Gene expression analyses reveal differences in children's response to malaria according to their age.

In Bandiagara, Mali, children experience on average two clinical malaria episodes per year. However, even in the same transmission area, the number of uncomplicated symptomatic infections, and their parasitemia, can vary dramatically among children. We simultaneously characterize host and parasite gene expression profiles from 136 Malian children with symptomatic falciparum malaria and examine differences in the relative proportion of immune cells and parasite stages, as well as in gene expression, associated with infection and or patient characteristics. Parasitemia explains much of the variation in host and parasite gene expression, and infections with higher parasitemia display proportionally more neutrophils and fewer T cells, suggesting parasitemia-dependent neutrophil recruitment and/or T cell extravasation to secondary lymphoid organs. The child's age also strongly correlates with variations in gene expression: Plasmodium falciparum genes associated with age suggest that older children carry more male gametocytes, while variations in host gene expression indicate a stronger innate response in younger children and stronger adaptive response in older children. These analyses highlight the variability in host responses and parasite regulation during P. falciparum symptomatic infections and emphasize the importance of considering the children's age when studying and treating malaria infections.

Randomized Field Trial to Assess the Safety and Efficacy of Dihydroartemisinin-Piperaquine for Seasonal Malaria Chemoprevention in School-Aged Children in Bandiagara, Mali.

BACKGROUND: Owing to the increased cases of malaria in older children, the World Health Organization has recently recommended extending seasonal malaria chemoprevention (SMC) to children >5 years of age and using other effective drugs for malaria. In this study, we report the safety and efficacy of dihydroartemisinin-piperaquine (DHA-PQ) for SMC in school-aged children in Mali. METHOD: This randomized, controlled trial included 345 participants aged 6-15 years randomized to receive DHA-PQ, sulfadoxine-pyrimethamine plus amodiaquine (SP-AQ), or no chemoprevention (albendazole) at a 1:1:1 ratio. Four rounds of SMC were conducted from September to December 2021. The participants were assessed 7 days after each round for safety and efficacy of the interventions. RESULTS: Abdominal pain (11.8% vs 29.2%), headache (11.2% vs 19.2%), and vomiting (5.7% vs 15.2%) were frequently reported in the DHA-PQ and SP-AQ arms. On Day 120 of follow up, the incidence of clinical malaria was 0.01 episodes/person-month in the DHA-PQ and SP-AQ arms and 0.17 episodes/person-month in the control arm (P < .0001). Gametocytes were detected in 37 participants in all arms. CONCLUSIONS: Children in DHA-PQ arm reported less adverse events compared to the SP-AQ arm. Both drugs were effective against clinical malaria and infection.

Gene expression analyses reveal differences in children's response to malaria according to their age.

In Bandiagara, Mali, children experience on average two clinical malaria episodes per season. However, even in the same transmission area, the number of uncomplicated symptomatic infections, and their parasitemia, vary dramatically among children. To examine the factors contributing to these variations, we simultaneously characterized the host and parasite gene expression profiles from 136 children with symptomatic falciparum malaria and analyzed the expression of 9,205 human and 2,484 Plasmodium genes. We used gene expression deconvolution to estimate the relative proportion of immune cells and parasite stages in each sample and to adjust the differential gene expression analyses. Parasitemia explained much of the variation in both host and parasite gene expression and revealed that infections with higher parasitemia had more neutrophils and fewer T cells, suggesting parasitemia-dependent neutrophil recruitment and/or T cell extravasation to secondary lymphoid organs. The child's age was also strongly correlated with gene expression variations. Plasmodium falciparum genes associated with age suggested that older children carried more male gametocytes, while host genes associated with age indicated a stronger innate response (through TLR and NLR signaling) in younger children and stronger adaptive immunity (through TCR and BCR signaling) in older children. These analyses highlight the variability in host responses and parasite regulation during P. falciparum symptomatic infections and emphasize the importance of considering the children's age when studying and treating malaria infections.

Gene expression analyses reveal differences in children's response to malaria according to their age.

