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Background Schistosoma antigen detection in urine is a valuable diagnostic approach for schistosomiasis control programmes because of the higher sensitivity compared to parasitological methods and preferred sampling of urine over stool. Highly accurate diagnostics are important in low Schistosoma transmission areas. Pregnant women and young children could particularly benefit from antigen testing as praziquantel (PZQ) can be given to only confirmed Schistosoma cases. This prevents the unborn baby from unnecessary exposure to PZQ. We present here the protocol of a diagnostic study that forms part of the freeBILy project. The aim is to evaluate the accuracy of circulating anodic antigen (CAA) detection for diagnosis of Schistosoma haematobium infections in pregnant women and to validate CAA as an endpoint measure for anti-Schistosoma drug efficacy. The study will also investigate Schistosoma infections in infants. Methods A set of three interlinked prospective, observational studies is conducted in Gabon. The upconverting phosphor lateral flow (UCP-LF) CAA test is the index diagnostic test that will be evaluated. The core trial, sub-study A, comprehensively evaluates the accuracy of the UCP-LF CAA urine test against a set of other Schistosoma diagnostics in a cross-sectional trial design. Women positive for S. haematobium will proceed with sub-study B and will be randomised to receive PZQ treatment immediately or after delivery followed by weekly sample collection. This approach includes comparative monitoring of CAA levels following PZQ intake and will also contribute further data for safety of PZQ administration during pregnancy. Sub-study C is a longitudinal study to determine the incidence of S. haematobium infection as well as the age for first infection in life-time. Discussion The freeBILy trial in Gabon will generate a comprehensive set of data on the accuracy of the UCP-LF CAA test for the detection of S. haematobium infection in pregnant women and newborn babies and for the use of CAA as a marker to determine PZQ efficacy. Furthermore, incidence of Schistosoma infection in infants will be reported. Using the ultrasensitive diagnostics, this information will be highly relevant for Schistosoma prevalence monitoring by national control programs as well as for the development of medicaments and vaccines. Trial registration The registration number of this study is NCT03779347 ( clinicaltrials.gov , date of registration: 19 December 2018).

Two hookworm vaccine candidates, Na-GST-1 and Na-APR-1, formulated with Glucopyranosyl Lipid A (GLA-AF) adjuvant, have been shown to be safe, well tolerated, and to induce antibody responses in a Phase 1 clinical trial (Clinicaltrials.gov NCT02126462) conducted in Gabon. Here, we characterized T cell responses in 24 Gabonese volunteers randomized to get vaccinated three times with Na-GST-1 and Na-APR-1 at doses of 30μg (n = 8) or 100μg (n = 10) and as control Hepatitis B (n = 6). Blood was collected pre- and post-vaccination on days 0, 28, and 180 as well as 2-weeks after each vaccine dose on days 14, 42, and 194 for PBMCs isolation. PBMCs were stimulated with recombinant Na-GST-1 or Na-APR-1, before (days 0, 28 and 180) and two weeks after (days 14, 42 and 194) each vaccination and used to characterize T cell responses by flow and mass cytometry. A significant increase in Na-GST-1 -specific CD4+ T cells producing IL-2 and TNF, correlated with specific IgG antibody levels, after the third vaccination (day 194) was observed. In contrast, no increase in Na-APR-1 specific T cell responses were induced by the vaccine. Mass cytometry revealed that, Na-GST-1 cytokine producing CD4+ T cells were CD161+ memory cells expressing CTLA-4 and CD40-L. Blocking CTLA-4 enhanced the cytokine response to Na-GST-1. In Gabonese volunteers, hookworm vaccine candidate, Na-GST-1, induces detectable CD4+ T cell responses that correlate with specific antibody levels. As these CD4+ T cells express CTLA-4, and blocking this inhibitory molecules resulted in enhanced cytokine production, the question arises whether this pathway can be targeted to enhance vaccine immunogenicity., Author summary Two hookworm vaccine candidate (Na-GST-1 and Na-APR-1) have been tested in Gabonese and found to be safe and to induce antibody response. We aimed to study the cellular immune responses among vaccinated and unvaccinated volunteers. We found that Na-GST-1 induced CD4+ T cell responses (IL-2, TNF) among the vaccinated volunteers that received the high vaccine dose (100 ug). Furthermore Na-GST-1 specific memory T cells were found to express the inhibitory molecule CTLA-4. These responses was not observed in those who received the low dose of the Na-GST-1 vaccine, or those who received Na-APR-1 or HBV. By blocking CTLA-4, we observed an increase in TNF production. Our data suggest that an intervention involving blockage of the CTLA-4 molecule in the vaccinated could be beneficial in endemic settings where vaccine responses have been shown to be lower compared to non-endemic settings.

