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Most patients with Post COVID Syndrome (PCS) present with a plethora of symptoms without clear evidence of organ dysfunction. A subset of them fulfills diagnostic criteria of myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS). Symptom severity of ME/CFS correlates with natural regulatory autoantibody (AAB) levels targeting several G-protein coupled receptors (GPCR). In this exploratory study, we analyzed serum AAB levels against vaso- and immunoregulatory receptors, mostly GPCRs, in 80 PCS patients following mild-to-moderate COVID-19, with 40 of them fulfilling diagnostic criteria of ME/CFS. Healthy seronegative (n=38) and asymptomatic post COVID-19 controls (n=40) were also included in the study as control groups. We found lower levels for various AABs in PCS compared to at least one control group, accompanied by alterations in the correlations among AABs. Classification using random forest indicated AABs targeting ADRB2, STAB1, and ADRA2A as the strongest classifiers (AABs stratifying patients according to disease outcomes) of post COVID-19 outcomes. Several AABs correlated with symptom severity in PCS groups. Remarkably, severity of fatigue and vasomotor symptoms were associated with ADRB2 AAB levels in PCS/ME/CFS patients. Our study identified dysregulation of AAB against various receptors involved in the autonomous nervous system (ANS), vaso-, and immunoregulation and their correlation with symptom severity, pointing to their role in the pathogenesis of PCS.

Highlights • Malaria cases in UK and Ireland travellers can give false-negative HRP-RDT results. • Histidine-rich protein (HRP2/3) deletions can cause false-negative HRP-RDT results. • High parasitemia may give false-negative RDT results due to a prozone-like effect. • False-negative RDT results may also be a result of low parasite density. • Next-generation sequencing can elaborate the breakpoints of HRP2/3 deletions., Objectives Our objective was to systematically investigate false-negative histidine-rich protein 2 rapid diagnostic tests (HRP2-RDT) in imported Plasmodium falciparum malaria cases from travelers to the UK and the Republic of Ireland (RoI). Methods Five imported malaria cases in travellers returning to the UK and RoI from East Africa were reported to the PHE Malaria Reference Laboratory as negative according to histidine-rich protein (HRP2)-RDT. The cases were systematically investigated using microscopic, RDT, molecular, genomic, and in in vitro approaches. Results In each case, HRP2-RDT was negative, whereas microscopy confirmed the presence of P. falciparum. Further analysis revealed that the genes encoding HRP2 and HRP3 were deleted in three of the five cases. Whole-genome sequencing in one of these isolates confirmed deletions in P. falciparum chromosomes 8 and 13. Our study produced evidence that the fourth case, which had high parasitemia at clinical presentation, was a rare example of antigen saturation (‘prozone-like effect’), leading to a false negative in the HRP2-RDT, while the fifth case was due to low parasitemia. Conclusions False-negative HRP2-RDT results with P. falciparum are concerning. Our findings emphasise the necessity of supporting the interpretation of RDT results with microscopy, in conjunction with clinical observations, and sets out a systematic approach to identifying parasites carrying pfhrp2 and pfhrp3 deletions.

In November 2015, cases of Zika virus infection were recorded in Cabo Verde (Africa), originating from Brazil. The outbreak subsided after seven months with 7580 suspected cases. We performed a serological survey (n = 431) in Praia, the capital city, 3 months after transmission ceased. Serum samples were screened for arbovirus antibodies using ELISA techniques and revealed seroconverted individuals with Zika (10.9%), dengue (1–4) (12.5%), yellow fever (0.2%) and chikungunya (2.6%) infections. Zika seropositivity was predominantly observed amongst females (70%). Using a logistic model, risk factors for increased odds of Zika seropositivity included age, self-reported Zika infection, and dengue seropositivity. Serological data from Zika and dengue virus assays were strongly correlated (Spearman’s rs = 0.80), which reduced when using a double antigen binding ELISA (Spearman’s rs = 0.54). Overall, our work improves an understanding of how Zika and other arboviruses have spread throughout the Cabo Verde population. It also demonstrates the utility of serological assay formats for outbreak investigations.

Highlights • In Vanuatu, ocular Chlamydia infection prevalence is low; in Kiribati it is high. • In Vanuatu, Pgp3 seroprevalence does not increase in childhood; in Kiribati it does. • Conjunctival scarring is more common in adults in Kiribati than in Vanuatu. • Trachomatous inflammation—follicular lacks specificity for ocular Chlamydia infection. • Non-TF markers may help to determine need for interventions against active trachoma., Introduction In the peri-elimination setting, the positive predictive value of trachomatous inflammation–follicular (TF), the primary marker used to determine need for antibiotics for trachoma, is suboptimal. Here, three non-TF measures are used to compare two regions where TF prevalence exceeds the threshold for intervention, but where the Chlamydia trachomatis (Ct) prevalence is different. Methods Population prevalence of trachoma was measured in Vanuatu (n = 3470) and Kiribati (n = 2922). Dried blood spots (DBS) and conjunctival photographs were collected from every survey participant, and conjunctival swabs were collected from those aged 1–9 years. Individuals were tested for blood anti-Pgp3 antibodies, Ct DNA at the conjunctiva and severity of conjunctival scarring. Results The prevalence of TF in 1–9-year-olds was 16.5% in Vanuatu and 38.2% in Tarawa. 7% of people aged ≥1 year in Vanuatu had conjunctival scarring compared to 27% in Tarawa. The prevalence of ocular Ct infection in 1–9-year-olds was 1.5% in Vanuatu and 27.4% in Tarawa. The seroconversion rate amongst 1–9-year-old children in Vanuatu and Tarawa was 0.018 and 0.197 events per child per year, respectively. Conclusions Comparing Vanuatu to Tarawa demonstrates several markers that could be used to differentiate the trachoma status of populations in these (and other) locations.

