23 samples (7%). 16S ONT sequencing identified 17 pathogens not detected by CSF culture, including Streptococcus suis and Acinetobacter baumannii. Based on 16S ONT findings, 61% of patients were found to have received inappropriate empirical antibiotic therapy and could have benefited from improved antimicrobial management, including de-escalation in 11, escalation in 5, and adjustments in 2 cases. Conclusions 16S ONT sequencing showed higher sensitivity and diagnostic yield than CSF culture, providing clinical insights for managing CNS infections through targeted antibiotic use and enhanced antimicrobial stewardship in resource-limited settings.

OBJECTIVES:Vaccination against SARS-CoV-2 induces antibodies that reduce the risk of severe disease. Because IgG subclasses differ in their ability to activate complement, to bind Fc receptors and neutralize viruses, it is crucial to understand how IgG subclass responses differ between vaccine platforms. DESIGN:IgG1, IgG2, IgG3, and IgG4 binding antibodies against SARS-CoV-2 trimeric spike protein, receptor-binding domain, and S1/S2 subunits responses were quantified using a multiplex immunoassay, after a booster dose of either BNT162b2 (Pfizer/BioNTech) or mRNA-1273 (Moderna) in a healthy cohort (n = 165) who had received two previous vaccine doses. RESULTS:Boosting increased all subclass IgG levels, except for S1-specific IgG1 and S2-specific IgG2. However, IgG2 and IgG4 levels were significantly higher in BNT162b2 than in mRNA-1273 vaccinees (P = 0.0313 [IgG2 S] and P = 0.0106 [IgG4 RBD], P = 0.0070 [IgG4 S1]). Individuals who had previously received a non-mRNA vaccination showed no significant increase in IgG2 (P = 0.4909 [S]) and IgG4 (P = 0.0607 [S]) post-boost. CONCLUSIONS:Vaccine-specific differences post-booster vaccination were identified and may drive the class switch between IgG2 and IgG4 responses. Given their different roles, these subtle differences may ultimately also affect long-term immunity and protection.

Abstract To develop a Plasmodium falciparum (Pf) vaccine that precludes replication inside the host for improved vaccine safety, we tested chemo-attenuation (CVac) of sporozoites (SPZ) with atovaquone–proguanil (AP). In mice, P. berghei sporozoites administered with AP invaded hepatocytes, arrested early, and induced robust protection, which correlated with parasite-specific effector-memory CD8+ T cell responses. In a clinical trial of PfSPZ-CVac (AP), in which three doses of 5.12 × 104 or 1.5 × 105 PfSPZ were administered by direct venous inoculation combined with oral single-dose AP (1000/400 mg), blood stage infections were fully prevented during immunisation. 2/8 and 2/10 of vaccinees, respectively, were protected when challenged with 3.2 × 103 PfSPZ 10 weeks later, inferior to PfSPZ-CVac (chloroquine/CQ) that allows in-host replication. Comparative analysis of responses to 228 Pf proteins revealed that protection with PfSPZ-CVac (CQ) was associated with antibodies to two liver-stage antigens (LISP2, LSA1) and a multi-stage antigen (PfMSP5), but not to the major surface protein PfCSP. The complete arrest of high numbers of Pf sporozoites by single-dose AP should allow a significant dose-frequency reduction of the current daily AP malaria chemoprophylaxis regimen.

Objectives:This study aimed to investigate genotypic characteristics and drug resistance profiles of Mycobacterium tuberculosis complex (Mtbc) strains isolated from patients with suspected tuberculosis (TB) in Gabon. Methods:We performed whole genome sequencing of 430 Mtbc strains cultured between 2012 and 2022. Phylogenetic strain classification, genomic resistance prediction, and cluster analysis were also performed. Results:Strains from four major Mtbc lineages, L4 (n = 372; 65%), L5 (n = 46; 11%), L2 (n = 6; 1,4%), and L6 (n = 3; 0,7%), were observed. Interestingly, more than 10% of the strains were represented by Mycobacterium africanum strains (L5 and L6), with L4 strains being dominant. The subclassification of L4 strains showed that L4.6.2.2 (Cameroon, n = 116) was the most prevalent. The proportion of resistance (any resistance to first-line TB drugs) was 30%, with 12% (n = 52) being at least multidrug-resistant (MDR) and six showing additional fluoroquinolone resistance. The overall cluster rate was 64% in non-MDR and MDR Mtbc strains. Conclusion:Although most Mtbc infections are caused by L4 M. tuberculosis strains, M. africanum strains also contribute to TB epidemiology in Gabon. The MDR epidemic is mainly driven by one dominant L4 Haarlem outbreak clone that has been spreading for approximately a century in the country.