Background Hookworms cause substantial morbidity in children and women of reproductive age. The control strategy of mass drug administration is suboptimal, hence the need for a vaccine. Necator americanus aspartic protease-1 (Na-APR-1) and N americanus glutathione S-transferase-1 (Na-GST-1) are involved in the digestion and detoxification of haemoglobin in the hookworm digestive tract. In animal models, vaccination against these antigens resulted in protection from challenge infection. Both vaccine candidates were shown to be safe and well tolerated when administered separately to healthy adults. We assessed the safety and immunogenicity of co-administered Na-GST-1 and Na-APR-1 (M74) vaccines in healthy Gabonese adults.Methods This randomised, controlled, double-blind, phase 1, dose-escalation trial was done at the Centre de Recherches Medicales de Lambarene, in a region of Gabon where N americanus and other helminths are prevalent. Healthy adults aged 18-50 years and living in Lambarene or the surrounding areas were recruited to the study. Participants were enrolled consecutively into two dose cohorts (30 mu g or 100 mu g of the experimental vaccines) and randomly assigned in blocks (block size four) to receive three doses of either co-administered Na-GST-1 plus Na-APR-1 (M74; 30 mu g or 100 mu g of each), adjuvanted with Alhydrogel (aluminium hydroxide gel suspension) together with an aqueous formulation of glucopyranosyl lipid A, or hepatitis B vaccine plus saline (control group). Vaccines were administered intramuscularly on days 0, 28, and 180. The primary endpoint was safety, with immunogenicity a secondary endpoint. The intention-to-treat population was used for safety analyses, whereas for immunogenicity analyses, the per-protocol population was used (participants who received all scheduled vaccinations). Control vaccine recipients for both dose cohorts were combined for the analyses. The trial is registered with ClinicalTrials.gov, NCT02126462.Findings Between Oct 27, 2014, and Jan 31, 2015, 56 individuals were screened for eligibility, of whom 32 were enrolled and randomly assigned to one of the three study groups (12 each in the 30 mu g and 100 mu g experimental vaccine groups and eight in the control group). Both study vaccines were well tolerated in both dose groups. The most common adverse events were mild-to-moderate injection-site pain, headache, myalgia, and nausea. No severe or serious adverse events related to the vaccines were recorded. 52 unsolicited vaccine-related adverse events occurred during the study, but there was no difference in frequency between vaccine groups. IgG antibodies were induced to each of the vaccine antigens, with mean IgG levels increasing after each vaccination. Vaccination with 100 mu g of each vaccine antigen consistently induced IgG seroconversion (IgG levels above the reactivity threshold). Peak IgG responses were observed 2 weeks after the third vaccine dose for both antigens, with all participants who received the 100 mu g doses seroconverting at that timepoint. IgG levels steadily declined until the final study visit 6 months after the third vaccination, although they remained significantly higher than baseline in the 100 mu g dose group.Interpretation Vaccination with recombinant Na-GST-1 and Na-APR-1 (M74) in healthy adults living in N americanusendemic areas of Gabon was safe and induced IgG to each antigen. To our knowledge, this study is the first to report results of Na-APR-1 (M74) co-administered with Alhydrogel in participants from an N americanus-endemic area. Further clinical development of these vaccines should involve efficacy studies. Funding European Union Seventh Framework Programme. Copyright (C) 2020 Elsevier Ltd. All rights reserved.