Background: \ud Passively collected malaria case data are the foundation for public health decision making. However, because of population-level immunity, infections might not always be sufficiently symptomatic to prompt individuals to seek care. Understanding the proportion of all Plasmodium spp infections expected to be detected by the health system becomes particularly paramount in elimination settings. The aim of this study was to determine the association between the proportion of infections detected and transmission intensity for Plasmodium falciparum and Plasmodium vivax in several global endemic settings.\ud \ud Methods: \ud The proportion of infections detected in routine malaria data, P(Detect), was derived from paired household cross-sectional survey and routinely collected malaria data within health facilities. P(Detect) was estimated using a Bayesian model in 431 clusters spanning the Americas, Africa, and Asia. The association between P(Detect) and malaria prevalence was assessed using log-linear regression models. Changes in P(Detect) over time were evaluated using data from 13 timepoints over 2 years from The Gambia.\ud \ud Findings: \ud The median estimated P(Detect) across all clusters was 12·5% (IQR 5·3–25·0) for P falciparum and 10·1% (5·0–18·3) for P vivax and decreased as the estimated log-PCR community prevalence increased (adjusted odds ratio [OR] for P falciparum 0·63, 95% CI 0·57–0·69; adjusted OR for P vivax 0·52, 0·47–0·57). Factors associated with increasing P(Detect) included smaller catchment population size, high transmission season, improved care-seeking behaviour by infected individuals, and recent increases (within the previous year) in transmission intensity.\ud \ud Interpretation: \ud The proportion of all infections detected within health systems increases once transmission intensity is sufficiently low. The likely explanation for P falciparum is that reduced exposure to infection leads to lower levels of protective immunity in the population, increasing the likelihood that infected individuals will become symptomatic and seek care. These factors might also be true for P vivax but a better understanding of the transmission biology is needed to attribute likely reasons for the observed trend. In low transmission and pre-elimination settings, enhancing access to care and improvements in care-seeking behaviour of infected individuals will lead to an increased proportion of infections detected in the community and might contribute to accelerating the interruption of transmission.\ud \ud Funding: \ud Wellcome Trust.

Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is a complex disease characterized by unexplained debilitating fatigue. Although the etiology is unknown, evidence supports immunological abnormalities, such as persistent inflammation and immune-cell activation, in a subset of patients. Since the interplay between inflammation and vascular alterations is well-established in other diseases, endothelial dysfunction has emerged as another player in ME/CFS pathogenesis. Endothelial nitric oxide synthase (eNOS) generates nitric oxide (NO) that maintains endothelial homeostasis. eNOS is activated by silent information regulator 1 (Sirt1), an anti-inflammatory protein. Despite its relevance, no study has addressed the Sirt1/eNOS axis in ME/CFS. The interest in circulating microRNAs (miRs) as potential biomarkers in ME/CFS has increased in recent years. Accordingly, we analyze a set of miRs reported to modulate the Sirt1/eNOS axis using plasma from ME/CFS patients. Our results show that miR-21, miR-34a, miR-92a, miR-126, and miR-200c are jointly increased in ME/CFS patients compared to healthy controls. A similar finding was obtained when analyzing public miR data on peripheral blood mononuclear cells. Bioinformatics analysis shows that endothelial function-related signaling pathways are associated with these miRs, including oxidative stress and oxygen regulation. Interestingly, histone deacetylase 1, a protein responsible for epigenetic regulations, represented the most relevant node within the network. In conclusion, our study provides a basis to find endothelial dysfunction-related biomarkers and explore novel targets in ME/CFS.

Background Many health facilities in malaria endemic countries are dependent on Rapid diagnostic tests (RDTs) for diagnosis and some National Health Service (NHS) hospitals without expert microscopists rely on them for diagnosis out of hours. The emergence of P. falciparum lacking the gene encoding histidine-rich protein 2 and 3 (HRP2 and HRP3) and escaping RDT detection threatens progress in malaria control and elimination. Currently, confirmation of RDT negative due to the deletion of pfhrp2 and pfhrp3, which encodes a cross-reactive protein isoform, requires a series of PCR assays. These tests have different limits of detection and many laboratories have reported difficulty in confirming the absence of the deletions with certainty. Methods We developed and validated a novel and rapid multiplex real time quantitative (qPCR) assay to detect pfhrp2, pfhrp3, confirmatory parasite and human reference genes simultaneously. We also applied the assay to detect pfhrp2 and pfhrp3 deletion in 462 field samples from different endemic countries and UK travellers. Results The qPCR assay demonstrated diagnostic sensitivity of 100% (n = 19, 95% CI= (82.3%; 100%)) and diagnostic specificity of 100% (n = 31; 95% CI= (88.8%; 100%)) in detecting pfhrp2 and pfhrp3 deletions. In addition, the assay estimates P. falciparum parasite density and accurately detects pfhrp2 and pfhrp3 deletions masked in polyclonal infections. We report pfhrp2 and pfhrp3 deletions in parasite isolates from Kenya, Tanzania and in UK travellers. Interpretation The new qPCR is easily scalable to routine surveillance studies in countries where P. falciparum parasites lacking pfhrp2 and pfhrp3 are a threat to malaria control.

Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is a debilitating disease characterized by severe and persistent fatigue. Along with clinical studies showing endothelial dysfunction (ED) in a subset of ME/CFS patients, we have recently reported altered ED-related microRNAs in plasma from affected individuals. Inadequate nitric oxide (NO), mainly produced by the endothelial isoform of nitric oxide synthase (eNOS) in endothelial cells (ECs), is a major cause of ED. In this study, we hypothesized that plasma from that cohort of ME/CFS patients induces eNOS-related ED in vitro. To test this, we cultured human umbilical vein endothelial cells (HUVECs) in the presence of plasma from either ME/CFS patients (ME/CFS-plasma, n = 11) or healthy controls (HC-plasma, n = 12). Then, we measured the NO production in the absence and presence of tyrosine kinase and G protein-coupled receptors agonists (TKRs and GPCRs, respectively), well-known to activate eNOS in ECs. Our data showed that HUVECs incubated with ME/CFS-plasma produced less NO either in the absence or presence of eNOS activators compared to ones in presence of HC-plasma. Also, the NO production elicited by bradykinin, histamine, and acetylcholine (GPCRs agonists) was more affected than the one triggered by insulin (TKR agonist). Finally, inhibitory eNOS phosphorylation at Thr