In Bandiagara, Mali, children experience on average two clinical malaria episodes per season. However, even in the same transmission area, the number of uncomplicated symptomatic infections, and their parasitemia, vary dramatically among children. To examine the factors contributing to these variations, we simultaneously characterized the host and parasite gene expression profiles from 136 children with symptomatic falciparum malaria and analyzed the expression of 9,205 human and 2,484 Plasmodium genes. We used gene expression deconvolution to estimate the relative proportion of immune cells and parasite stages in each sample and to adjust the differential gene expression analyses. Parasitemia explained much of the variation in both host and parasite gene expression and revealed that infections with higher parasitemia had more neutrophils and fewer T cells, suggesting parasitemia-dependent neutrophil recruitment and/or T cell extravasation to secondary lymphoid organs. The child's age was also strongly correlated with gene expression variations. Plasmodium falciparum genes associated with age suggested that older children carried more male gametocytes, while host genes associated with age indicated a stronger innate response (through TLR and NLR signaling) in younger children and stronger adaptive immunity (through TCR and BCR signaling) in older children. These analyses highlight the variability in host responses and parasite regulation during P. falciparum symptomatic infections and emphasize the importance of considering the children's age when studying and treating malaria infections.

Natural immunity to malaria preferentially targets the endothelial protein C receptor-binding regions of PfEMP1s.

Antibody responses to variant surface antigens (VSAs) produced by the malaria parasite Plasmodium falciparum may contribute to age-related natural immunity to severe malaria. One VSA family, P. falciparum erythrocyte membrane protein-1 (PfEMP1), includes a subset of proteins that binds endothelial protein C receptor (EPCR) in human hosts and potentially disrupts the regulation of inflammatory responses, which may lead to the development of severe malaria. We probed peptide microarrays containing segments spanning five PfEMP1 EPCR-binding domain variants with sera from 10 Malian adults and 10 children to determine the differences between adult and pediatric immune responses. We defined serorecognized peptides and amino acid residues as those that elicited a significantly higher antibody response than malaria-naïve controls. We aimed to identify regions consistently serorecognized among adults but not among children across PfEMP1 variants, potentially indicating regions that drive the development of immunity to severe malaria. Adult sera consistently demonstrated broader and more intense serologic responses to constitutive PfEMP1 peptides than pediatric sera, including peptides in EPCR-binding domains. Both adults and children serorecognized a significantly higher proportion of EPCR-binding peptides than peptides that do not directly participate in receptor binding, indicating a preferential development of serologic responses at functional residues. Over the course of a single malaria transmission season, pediatric serological responses increased between the start and the peak of the season, but waned as the transmission season ended. IMPORTANCE Severe malaria and death related to malaria disproportionately affect sub-Saharan children under 5 years of age, commonly manifesting as cerebral malaria and/or severe malarial anemia. In contrast, adults in malaria-endemic regions tend to experience asymptomatic or mild disease. Our findings indicate that natural immunity to malaria targets specific regions within the EPCR-binding domain, particularly peptides containing EPCR-binding residues. Epitopes containing these residues may be promising targets for vaccines or therapeutics directed against severe malaria. Our approach provides insight into the development of natural immunity to a binding target linked to severe malaria by characterizing an "adult-like" response as recognizing a proportion of epitopes within the PfEMP1 protein, particularly regions that mediate EPCR binding. This "adult-like" response likely requires multiple years of malaria exposure, as increases in pediatric serologic response over a single malaria transmission season do not appear significant.

Immunogenomic profile at baseline predicts host susceptibility to clinical malaria.

Introduction: Host gene and protein expression impact susceptibility to clinical malaria, but the balance of immune cell populations, cytokines and genes that contributes to protection, remains incompletely understood. Little is known about the determinants of host susceptibility to clinical malaria at a time when acquired immunity is developing. Methods: We analyzed peripheral blood mononuclear cells (PBMCs) collected from children who differed in susceptibility to clinical malaria, all from a small town in Mali. PBMCs were collected from children aged 4-6 years at the start, peak and end of the malaria season. We characterized the immune cell composition and cytokine secretion for a subset of 20 children per timepoint (10 children with no symptomatic malaria age-matched to 10 children with >2 symptomatic malarial illnesses), and gene expression patterns for six children (three per cohort) per timepoint. Results: We observed differences between the two groups of children in the expression of genes related to cell death and inflammation; in particular, inflammatory genes such as CXCL10 and STAT1 and apoptotic genes such as XAF1 were upregulated in susceptible children before the transmission season began. We also noted higher frequency of HLA-DR+ CD4 T cells in protected children during the peak of the malaria season and comparable levels cytokine secretion after stimulation with malaria schizonts across all three time points. Conclusion: This study highlights the importance of baseline immune signatures in determining disease outcome. Our data suggests that differences in apoptotic and inflammatory gene expression patterns can serve as predictive markers of susceptibility to clinical malaria.

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

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

Malian children infected with Plasmodium ovale and Plasmodium falciparum display very similar gene expression profiles.