GMZ2 is a recombinant protein malaria vaccine, comprising two blood-stage antigens of Plasmodium falciparum, glutamate-rich protein and merozoite surface protein 3. We assessed efficacy of GMZ2 in children in Burkina Faso, Gabon, Ghana and Uganda. : Children 12-60months old were randomized to receive three injections of either 100μg GMZ2 adjuvanted with aluminum hydroxide or a control vaccine (rabies) four weeks apart and were followed up for six months to measure the incidence of malaria defined as fever or history of fever and a parasite density ⩾5000/μL. : A cohort of 1849 children were randomized, 1735 received three doses of vaccine (868 GMZ2, 867 control-vaccine). There were 641 malaria episodes in the GMZ2/Alum group and 720 in the control group. In the ATP analysis, vaccine efficacy (VE), adjusted for age and site was 14% (95% confidence interval [CI]: 3.6%, 23%, p-value=0.009). In the ITT analysis, age-adjusted VE was 11.3% (95% CI 2.5%, 19%, p-value=0.013). VE was higher in older children. In GMZ2-vaccinated children, the incidence of malaria decreased with increasing vaccine-induced anti-GMZ2 IgG concentration. There were 32 cases of severe malaria (18 in the rabies vaccine group and 14 in the GMZ2 group), VE 27% (95% CI -44%, 63%). : GMZ2 is the first blood-stage malaria vaccine to be evaluated in a large multicenter trial. GMZ2 was well tolerated and immunogenic, and reduced the incidence of malaria, but efficacy would need to be substantially improved, using a more immunogenic formulation, for the vaccine to have a public health role.

An estimated 40 million women of childbearing age suffer from schistosomiasis. Animal models indicate a deleterious effect of maternal schistosomiasis on pregnancy outcomes. To date there is a lack of epidemiological evidence evaluating schistosomiasis-related morbidity in pregnancy. This study was designed to describe the impact of urogenital schistosomiasis on pregnancy outcomes in a highly endemic region of central Africa. Pregnant women attending antenatal clinics in Fougamou and Lambaréné, Gabon, were consecutively screened for the presence of Schistosoma haematobium eggs in diurnal urine samples. Maternal and newborn characteristics assessed at delivery were compared between infected and uninfected mothers. The impact of maternal schistosomiasis on low birth weight and preterm delivery was assessed using logistic regression analysis. Urogenital schistosomiasis was diagnosed in 103 (9%) of 1115 pregnant women. Maternal age was inversely associated with the prevalence of urogenital schistosomiasis, with a higher burden amongst nulliparous women. Low birth weight was more common amongst infants of S. haematobium-infected mothers. This association was unaffected by controlling for demographic characteristics, gestational age and Plasmodium infection status (adjusted Odds Ratio 1.93; 95% confidence interval: 1.08-3.42). Other risk factors associated with low birth weight delivery were underweight mothers (adjusted Odds Ratio 2.34; 95% confidence interval: 1.12-4.92), peripheral or placental Plasmodium falciparum infection (adjusted Odds Ratio 2.04; 95% confidence interval: 1.18-3.53) and preterm birth (adjusted Odds Ratio 3.12; 95% confidence interval: 1.97-4.96). Preterm delivery was not associated with S. haematobium infection (adjusted Odds Ratio 1.07 95% confidence interval: 0.57-1.98). In conclusion, this study indicates that pregnant women with urogenital schistosomiasis are at an increased risk for low birth weight deliveries. Further studies evaluating targeted treatment and prevention programmes for urogenital schistosomiasis in pregnant women and their impact on delivery outcomes are warranted.

In many tropical areas schistosomiasis is a major health problem causing hepatosplenic, intestinal or urogenital complaints. Hepatosplenic schistosomiasis mansoni is also characterized by blood coagulation abnormalities. Liver pathology plays a role in the development of haemostatic changes and the parasitic infection may directly affect coagulation. However, these contributing factors cannot be studied separately in hepatosplenic schistosomiasis infections. This pilot study provides insight in haemostatic changes in urinary schistosomiasis by studying coagulation parameters in schistosomiasis haematobium-infected Gabonese schoolchildren. Selection on urinary schistosomiasis patients without hepatosplenic complaints allows for the investigation of the direct effects of the parasite on haemostasis. Levels of von Willebrand Factor (VWF) antigen, active VWF and osteoprotegerin were elevated, indicating inflammation-mediated endothelial activation. In contrast to hepatosplenic schistosomiasis, thrombin–antithrombin complex and D-dimer levels were not affected. Despite its small sample size, this study clearly indicates that Schistosoma haematobium directly alters the activation status of the endothelium, without initiation of coagulation.