Background Aedes spp. are responsible for the transmission of many arboviruses, which contribute to rising human morbidity and mortality worldwide. The Aedes aegypti mosquito is a main vector for chikungunya, dengue and yellow fever infections, whose incidence have been increasing and distribution expanding. This vector has also driven the emergence of the Zika virus (ZIKV), first reported in Africa which spread rapidly to Asia and more recently across the Americas. During the outbreak in the Americas, Cape Verde became the first African country declaring a Zika epidemic, with confirmed cases of microcephaly. Here we investigate the prevalence of ZIKV and dengue (DENV) infected Ae. aegypti mosquitoes in the weeks following the outbreak in Cape Verde, and the presence of insecticide resistance in the circulating vector population. Genetic diversity in the mosquito population was also analysed. Methods From August to October 2016, 816 Ae. aegypti mosquitoes were collected in several locations across Praia, Cape Verde, the major hot spot of reported ZIKV cases in the country. All mosquitoes were screened by reverse transcription PCR for ZIKV and DENV, and a subset (n = 220) were screened for knockdown insecticide resistance associated mutations in the voltage gated sodium channel (VGSC) gene by capillary sequencing. The mitochondrial NADH dehydrogenase subunit 4 (nad4) gene was sequenced in 100 mosquitoes. These data were compared to 977 global sequences in a haplotype network and a phylogenetic tree analysis. Results Two Ae. aegypti mosquitoes were ZIKV positive (0.25%). There were no SNP mutations found in the VGSC gene associated with insecticide resistance. Analysis of the nad4 gene revealed 11 haplotypes in the Cape Verdean samples, with 5 being singletons. Seven haplotypes were exclusive to Cape Verde. Several of the remaining haplotypes were frequent in the global dataset, being present in several countries (including Cape Verde) across five different continents. The most common haplotype in Cape Verde (50.6 %) was also found in Africa and South America. Conclusions There was low-level Zika virus circulation in mosquitoes from Praia shortly after the outbreak. The Ae. aegypti population did not appear to have the kdr mutations associated with pyrethroid resistance. Furthermore, haplotype and phylogenetic analyses revealed that Cape Verde Ae. aegypti mosquitoes are most closely related to those from other countries in Africa and South America.

Background: Antimalarial antibody measurements are useful because they reflect historical and recent exposure to malaria. As such, they may provide additional information to assess ongoing transmission in low endemic or pre-elimination settings where cases are rare. In addition, the absence of antibody responses in certain individuals can indicate the cessation of transmission. Commercial malaria enzyme-linked immunosorbent assays (ELISA) detect antimalarial antibodies and are commonly used to screen blood donations for possible malaria infection. However, there is no standardized test to detect antimalarial antibodies for epidemiological use. Here we compared five commercially available ELISA kits (Trinity Biotech, newbio, DiaPro, Cellabs, and NovaTec) in search of a standardized tool for supporting claims of absence of malaria transmission. For comparison, a research-based (RB) ELISA protocol was performed alongside the commercial kits.\ud \ud Results: The commercial kits were first compared using serum samples from known malaria-unexposed individuals (n = 223) and Toxoplasma-infected individuals (n = 191) to assess specificity and cross-reactivity against non-malaria infections. In addition, 134 samples from ≥10-year-olds collected in a hyperendemic region in the Gambia in the early 1990s were used to assess sensitivity. Three out of five kits showed high sensitivity (90–92%), high specificity (98–99%), low cross-reactivity (0–3%) and were considered user-friendly (Trinity Biotech, newbio and NovaTec). Two of these kits (Trinity Biotech and NovaTec) were taken forward for epidemiological evaluation and results were compared to those using the RB-ELISA. Samples from two pre-elimination settings (Praia, Cape Verde; n = 1,396, and Bataan, the Philippines; n = 1,824) were tested. Serological results from both the Trinity Biotech kit and the RB-ELISA concurred with recent passively detected case counts in both settings. Results from the Trinity Biotech kit reflected a significant decrease in the number of reported cases in Bataan in the 1990s better than the RB-ELISA. Results from the NovaTec kit did not reflect transmission patterns in either setting.\ud \ud Conclusion: The Trinity Biotech commercial ELISA kit was considered reliable for epidemiological use and accurately described transmission patterns in two (previously) malaria endemic settings. The use of this simple and standardized serological tool may aid national control and elimination programs by confirming that regions are free from malaria.

Measuring antimalarial antibodies can estimate transmission in a population. To compare outputs, standardized laboratory testing is required. Here we describe the in-country establishment and quality control (QC) of a multiplex bead assay (MBA) for three sero-surveys in Haiti. Total IgG data against 21 antigens were collected for 32,758 participants. Titration curves of hyperimmune sera were included on assay plates, assay signals underwent 5-parameter regression, and inspection of the median and interquartile range (IQR) for the y-inflection point was used to determine assay precision. The medians and IQRs were similar for Surveys 1 and 2 for most antigens, while the IQRs increased for some antigens in Survey 3. Levey-Jennings charts for selected antigens provided a pass/fail criterion for each assay plate and, of 387 assay plates, 13 (3.4%) were repeated. Individual samples failed if IgG binding to the generic glutathione-S-transferase protein was observed, with 659 (2.0%) samples failing. An additional 455 (1.4%) observations failed due to low bead numbers (

Data Availability: The datasets generated for this study are available on request to the corresponding author. Copyright © 2019 Cliff, King, Lee, Sepúlveda, Wolf, Kingdon, Bowman, Dockrell, Nacul, Lacerda and Riley. Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is a debilitating condition with unknown aetiology, Myalgic encephalomyelitis unclear pathophysiology and with no diagnostic test or biomarker available. Many patients report their ME/CFS began after an acute infection, and subsequent increased frequency of infections, such as colds or influenza, is common. These factors imply an altered immunological status exists in ME/CFS, in at least a proportion of patients, yet previous studies of peripheral immunity have been discrepant and inconclusive. The UK ME/CFS Biobank, which has collected blood samples from nearly 300 clinically-confirmed ME/CFS patients, enables large-scale studies of immunological function in phenotypically well-characterised participants. In this study, herpes virus serological status and T cell, B cell, NK cell and monocyte populations were investigated in 251 ME/CFS patients, including 54 who were severely affected, and compared with those from 107 healthy participants and with 46 patients with Multiple Sclerosis. There were no differences in seroprevalence for six human herpes viruses between ME/CFS and healthy controls, although seroprevalence for the Epstein-Barr virus was higher in multiple sclerosis patients. Contrary to previous reports, no significant differences were observed in NK cell numbers, subtype proportions or in vitro responsiveness between ME/CFS patients and healthy control participants. In contrast, the T cell compartment was altered in ME/CFS, with increased proportions of effector memory CD8+ T cells and decreased proportions of terminally differentiated effector CD8+ T cells. Conversely, there was a significantly increased proportion of mucosal associated invariant T cells (MAIT) cells, especially in severely affected ME/CFS patients. These abnormalities demonstrate that an altered immunological state does exist in ME/CFS, particularly in severely affected people. This may simply reflect ongoing or recent infection, or may indicate future increased susceptibility to infection. Longitudinal studies of ME/CFS patients are needed to help to determine cause and effect and thus any potential benefits of immuno-modulatory treatments for ME/CFS. We thank all the study participants for their time and energy and for donating their blood to the UK ME/CFS Biobank (UKMEB). We also thank the support from the charities The ME Association, Action for ME, and ME Research UK, as well as a private donor, who funded the UKMEB start-up and to the ME Association for continued funding. National Institute of Allergy and Infectious Diseases (NIAID) of the National Institutes of Health (NIH) under Award Number: R01AI103629. NS acknowledges funding from Fundação para a Ciência e Tecnologia, Portugal (grant ref. UID/MAT/00006/2013).