BACKGROUND:The pathogenesis of dengue is attributed to a complex interaction between the dengue virus (DENV) and the host immune system. The aim of this study is to investigate the clinical, virological, and Interleukin-10 (IL-10) profiles of dengue patients in Vietnam from two consecutive outbreaks in 2021 and 2022. METHODS:A total of n=306 dengue patients were examined, who were clinically stratified according to dengue without warning signs (DF; n=178), dengue with warning signs (DWS; n=115) and severe dengue (SD; n=13). Patients were screened for dengue, Zika and chikungunya viruses. DENV were subjected to serotype specific real-time RT-PCR. Interleukin-10 (IL-10) levels were measured by ELISA, and IL-10 promoter variants (-1082G/A; -819C/T; -592C/A) were genotyped by direct Sanger sequencing to determine a possible association with susceptibility to dengue and disease severity. RESULTS:No chikungunya or Zika viruses were detected. Patients were infected by one of the three different DENV serotypes (DENV-1, -2, -4). Plasma IL-10 levels were significantly elevated in patients (DF vs. DWS, p=0.004; DF vs. SD, p=0.001; DWS vs. SD, p=0.015). While the IL-10 allele -819C contributed to an increased risk of dengue (OR = 1.5, 95% CI = 1.1-2.0, p=0.04), genotype -1082GA showed a protective role against the disease (OR = 0.45, 95% CI = 0.27-0.72, p=0.009), and allele -1082G showed a protective role against DWS (OR = 0.44, 95% CI = 0.22-0.81, p=0.049). Also, the IL-10 GTA (-1082G/-819T/-592A) haplotype was observed to confer protection (OR = 0.31, 95% CI = 0.14-0.67, p< 0.003). CONCLUSION:While DENV-1 and DENV-2 were the predominant serotypes in circulation, plasma IL-10 levels and IL-10 promoter variants were also significantly associated with dengue and its severity.

Dengue pathogenesis involves immune-driven inflammation that contributes to severe disease progression. This study assessed a machine learning model to identify a minimal, yet highly predictive biomarker set, aiming to support clinical decision-making and patient triage. A total of 48 inflammatory mediators were quantified from plasma samples collected at admission from confirmed dengue patients, classified as either dengue without warning signs (DF) or dengue with warning signs/severe dengue (DWS/SD). A random forest approach was applied to identify the most predictive biomarkers associated with disease severity requiring hospitalization, based on admission-time variables. Among the 48 immune mediators, 43 were differentially expressed in dengue patients versus healthy controls, and 26 showed significant differences between DF and DWS/SD cases. Lymphocyte counts negatively correlated with IL-1RA, while liver enzymes showed positive correlations with HGF and SCGF-beta; platelet counts also negatively correlated with these markers. Key severity-associated markers included HGF, TNF-beta, MIP-1-beta, and SCGF-beta. A model incorporating these markers and fever duration achieved nearly 80% accuracy in distinguishing DWS/SD from DF cases, independent of clinical examination. The findings suggest that targeted cytokine profiling may guide early hospitalization decisions and ease healthcare burdens in dengue-endemic regions.

The intestinal microbiota is thought to modulate immune responsiveness to vaccines. Human studies on this topic, however, have yielded inconsistent results. We hypothesized that the microbiome would influence innate immune responses, and thus vaccine reactogenicity, more directly than vaccine immunogenicity. To test this, we established the uHEAT (Microbial-Human Ecology And Temperature) study, which longitudinally profiled the fecal microbiota, oral body temperature and serum antibody responses of 171 healthy adults (18-40 years old) before and after vaccination for SARS-CoV-2. Increased temperature after vaccination (delta-T) was associated with habitual diet and with baseline metabolic and immune markers. The microbiomes of delta-T-high (delta-Thi) participants were characterized by high expression of flagellin and an overabundance of the flagellated bacterium Waltera. Fecal samples from delta-Thi participants induced more inflammation in human cells and stronger post-vaccine temperature responses in mice compared to delta-Tlo samples, suggesting a causal role for the microbiome. Moreover, Waltera flagellin replicated the inflammatory phenotypes in vitro and was modulable via a dietary additive. Overall, these data suggest that flagellin from the gut microbiome stimulates innate immunity and vaccine reactogenicity, and that this axis can be manipulated via diet. These findings have implications for improving human vaccine tolerance and immunogenicity. ### Competing Interest Statement The authors have declared no competing interest.