BackgroundIn sub-Saharan Africa, Plasmodium spp. infection prevalence very often overlaps with helminth infections, particularly with schistosomiasis which is reported to be the second parasitic infection after malaria in terms of prevalence. Interaction between both infections has been reported earlier. Schistosomiasis is typically a chronic disease, whereas malaria occurs in episodes, particularly in children. In this study, we assessed the effect of Schistosoma haematobium infection on clinical malaria among children.MethodsA longitudinal study was conducted from June 2016 to February 2018. Volunteers without any known chronic condition were included. Thick blood smear (TBS) was performed monthly at participants’ homes. For any medical concern including malaria-like symptoms and visible haematuria, participants were invited to come to CERMEL for diagnosis and treatment. Light microscopy was performed to detect malaria parasites and Schistosoma eggs, using TBS and urine filtration technique, respectively. Over the study course, participants found to be infected were treated accordingly. Schistosomiasis status was determined at the end of the follow-up.ResultsAmong the 351 volunteers included in the study, schistosomiasis status was available for 260. Mean age was 12.3 year (SD 4.6) with a 0.96 women-to-men sex ratio. Of those, 112/260 (43.1%) [37.0%–49.3%] participants were positive for S. haematobium, and a total of 132 (51%) children developed 230 malaria attacks. Those with schistosomiasis had a 1.5:1 [1.1–2.0] risk to develop malaria compared to their uninfected counterparts. The mean number of malaria episodes per child over the study course was higher among children with schistosomiasis compared to those without (2.03 vs 1.57, p-value=0.015).ConclusionS. haematobium infection was associated with increased susceptibility to develop malaria (by increasing the risk to develop a malaria episode) and, consequently, a higher malaria incidence.

BackgroundSchistosomiasis is a highly prevalent parasitic infection in Central Africa, where co-endemicity with other parasitic infections is common, and schistosomiasis outcomes can be affected by those other infections. Therefore, proper schistosomiasis control needs epidemiological data accounting for co-infections, too. In this present study, our objective was to determine the epidemiological situation around schistosomiasis in Lambaréné.MethodsA cross-sectional study was conducted among school-aged children living in Lambaréné. Urine filtration exam was performed for the detection of Schistosoma eggs. Kato-Katz and stool culture (Coproculture and Harada-Mori) techniques were used for the detection of soil-transmitted helminths. Detection of Plasmodium spp. and blood microfilariae was performed applying light microscopy. Risk factors for schistosomiasis and factors associated with schistosomiasis were investigated; haematology parameters evaluated.ResultsA total of 614 school children with available schistosomiasis status were included in the analysis. Mean age was 10.9 (SD=2.7) years, with a 0.95 boy-to-girl sex ratio. The prevalence of schistosomiasis was 26%. No risk factors except human-water contact were associated with schistosomiasis. Only Trichuris trichiura co-infection was associated with an increased odd (aOR=2.3, p-value=0.048) to be infected with schistosomiasis. Full blood counts showed a decrease of haemoglobin level and increase of WBC and platelet levels among the schistosoma-infected children. Haematuria was found associated with schistosomiasis (aOR=14.5, p-valueConclusionThe prevalence of schistosomiasis is moderate in Lambaréné where human-water contact remains the main risk factor and praziquantel is available for treatment. Trichuriasis is associated with increased risk to be infected. Children with schistosomiasis exhibit a distinct full blood count profile and haematuria is found to be more suitable to predict infection. However, it is desirable to implement comprehensive approaches beyond chemotherapy for schistosomiasis control in this area as recommended by WHO.