Autoimmunity and chronic viral infections are recurrent clinical observations in Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS), a complex disease with an unknown cause. Given these observations, the regulatory CD4+ T cells (Tregs) show promise to be good candidates for the underlying pathology due to their capacity to suppress the immune responses against both self and microbial antigens. Here, we discussed the overlooked role of these cells in the chronicity of Human Herpes Virus 6 (HHV6), Herpes Simplex 1 (HSV1), and Epstein-Barr virus (EBV), as often reported as triggers of ME/CFS. Using simulations of the cross-regulation model for the dynamics of Tregs, we illustrated that mild infections might lead to a chronically activated immune responses under control of Tregs if the responding clone has a high autoimmune potential. Such infections promote persistent inflammation and possibly fatigue. We then hypothesized that ME/CFS is a condition characterized by a predominance of this type of infections under control of Tregs. In contrast, healthy individuals are hypothesized to trigger immune responses of a virus-specific clone with a low autoimmune potential. According to this hypothesis, simple model simulations of the CD4+ T-cell repertoire could reproduce the increased density and percentages of Tregs observed in patients suffering from the disease, when compared to healthy controls. A deeper analysis of Tregs in the pathogenesis of ME/CFS will help to assess the validity of this hypothesis.

International audience; In this trans-ethnic multi-omic study, we reinterpret the genetic architecture of blood pressure to identify genes, tissues, phenomes and medication contexts of blood pressure homeostasis. We discovered 208 novel common blood pressure SNPs and 53 rare variants in genome-wide association studies of systolic, diastolic and pulse pressure in up to 776,078 participants from the Million Veteran Program (MVP) and collaborating studies, with analysis of the blood pressure clinical phenome in MVP. Our transcriptome-wide association study detected 4,043 blood pressure associations with genetically predicted gene expression of 840 genes in 45 tissues, and mouse renal single-cell RNA sequencing identified upregulated blood pressure genes in kidney tubule cells.

Background: Plasmodium ovale and Plasmodium malariae infections are scarcely studied in sub-Saharan Africa, where the Plasmodium falciparum species predominates. The objective of this study is to investigate the prevalence of P. ovale and P. malariae infections and their relationship with common red blood cell polymorphisms in a cohort of 509 individuals from Uganda. Methods: Three cross-sectional surveys were conducted in individuals of 1–10 and >20 y of age from the Apac district at baseline and 6 and 16 weeks after drug treatment. Malaria infections were assessed by polymerase chain reaction and genotyping was performed for the sickle-cell allele, α-thalassaemia and glucose-6-phosphate dehydrogenase. Results: At baseline, the prevalence of infection was 7.5%, 12.6% and 57.4% for P. ovale, P. malariae and P. falciparum species, respectively. Co-infections were present in 14.1% of individuals, all including P. falciparum parasites. In children 1–5 y of age, the prevalence of P. ovale mono-infections increased significantly from 1.7% to 7.3% over time (p=0.004) while the prevalence of P. malariae and P. falciparum infections declined significantly during this study. After adjusting for confounding and multiple testing, only α-thalassaemia had a statistically significant increase in the odds of P. falciparum infections (odds ratio 1.93 [95% confidence interval 1.26 to 2.94]). Conclusions: Common red blood cell polymorphisms do not show strong effects on mild Plasmodium infections in this Ugandan population. To understand the extent of this result, similar studies should be carried out in other populations using larger cohorts.

Significant selection pressure has been exerted on the genomes of human populations exposed to Plasmodium falciparum infection, resulting in the acquisition of mechanisms of resistance against severe malarial disease. Many host genetic factors, including sickle cell trait, have been associated with reduced risk of developing severe malaria, but do not account for all of the observed phenotypic variation. Identification of novel inherited risk factors relies upon high-resolution genome-wide association studies (GWAS). We present findings of a GWAS of severe malaria performed in a Tanzanian population (n = 914, 15.2 million SNPs). Beyond the expected association with the sickle cell HbS variant, we identify protective associations within two interleukin receptors (IL-23R and IL-12RBR2) and the kelch-like protein KLHL3 (all P0.3). Our approach demonstrates the potential of a joint genotyping-sequencing strategy to identify as-yet unknown susceptibility loci in an African population with well-characterised malaria phenotypes. The regions encompassing these loci are potential targets for the design of much needed interventions for preventing or treating malarial disease., Author summary Malaria, caused by Plasmodium falciparum parasites, is a major cause of mortality and morbidity in endemic countries of sub-Saharan Africa, including Tanzania. Some gene mutations in the human genome, including sickle cell trait, have been associated with reduced risk of developing severe malaria, and have increased in frequency through natural selection over generations. However, new genetic mutations remain to be discovered, and recent advances in human genome research technologies such as genome-wide association studies (GWAS) and fine-scale molecular genotyping tools, are facilitating their identification. Here, we present findings of a GWAS of severe malaria performed in a well characterised Tanzanian population (n = 914). We confirm the expected association with the sickle cell trait, but also identify new gene targets in immunological pathways, some under natural selection. Our approach demonstrates the potential of using GWAS to identify as-yet unknown susceptibility genes in endemic populations with well-characterised malaria phenotypes. The genetic mutations are likely to form potential targets for the design of much needed interventions for preventing or treating malarial disease.

Trachoma is caused byChlamydia trachomatis(Ct). It is targeted for global elimination as a public health problem. In 2014, a population-based cross-sectional study was performed in two previously trachoma-endemic areas of The Gambia. Participants of all ages from Lower River Region (LRR) (N = 1028) and Upper River Region (URR) (N = 840) underwent examination for trachoma and had blood collected for detection of antibodies against the Ct antigen Pgp3, by ELISA. Overall, 30 (1.6%) individuals had active trachoma; the prevalence in children aged 1–9 years was 3.4% (25/742) with no statistically significant difference in prevalence between the regions. There was a significant difference in overall seroprevalence by region: 26.2% in LRR and 17.1% in URR (p

Summary Genetic association within several malaria candidate genes was examined in 24 villages of northeast Tanzania, using different measures of malaria transmission intensity. We demonstrate that the classic hemoglobinopathies were associated with measures of transmission intensity that had longer time scales., Background. Human malaria susceptibility is determined by multiple genetic factors. It is unclear, however, which genetic variants remain important over time. Methods. Genetic associations of 175 high-quality polymorphisms within several malaria candidate genes were examined in a sample of 8096 individuals from northeast Tanzania using altitude, seroconversion rates, and parasite rates as proxies of historical, recent, and current malaria transmission intensity. A principal component analysis was used to derive 2 alternative measures of overall malaria propensity of a location across different time scales. Results. Common red blood cell polymorphisms (ie, hemoglobin S, glucose-6-phosphate dehydrogenase, and α-thalassemia) were the only ones to be associated with all 3 measures of transmission intensity and the first principal component. Moderate associations were found between some immune response genes (ie, IL3 and IL13) and parasite rates, but these could not be reproduced using the alternative measures of malaria propensity. Conclusions. We have demonstrated the potential of using altitude and seroconversion rate as measures of malaria transmission capturing medium- to long-term time scales to detect genetic associations that are likely to persist over time. These measures also have the advantage of minimizing the deleterious effects of random factors affecting parasite rates on the respective association signals.