Abstract Background Insecticides are a crucial component of vector control. However, resistance constitute a threat on their efficacy and the gains obtained over the years through malaria vector control. In Gabon, little data on phenotypic insecticide resistance in Anopheles vectors are published, compromising the rational implementation of resistance management strategies. We assessed the susceptibility to pyrethroids, carbamates and organophosphates of Anopheles gambiae sensu lato (s.l.) and discuss the mechanisms involved in the pyrethroid resistance-phenotype. Methods A. gambiae s.l. larvae were collected from breeding sites in Lambaréné. Emerging adults were used in WHO tube assays at an insecticide concentration that defines resistance (diagnostic concentration). Subsequently, deltamethrin and permethrin were used at 5x and 10x diagnostic concentrations and after preexposure with the cytochrome p450 (and glutathione S-transferase) inhibitor piperonyl butoxide (PBO). A subset of mosquitoes was typed by molecular methods and screened using Taqman assays for mutations conferring target site resistance at the Voltage-gated sodium channel 1014 (Vgsc-1014) locus and the acetylcholinesterase (Ace-1) gene. Results All mosquitoes were A. gambiae sensu stricto (s.s.) and resistant to permethrin, deltamethrin and alphacypermethrin (mortality less than 98%). However, mosquitoes were susceptible to malathion but resistant to bendiocarb. The level of resistance was high for permethrin and at least moderate for deltamethrin. Pre-exposure to PBO significantly increased the mortality of resistant mosquitoes (P < 0.0001). They became fully susceptible to deltamethrin and permethrin-induced mortality increased 4-fold. The G119S Ace-1 resistance allele, which confers resistance to both organophosphates and carbamates, was not present. All sampled mosquitoes were either homozygous for the Vgsc-L1014F or heterozygous for Vgsc-L1014F/L1014S, a marker for resistance to pyrethroids and organochlorides. Conclusion These findings demonstrate a role of cytochrome P450 monooxygenases in the pyrethroid-resistance of A. gambiae s.s. from Lambaréné. Combining PBO with pyrethroids, as done in second generation bednets, may be used to revert resistance. In addition, malathion could also be used in combination with pyrethroids-based methods for resistance management.

Background More than 20 million people are infected with L. loa, and around 40 million live in high or intermediate-risk areas in West- and Central Africa. Although loiasis is associated with significant morbidity and excess mortality, little is known about the perception of loiasis by affected communities. This study assessed the knowledge, attitudes, and practices in the rural population of Sindara, Gabon, a region characterized by high loiasis prevalence. Methods A community-based cross-sectional survey was conducted in Gabon between January and June 2022. During systematic door-to-door visits, randomly selected inhabitants were invited to participate in this questionnaire based survey. Venous blood was collected at midday from all participants for microscopic detection of filarial infection and clinical signs of loiasis were assessed. Results A total of 150 participants were recruited, of which 66% were infected by L. loa. While almost everyone had some knowledge about L. loa, 72% of the participants understood that L. loa is a parasitic worm. The transmission of L. loa via the deer fly was known to only 21% of participants. The most frequently mentioned clinical symptoms attributed to loiasis were itching (84%), eye worm migration (59%), and conjunctivitis-like symptoms (53%). Participants who experienced migratory loiasis had better knowledge of loiasis and considered it as more serious. Traditional and herbal medicine was reported most often as an available treatment option (72%). While the formal healthcare sector was mentioned as the preferred treatment provider, 60% of the reported infections were treated by traditional medical practitioners. Conclusion Loiasis is in general well known by this community residing in a region of high L. loa transmission. Important gaps in knowledge were discovered foremost regarding the mode of transmission. The available healthcare system does not seem to provide adequate management for loiasis.