BackgroundDiarrhoeal disease remains the second leading cause of death in children under five years, being associated with about 525,000 deaths every year. The most common pathogens worldwide are Shigella spp/EIEC, rotavirus, adenovirus 40/41, ST-ETEC and Cryptosporidium spp. Public health interventions rely on estimates of pathogen-specific burden for prioritisation. Sadly, comprehensive data on the aetiology of diarrhoea in children is lacking for Gabon. This study aimed to identify the spectrum of pathogens found in Lambaréné, Gabon and provide baseline data on their prevalence, needed for implementation of effective control measures.MethodsA cross-sectional study was conducted at Albert Schweitzer and Georges Rawiri Regional hospitals in Lambaréné from February 2017 to February 2018. A consecutive sample of children under 5 year old with diarrhoea or a history of diarrhoea within the previous three days were prospectively studied. A single stool sample was collected from each study participant and processed using commercial rapid immunoassays to detect antigens of rotavirus, adenovirus, and Cryptosporidium spp. Multiplex PCR was used for Cryptosporidium spp., Giardia lamblia and Cyclospora cayetanensis detection, and characterisation of E. coli strains.ResultsOut of 188 participants who provided stool samples, one or more pathogens could be detected in 34.6% of the cases. The most prevalent parasites were Giardia lamblia (14.9%), Cryptosporidium spp. (11.7%), and Cyclospora cayetanensis (2.7%). Enteric viruses also were identified in these children: 10.6% and 1.6% of rotavirus and adenovirus, respectively. Multiple pathogens were detected in 5.3% of samples.ConclusionThis analysis of the causes of diarrhoea in children under 5 years of age in our setting showed three main pathogens: Giardia lamblia, Cryptosporidium spp. and rotavirus. Our study confirms major agents of acute diarrhoeal diseases in children, highlights research needs (Cryptosporidium) and supports the introduction of new tools such as the implementation of the rotavirus vaccine in the national immunisation programme.

Contains fulltext : 171915.pdf (Publisher’s version ) (Open Access) Infections with helminths and Plasmodium spp. overlap in their geographical distribution. It has been postulated that helminth infections may influence malarial transmission by altering Plasmodium falciparum gametocytogenesis. This cross-sectional study assessed the effect of helminth infections on P. falciparum gametocyte carriage and on humoral immune responses to sexual stage antigens in Gabon. Schistosoma haematobium and filarial infections as well as P. falciparum asexual forms and gametocyte carriage were determined. The antibody responses measured were to sexual (Pfs230, Pfs48/45) and asexual P. falciparum antigens (AMA1, MSP1, and GLURP). A total of 287 subjects were included. The prevalence of microscopically detectable P. falciparum asexual parasites was higher in S. haematobium-infected subjects in comparison to their uninfected counterparts (47% versus 26%, P = 0.003), but this was not different when filarial infections were considered. Plasmodium falciparum gametocyte carriage was similar between Schistosoma- or filaria-infected and uninfected subjects. We observed a significant decrease of Pfs48/45 immunoglobulin G titer in S. haematobium-infected subjects (P = 0.037), whereas no difference was seen for Pfs230 antibody titer, nor for antibodies to AMA1, MSP1, or GLURP. Our findings suggest an effect of S. haematobium on antibody responses to some P. falciparum gametocyte antigens that may have consequences for transmission-blocking immunity.

Background Characterising the effect of natural acquired immunity and sickle cell anaemia on the pattern of Plasmodium falciparum parasitaemia may be useful to understand the pathophysiological mechanisms of protection against malaria. Controlled human malaria infection (CHMI) by direct venous inoculation of aseptic, purified, cryopreserved sporozoites (Pf-SPZ challenge) is a new tool which can be used to investigate the pathophysiology of malaria. Methods The study was performed in Lambarene, Gabon, one of seven African partners in the EDCTP-funded CHMI platform. Adults aged 18–35 from three groups NI: 5 non-immune (NI), 11 semi-immunes with haemoglobin AA (IA), and 9 semi-immunes with haemoglobin AS (IS) received 3,200 sporozoites after a curative treatment course with clindamycin. Capillary blood samples were taken daily up to Day 28 to determine parasitaemia by real-time quantitative polymerase chain reaction (RT-qPCR). Treatment was administered for a malaria episode or at Day 28, whichever came first. Results Parasitaemia was detected in 5 (100%) subjects in the NI group, 9 (82%) in the IA group and 7 (78%) in the IS group. All volunteers in the NI group showed similar patterns with parasitaemia starting around Day 8 and rising quickly. Patterns for parasitaemia in the immune groups (IA and IS) were highly heterogeneous. Although time points of initial parasitaemia and duration of parasitaemia varied, all semi-immunes managed to control parasitaemia for at least several days. There were no discernible differences in patterns between the IS and IA group, including the area under curve of parasitaemia over time. Conclusions No parasitaemia was detected in 20% of the semi-immunes, likely due to liver stage immunity. The highly variable patterns of parasitaemia did not allow us to discern immune mechanisms against blood stages. Haemoglobin AS had no visible effect on parasite dynamics at the low parasitaemia encountered.