Background Sri Lanka achieved the WHO certificate as a malaria free country in September 2016, thus monitoring of malaria transmission using sensitive and effective tools is an important need. Use of age-specific antibody prevalence as a serological tool to predict transmission intensity is proven to be a cost effective and reliable method under elimination settings. This paper discusses the correlation of four anti-malarial antibodies against vivax and falciparum malaria with the declining transmission intensities in two previously high malaria endemic districts i.e. Kurunegala and Moneragala of Sri Lanka. Methods Sera was collected from 1,186 individuals from the two districts and were subjected to standard ELISA together with control sera from non-immune individuals to obtain Optical Density (OD) values for four anti-malarial antibodies i.e. anti-MSP1 and anti-AMA1 for both Plasmodium vivax and Plasmodium falciparum. The sero-positive samples were determined as mean OD + 3SD of the negative controls. The sero-prevalence was analyzed against the demographic characteristics of the population. A simple reversible catalytic model was fitted into sero-prevalence data to predict the sero-conversion and sero-reversion rates. Results Over 60% of the population was sero-positive for one or more antibodies except young children (

Deletions of the Plasmodium falciparum hrp2 and hrp3 genes can affect the performance of HRP2-based malaria rapid diagnostic tests (RDTs). Such deletions have been reported from South America, India and Eritrea. Whether these parasites are widespread in East Africa is unknown. A total of 274 samples from asymptomatic children in Mbita, western Kenya, and 61 genomic data from Kilifi, eastern Kenya, were available for analysis. PCR-confirmed samples were investigated for the presence of pfhrp2 and pfhrp3 genes. In samples with evidence of deletion, parasite presence was confirmed by amplifying three independent genes. We failed to amplify pfhrp2 from 25 of 131 (19.1%) PCR-confirmed samples. Of these, only 8 (10%) samples were microscopic positive and were classified as pfhrp2-deleted. Eight microscopically-confirmed pfhrp2-deleted samples with intact pfhrp3 locus were positive by HRP2-based RDT. In addition, one PCR-confirmed infection showed a deletion at the pfhrp3 locus. One genomic sample lacked pfhrp2 and one lacked pfhrp3. No sample harbored parasites lacking both genes. Parasites lacking pfhrp2 are present in Kenya, but may be detectable by HRP-based RDT at higher parasitaemia, possibly due to the presence of intact pfhrp3. These findings warrant further systematic study to establish prevalence and diagnostic significance.

Many human genetic associations with resistance to malaria have been reported, but few have been reliably replicated. We collected data on 11,890 cases of severe malaria due to Plasmodium falciparum and 17,441 controls from 12 locations in Africa, Asia and Oceania. We tested 55 SNPs in 27 loci previously reported to associate with severe malaria. There was evidence of association at P < 1 × 10(-4) with the HBB, ABO, ATP2B4, G6PD and CD40LG loci, but previously reported associations at 22 other loci did not replicate in the multicenter analysis. The large sample size made it possible to identify authentic genetic effects that are heterogeneous across populations or phenotypes, with a striking example being the main African form of G6PD deficiency, which reduced the risk of cerebral malaria but increased the risk of severe malarial anemia. The finding that G6PD deficiency has opposing effects on different fatal complications of P. falciparum infection indicates that the evolutionary origins of this common human genetic disorder are more complex than previously supposed.

It has been previously shown that there are some interethnic differences in susceptibility to malaria between two sympatric ethnic groups of Mali, the Fulani and the Dogon. The lower susceptibility to Plasmodium falciparum malaria seen in the Fulani has not been fully explained by genetic polymorphisms previously known to be associated with malaria resistance, including haemoglobin S (HbS), haemoglobin C (HbC), alpha-thalassaemia and glucose-6-phosphate dehydrogenase (G6PD) deficiency. Given the observed differences in the distribution of FcγRIIa allotypes among different ethnic groups and with malaria susceptibility that have been reported, we analysed the rs1801274-R131H polymorphism in the FcγRIIa gene in a study of Dogon and Fulani in Mali (n = 939). We confirm that the Fulani have less parasite densities, less parasite prevalence, more spleen enlargement and higher levels of total IgG antibodies (anti-CSP, anti-AMA1, anti-MSP1 and anti-MSP2) and more total IgE (P

Pakistan faces critical challenges pertaining to cutaneous leishmaniasis (CL), where it's distribution is more or less patchy. The goal of this study was to assess the incidence of CL as well as to identify potential risk factors in Peshawar region, Khyber Pakhtunkhwa, Pakistan. The study was conducted in the dermatology outpatient unit at Kuwait Teaching Hospital, Peshawar, Khyber Pakhtunkhwa, Pakistan. Longitudinal out-patient department visit data for 9,631 CL patients spanning a 42-mo (April 2011-October 2014) period was analyzed using autoregressive integrated moving average (ARIMA) time series models ARIMA(1,0,0)(0,1,0)12 and ARIMA(0,0,0)(0,1,0)12. The ARIMA concluded that the number of patients was increasing over time. Over the duration, frequency of male patients (58.2%) was higher. The mean age of CL patients was 16.4 (confidence interval = 16.14-16.70) yr and the majority of the patients were aged 5-20 yr (52.6%). Inflow of CL patients peaked close to February and March, followed by a decline until its lowest point in the months of August and September (P 20 yr) to derive 63 matched pairs. Using univariate conditional logistic regression analyses of the matched pairs, we found that living in congested rooms (>6 persons), having family members with lesions (active/scars), keeping cattle inside dwellings at night, and having in-door vegetation were established as factors that significantly increased the risk of CL. On the other hand, living in houses constructed with bricked walls or wooden roofs (thatched/beam), ownership of treated bed nets, and having meshed windows were proven to be protective against CL. It was evident that the disease incidence has been on a gradual rise over the past few years. It was concluded that household clustering, house construction, and conventional behavioral practices (living with cattle) greatly impact the epidemiology of CL in the region. Conclusions from this study have significant implications for prospective control programs.

Background Antigenic polymorphisms are considered as one of the main strategies employed by malaria parasites to escape from the host immune responses after infections. Merozoite surface protein-1 (MSP-1) of Plasmodium vivax, a promising vaccine candidate, is a highly polymorphic protein whose immune recognition is not well understood. Methods and results The IgG responses to conserved (MSP-119) and polymorphic (block 2 and block 10) epitopes of PvMSP-1 were evaluated in 141 P. vivax infected patients. Ten recombinant proteins corresponding to block 2 (variants BR07, BP29, BP39, BP30, BEL) and block 10 (BR07, BP29, BP39, BP01, BP13) often observed in Brazilian P. vivax isolates were assessed by ELISA in order to determine levels of specific antibodies and their respective seroprevalence. The magnitude and the frequency of variant-specific responses were very low, except for BR07 variant (>40%), which was the predominant haplotype as revealed by block 10 PvMSP-1 gene sequencing. By contrast, 89% of patients had IgG against the C-terminal conserved domain (PvMSP-119), confirming the high antigenicity of this protein. Using multiple linear and logistic regression models, there was evidence for a negative association between levels of haemoglobin and several IgG antibodies against block 2 variant antigens, with the strongest association being observed for BP39 allelic version. This variant was also found to increase the odds of anaemia in these patients. Conclusions These findings may have implications for vaccine development and represent an important step towards a better understanding of the polymorphic PvMSP-1 domain as potential targets of vaccine development. These data highlight the importance of extending the study of these polymorphic epitopes of PvMSP-1 to different epidemiological settings. Electronic supplementary material The online version of this article (doi:10.1186/s12936-016-1612-z) contains supplementary material, which is available to authorized users.

IntroductionMyalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is a chronic disease involving central nervous system and immune system disorders, as well as cardiovascular abnormalities. ME/CFS is characterised by severe chronic fatigue lasting for at least 6 months, including clinical symptoms such as tender cervical or axillary lymph nodes, muscle pain, joint pain without swelling or redness, post-exertional malaise for more than 24 hours and unrefreshing sleep. Studies on the epidemiology of ME/CFS in Europe only include single countries and, therefore, the prevalence and incidence of ME/CFS in Europe (as a whole) is unknown. One of the purposes of the European Network on ME/CFS (EUROMENE; European Union-funded COST Action; Reference number: 15111) is to address this gap in knowledge. We will systematically review the literature reporting figures from European countries to provide a robust summary and identify new challenges.Methods and analysisWe will systematically search the literature databases Scopus, PubMed and Web of Science for studies published in the last 10 years (ie, after 2007). No language restriction will be applied. Two independent reviewers will search, screen and select studies as well as extract data about their main characteristics and evaluate their methodological and reporting quality. When disagreements emerge, the reviewers will discuss to reach a consensus. We plan to produce a narrative summary of our findings as we anticipate that studies are scarce and heterogeneous. The possibility of performing meta-analyses will be discussed in a EUROMENE meeting.Ethics and disseminationEthical approval is not required as only publicly available data will be included. Findings will be described in EUROMENE reports, published in peer-reviewed journal(s) and presented at conferences. The findings will be also communicated to policy-makers, healthcare providers, people with ME/CFS and other sections of society through regular channels including the mass-media.PROSPERO registration numberCRD42017078688

Background Malaria morbidity and mortality has declined in recent years in a number of settings. The ability to describe changes in malaria transmission associated with these declines is important in terms of assessing the potential effects of control interventions, and for monitoring and evaluation purposes. Methods Data from five cross-sectional surveys conducted in Farafenni and surrounding villages on the north bank of River Gambia between 1988 and 2011 were compiled. Antibody responses to MSP-119 were measured in samples from all surveys, data were normalized and expressed as seroprevalence and seroconversion rates (SCR) using different mathematical models. Results Results showed declines in serological metrics with seroprevalence in children aged one to 5 years dropping from 19 % (95 % CI 15–23 %) in 1988 to 1 % (0–2 %) in 2011 (p value for trend in proportions

Summary Background The global prevalence of X-linked glucose-6-phosphate dehydrogenase (G6PD) deficiency is thought to be a result of selection by malaria, but epidemiological studies have yielded confusing results. We investigated the relationships between G6PD deficiency and both malaria and non-malarial illnesses among children in Kenya. Methods We did this study in Kilifi County, Kenya, where the G6PD c.202T allele is the only significant cause of G6PD deficiency. We tested the associations between G6PD deficiency and severe and complicated Plasmodium falciparum malaria through a case-control study of 2220 case and 3940 control children. Cases were children aged younger than 14 years, who visited the high dependency ward of Kilifi County Hospital with severe malaria between March 1, 1998, and Feb 28, 2010. Controls were children aged between 3–12 months who were born within the same study area between August 2006, and September 2010. We assessed the association between G6PD deficiency and both uncomplicated malaria and other common diseases of childhood in a cohort study of 752 children aged younger than 10 years. Participants of this study were recruited from a representative sample of households within the Ngerenya and Chonyi areas of Kilifi County between Aug 1, 1998, and July 31, 2001. The primary outcome measure for the case-control study was the odds ratio for hospital admission with severe malaria (computed by logistic regression) while for the cohort study it was the incidence rate ratio for uncomplicated malaria and non-malaria illnesses (computed by Poisson regression), by G6PD deficiency category. Findings 2863 (73%) children in the control group versus 1643 (74%) in the case group had the G6PD normal genotype, 639 (16%) versus 306 (14%) were girls heterozygous for G6PD c.202T, and 438 (11%) versus 271 (12%) children were either homozygous girls or hemizygous boys. Compared with boys and girls without G6PD deficiency, we found significant protection from severe malaria (odds ratio [OR] 0·82, 95% CI 0·70–0·97; p=0·020) among G6PD c.202T heterozygous girls but no evidence for protection among G6PD c.202T hemizygous boys and homozygous girls (OR 1·18, 0·99–1·40; p=0·056). Median follow-up for the mild disease cohort study was 2·24 years (IQR 2·22–2·85). G6PD c.202T had no effect on other common diseases of childhood in heterozygous girls (incidence rate ratio 0·98, 95% CI 0·86–1·11; p=0·82) or homozygous girls or hemizygous boys (0·93, 0·82–1·04; p=0·25), with the sole exception of a marginally significant increase in the incidence of helminth infections among heterozygous girls. Interpretation Heterozygous girls might be the driving force for the positive selection of G6PD deficiency alleles. Further studies are needed to definitively establish the mechanisms by which G6PD deficiency confers an advantage against malaria in heterozygous individuals. Such studies could lead to the development of new treatments. Funding Wellcome Trust, UK Medical Research Council, European Union, and Foundation for the National Institutes of Health (as part of the Bill & Melinda Gates Grand Challenges in Global Health Initiative).