Background Malaria remains the first cause of death in Africa. In endemic area, it overlaps with other infections including helminths infections. It has been shown that there are interactions between the two parasites infection. Lambarene is the endemic area for urogenital schistosomiasis, which co-exist with P. falciparum malaria. Therefore, we decide to assess for the first time the effect of schistosomiasis infection on malaria infection burden. Methods In order to assess the effect of S. haematobium on malaria infection burden, a cross-sectional study was conducted in school children aged 6–16 years old. One blood smear was performed and 3 urine samples were obtained to assess the presence of infections. Chi-square test and generalised linear model were used to compare the risk to be infected by P. falciparum parasite and Mann-Whitney-Wilcoxon test to compare the parasitaemia of P. falciparum . Demographic data was also collected. Results A total of 741 children were included. The overall prevalence was 20% and 31% for P. falciparum microscopic carriage and S. haematobium infection, respectively. Co-infection of both was found in 65 (9%) participants. S. haematobium and P. falciparum are highly prevalent in PK compared to Bindo and Makouke areas. At univariable analysis, schistosomiasis-infected subjects have an odd of 2.11 [1.46–3.07] to be infected by P. falciparum parasite compared to non-infected subjects. Locality was found to confound the association which remains significant after adjustment for age, gender and locality (aOR=1.69, [1.13–2.59]). The effect of S. haematobium on the P. falciparum parasitaemia outcome was also assessed. There is no effect of Schistosoma infection on malaria parasite density (p-value=0.92). Conclusions S. haematobium infection increases the risk of being infected with P. falciparum but doesn9t affect the parasitaemia density of P. falciparum malaria in our study population.

In many regions where soil-transmitted helminth infections are endemic, single-dose albendazole is used in mass drug administration programs to control infections. There are little data on the efficacy of the standard single-dose administration compared to that of alternative regimens. We conducted a randomized, controlled, assessor-blinded clinical trial to determine the efficacies of standard and extended albendazole treatment against soil-transmitted helminth infection in Gabon. A total of 175 children were included. Adequate cure rates and egg reduction rates above 85% were found with a single dose of albendazole for Ascaris infection, 85% (95% confidence interval [CI], 73, 96) and 93.8% (CI, 87.6, 100), respectively, while two doses were necessary for hookworm infestation (92% [CI, 78, 100] and 92% [CI, 78, 100], respectively). However, while a 3-day regimen was not sufficient to cure Trichuris (cure rate, 83% [CI, 73, 93]), this regimen reduced the number of eggs up to 90.6% (CI, 83.1, 100). The rate ratios of two- and three-dose regimens compared to a single-dose treatment were 1.7 (CI, 1.1, 2.5) and 2.1 (CI, 1.5, 2.9) for Trichuris and 1.7 (CI, 1.0, 2.9) and 1.7 (CI, 1.0, 2.9) for hookworm. Albendazole was safe and well tolerated in all regimens. A single-dose albendazole treatment considerably reduces Ascaris infection but has only a moderate effect on hookworm and Trichuris infections. The single-dose option may still be the preferred regimen because it balances efficacy, safety, and compliance during mass drug administration, keeping in mind that asymptomatic low-level helminth carriage may also have beneficial effects. (This study has been registered at ClinicalTrials.gov under registration number NCT01192802.)