The last decade has witnessed a steady reduction of the malaria burden worldwide. With various countries targeting disease elimination in the near future, the popular parasite infection or entomological inoculation rates are becoming less and less informative of the underlying malaria burden due to a reduced number of infected individuals or mosquitoes at the time of sampling. To overcome such problem, alternative measures based on antibodies against specific malaria antigens have gained recent interest in malaria epidemiology due to the possibility of estimating past disease exposure in absence of infected individuals. This paper aims then to review current mathematical models and corresponding statistical approaches used in antibody data analysis. The application of these models is illustrated with three data sets from Equatorial Guinea, Brazilian Amazonia region, and western Kenyan highlands. A brief discussion is also carried out on the future challenges of using these models in the context of malaria elimination.

Populations exposed to Plasmodium falciparum infection develop genetic mechanisms of protection against severe malarial disease. Despite decades of genetic epidemiological research, the sickle cell trait (HbAS) sickle cell polymorphism, ABO blood group, and other hemoglobinopathies remain the few major determinants in severe malaria to be replicated across different African populations and study designs. Within a case-control study in a region of high transmission in Tanzania (n = 983), we investigated the role of 40 new loci identified in recent genome-wide studies. In 32 loci passing quality control procedures, we found polymorphisms in USP38, FREM3, SDC1, DDC, and LOC727982 genes to be putatively associated with differential susceptibility to severe malaria. Established candidates explained 7.4% of variation in severe malaria risk (HbAS polymorphism, 6.3%; alpha-thalassemia, 0.3%; ABO group, 0.3%; and glucose-6-phosphate dehydrogenase deficiency, 0.5%) and the new polymorphisms, another 4.3%. The regions encompassing the loci identified are promising targets for the design of future treatment and control interventions.

BACKGROUND: Measurement of malaria endemicity is typically based on vector or parasite measures. A complementary approach is the detection of parasite specific IgG antibodies. We determined the antibody levels and seroconversion rates to both P. vivax and P. falciparum merozoite antigens in individuals living in areas of varying P. vivax endemicity in Pará state, Brazilian Amazon region.METHODOLOGY/PRINCIPAL FINDINGS: The prevalence of antibodies to recombinant antigens from P. vivax and P. falciparum was determined in 1,330 individuals. Cross sectional surveys were conducted in the north of Brazil in Anajás, Belém, Goianésia do Pará, Jacareacanga, Itaituba, Trairão, all in the Pará state, and Sucuriju, a free-malaria site in the neighboring state Amapá. Seroprevalence to any P. vivax antigens (MSP1 or AMA-1) was 52.5%, whereas 24.7% of the individuals were seropositive to any P. falciparum antigens (MSP1 or AMA-1). For P. vivax antigens, the seroconversion rates (SCR) ranged from 0.005 (Sucuriju) to 0.201 (Goianésia do Pará), and are strongly correlated to the corresponding Annual Parasite Index (API). We detected two sites with distinct characteristics: Goianésia do Pará where seroprevalence curve does not change with age, and Sucuriju where seroprevalence curve is better described by a model with two SCRs compatible with a decrease in force of infection occurred 14 years ago (from 0.069 to 0.005). For P. falciparum antigens, current SCR estimates varied from 0.002 (Belém) to 0.018 (Goianésia do Pará). We also detected a putative decrease in disease transmission occurred ∼29 years ago in Anajás, Goianésia do Pará, Itaituba, Jacareacanga, and Trairão.CONCLUSIONS: We observed heterogeneity of serological indices across study sites with different endemicity levels and temporal changes in the force of infection in some of the sites. Our study provides further evidence that serology can be used to measure and monitor transmission of both major species of malaria parasite.

Multiple imputation based on chained equations (MICE) is an alternative missing genotype method that can use genetic and nongenetic auxiliary data to inform the imputation process. Previously, MICE was successfully tested on strongly linked genetic data. We have now tested it on data of the HBA2 gene which, by the experimental design used in a malaria association study in Tanzania, shows a high missing data percentage and is weakly linked with the remaining genetic markers in the data set. We constructed different imputation models and studied their performance under different missing data conditions. Overall, MICE failed to accurately predict the true genotypes. However, using the best imputation model for the data, we obtained unbiased estimates for the genetic effects, and association signals of the HBA2 gene on malaria positivity. When the whole data set was analyzed with the same imputation model, the association signal increased from 0.80 to 2.70 before and after imputation, respectively. Conversely, postimputation estimates for the genetic effects remained the same in relation to the complete case analysis but showed increased precision. We argue that these postimputation estimates are reasonably unbiased, as a result of a good study design based on matching key socio-environmental factors.

© 2012 Gomes-Santos et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited., Plasmodium sporozoites are transmitted by Anopheles mosquitoes and infect hepatocytes, where a single sporozoite replicates into thousands of merozoites inside a parasitophorous vacuole. The nature of the Plasmodium-host cell interface, as well as the interactions occurring between these two organisms, remains largely unknown. Here we show that highly dynamic hepatocyte actin reorganization events occur around developing Plasmodium berghei parasites inside human hepatoma cells. Actin reorganization is most prominent between 10 to 16 hours post infection and depends on the actin severing and capping protein, gelsolin. Live cell imaging studies also suggest that the hepatocyte cytoskeleton may contribute to parasite elimination during Plasmodium development in the liver., The work was supported by Fundação para a Ciência e Tecnologia (FCT) of the Portuguese Ministry of Science (PTDC/SAU-GMG/100313/2008), the Federal German Ministry of education and Science (Biofuture) C.S.S.G-S. was supported by an FCT fellowship (SFRH/BD/15888/2005). M.A.I. was funded by the EU FP 7 Marie Curie Initial Training Network ‘‘Intervention Strategies against Malaria (InterMalTraining)’’, contract no. 215281. L.F.M. is a Young Investigator from the Human Frontier Science Program and receives support from Fundação Luso-Americana para o Desenvolvimento and Fundação para a Ciência e a Tecnologia (PTDC/SAU-MII/69280/2006 and PTDC/SAU-MII/78333/2006). F.F. thanks the Chica and Heinz Schaller Foundation for support. M.M.M. and F.F. are members of the EU FP7 Network of Excellence EVIMalaR

© 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim, Treg cells, best identified by the expression of the transcription factor FOXP3, play a crucial role in maintaining self-tolerance. Natural Treg cells constitute an independent thymus-derived T-cell lineage whose developmental program in humans is still ill-defined. Here, we provide evidence of a Treg-cell differentiation pathway at the double positive (DP) stage, prior to commitment to the CD4(+) or CD8(+) lineage, in pediatric thymuses. FOXP3(+) DP cells displayed a functional IL-7 receptor and increased Bcl-2 levels that may protect them from cell death/negative selection, and an activated/suppressive phenotype that was lost as CD4 single positive (SP) cells matured and acquired egress markers. A subpopulation of FOXP3(+) DP thymocytes expressing CD103 likely represents the precursor of FOXP3(+) CD8SP cells, which homogeneously expressed this mucosal-homing molecule. Finally, co-cultures of DP thymocytes with primary thymic epithelial cells and multiple linear regression analyses support that FOXP3(+) SP cells are largely derived from FOXP3(+) DP thymocytes. Overall, our data suggest that human Treg-cell lineage commitment significantly occurs at the DP stage with possible implications for the diversity and autoreactivity of the natural Treg-cell repertoire., This work was supported by grant (PTDC/ SAU-MI/66248/2006) from Fundação para a Ciência e a Tecnologia (FCT) and by Programa Operacional Ciência e Inovação 2010 (POCI2010) to AES. H. N. C. received a scholarship from FCT co-financed by POCI 2010.

BACKGROUND: Cerebral malaria (CM) represents a severe outcome of the Plasmodium falciparum infection. Recent genetic studies have correlated human genes with severe malaria susceptibility, but there is little data on genetic variants that increase the risk of developing specific malaria clinical complications. Nevertheless, susceptibility to experimental CM in the mouse has been linked to host genes including Transforming Growth Factor Beta 2 (TGFB2) and Heme oxygenase-1 (HMOX1). Here, we tested whether those genes were governing the risk of progressing to CM in patients with severe malaria syndromes. METHODOLOGY/PRINCIPAL FINDINGS: We report that the clinical outcome of P. falciparum infection in a cohort of Angolan children (n = 430) correlated with nine TGFB2 SNPs that modify the risk of progression to CM as compared to other severe forms of malaria. This genetic effect was explained by two haplotypes harboring the CM-associated SNPs (Pcorrec. = 0.035 and 0.036). In addition, one HMOX1 haplotype composed of five CM-associated SNPs increased the risk of developing the CM syndrome (Pcorrec. = 0.002) and was under-transmitted to children with uncomplicated malaria (P = 0.036). Notably, the HMOX1-associated haplotype conferred increased HMOX1 mRNA expression in peripheral blood cells of CM patients (P = 0.012). CONCLUSIONS/SIGNIFICANCE: These results represent the first report on CM genetic risk factors in Angolan children and suggest the novel hypothesis that genetic variants of the TGFB2 and HMOX1 genes may contribute to confer a specific risk of developing the CM syndrome in patients with severe P. falciparum malaria. This work may provide motivation for future studies aiming to replicate our findings in larger populations and to confirm a role for these genes in determining the clinical course of malaria.

Low-grade polymicrobial infection induced by cecal ligation and puncture is lethal in heme oxygenase-1-deficient mice (Hmox1(-/-)), but not in wild-type (Hmox1(+/+)) mice. Here we demonstrate that the protective effect of this heme-catabolizing enzyme relies on its ability to prevent tissue damage caused by the circulating free heme released from hemoglobin during infection. Heme administration after low-grade infection in mice promoted tissue damage and severe sepsis. Free heme contributed to the pathogenesis of severe sepsis irrespective of pathogen load, revealing that it compromised host tolerance to infection. Development of lethal forms of severe sepsis after high-grade infection was associated with reduced serum concentrations of the heme sequestering protein hemopexin (HPX), whereas HPX administration after high-grade infection prevented tissue damage and lethality. Finally, the lethal outcome of septic shock in patients was also associated with reduced HPX serum concentrations. We propose that targeting free heme by HPX might be used therapeutically to treat severe sepsis.

Background Malawi experienced prolonged use of sulfadoxine/pyrimethamine (SP) as the front-line anti-malarial drug, with early replacement of chloroquine and delayed introduction of artemisinin-based combination therapy. Extended use of SP, and its continued application in pregnancy is impacting the genomic variation of the Plasmodium falciparum population. Methods Whole genome sequence data of P. falciparum isolates covering 2 years of transmission within Malawi, alongside global datasets, were used. More than 745,000 SNPs were identified, and differences in allele frequencies between countries assessed, as well as genetic regions under positive selection determined. Results Positive selection signals were identified within dhps, dhfr and gch1, all components of the parasite folate pathway associated with SP resistance. Sitting predominantly on a dhfr triple mutation background, a novel copy number increase of ~twofold was identified in the gch1 promoter. This copy number was almost fixed (96.8% frequency) in Malawi samples, but found at less than 45% frequency in other African populations, and distinct from a whole gene duplication previously reported in Southeast Asian parasites. Conclusions SP resistance selection pressures have been retained in the Malawian population, with known resistance dhfr mutations at fixation, complemented by a novel gch1 promoter duplication. The effects of the duplication on the fitness costs of SP variants and resistance need to be elucidated. Electronic supplementary material The online version of this article (doi:10.1186/s12936-016-1634-6) contains supplementary material, which is available to authorized users.

BACKGROUND: Glucose-6-phosphate dehydrogenase (G6PD) deficiency is associated with protection from severe malaria, and potentially uncomplicated malaria phenotypes. It has been documented that G6PD deficiency in sub-Saharan Africa is due to the 202A/376G G6PD A-allele, and association studies have used genotyping as a convenient technique for epidemiological studies. However, recent studies have shown discrepancies in G6PD202/376 associations with severe malaria. There is evidence to suggest that other G6PD deficiency alleles may be common in some regions of West Africa, and that allelic heterogeneity could explain these discrepancies. METHODS: A cross-sectional epidemiological study of malaria susceptibility was conducted during 2006 and 2007 in the Sahel meso-endemic malaria zone of Mali. The study included Dogon (n = 375) and Fulani (n = 337) sympatric ethnic groups, where the latter group is characterized by lower susceptibility to Plasmodium falciparum malaria. Fifty-three G6PD polymorphisms, including 202/376, were genotyped across the 712 samples. Evidence of association of these G6PD polymorphisms and mild malaria was assessed in both ethnic groups using genotypic and haplotypic statistical tests. RESULTS: It was confirmed that the Fulani are less susceptible to malaria, and the 202A mutation is rare in this group (