Plasmodium parasites caused 241 million cases of malaria and over 600,000 deaths in 2020. Both P. falciparum and P. ovale are endemic to Mali and cause clinical malaria, with P. falciparum infections typically being more severe. Here, we sequenced RNA from nine pediatric blood samples collected during infections with either P. falciparum or P. ovale, and characterized the host and parasite gene expression profiles. We found that human gene expression varies more between individuals than according to the parasite species causing the infection, while parasite gene expression profiles cluster by species. Additionally, we characterized DNA polymorphisms of the parasites directly from the RNA-seq reads and found comparable levels of genetic diversity in both species, despite dramatic differences in prevalence. Our results provide unique insights into host-pathogen interactions during malaria infections and their variations according to the infecting Plasmodium species, which will be critical to develop better elimination strategies against all human Plasmodium parasites.

The estimated burden of fungal diseases in Mali.

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

Shifts in the clinical epidemiology of severe malaria after scaling up control strategies in Mali.

A decrease in malaria incidence following implementation of control strategies such as use of artemisinin-based combination therapies, insecticide-impregnated nets, intermittent preventive treatment during pregnancy and seasonal malaria chemoprevention (SMC) has been observed in many parts of Africa. We hypothesized that changes in malaria incidence is accompanied by a change in the predominant clinical phenotypes of severe malaria. To test our hypothesis, we used data from a severe malaria case-control study that lasted from 2014-2019 to describe clinical phenotypes of severe forms experienced by participants enrolled in Bandiagara, Bamako, and Sikasso, in Mali. We also analyzed data from hospital records of inpatient children at a national referral hospital in Bamako. Among 97 cases of severe malaria in the case-control study, there was a predominance of severe malarial anemia (49.1%). The frequency of cerebral malaria was 35.4, and 16.5% of cases had a mixed clinical phenotype (concurrent cerebral malaria and severe anemia). National referral hospital record data in 2013-15 showed 24.3% of cases had severe malarial anemia compared to 51.7% with cerebral malaria. In the years after SMC scale-up, severe malarial anemia cases increased to 30.1%, (P = 0.019), whereas cerebral malaria cases decreased to 45.5% (P = 0.025). In addition, the predominant age group for each severe malaria phenotype was the 0-1-year-olds. The decrease in malaria incidence noted with the implementation of control strategies may be associated with a change in the clinical expression patterns of severe malaria, including a potential shift in severe malaria burden to age groups not receiving seasonal malaria chemoprevention.

PfSPZ-CVac malaria vaccine demonstrates safety among malaria-experienced adults: A randomized, controlled phase 1 trial.

Background: Plasmodium falciparum (Pf) Sporozoite (SPZ) Chemoprophylaxis Vaccine (PfSPZ-CVac) involves concurrently administering infectious PfSPZ and malaria drug, often chloroquine (CQ), to kill liver-emerging parasites. PfSPZ-CVac (CQ) protected 100% of malaria-naïve participants against controlled human malaria infection. We investigated the hypothesis that PfSPZ-CVac (CQ) is safe and efficacious against seasonal, endemic Pf in malaria-exposed adults. Methods: Healthy 18-45 year olds were enrolled in a double-blind, placebo-controlled trial in Bougoula-Hameau, Mali, randomized 1:1 to 2.048 × 105 PfSPZ (PfSPZ Challenge) or normal saline administered by direct venous inoculation at 0, 4, 8 weeks. Syringes were prepared by pharmacy staff using online computer-based enrolment that randomized allocations. Clinical team and participant masking was assured by identical appearance of vaccine and placebo. Participants received chloroquine 600mg before first vaccination, 10 weekly 300mg doses during vaccination, then seven daily doses of artesunate 200mg before 24-week surveillance during the rainy season. Safety outcomes were solicited adverse events (AEs) and related unsolicited AEs within 12 days of injections, and all serious AEs. Pf infection was detected by thick blood smears performed every four weeks and during febrile illness over 48 weeks. Primary vaccine efficacy (VE) endpoint was time to infection at 24 weeks. NCT02996695. Findings: 62 participants were enrolled in April/May 2017. Proportions of participants experiencing at least one solicited systemic AE were similar between treatment arms: 6/31 (19.4%, 95%CI 9.2-36.3) of PfSPZ-CVac recipients versus 7/31 (22.6%, 95%CI 29.2-62.2) of controls (p value = 1.000). Two/31 (6%) in each group reported related, unsolicited AEs. One unrelated death occurred. Of 59 receiving 3 immunizations per protocol, fewer vaccinees (16/29, 55.2%) became infected than controls (22/30, 73.3%). VE was 33.6% by hazard ratio (p = 0.21, 95%CI -27·9, 65·5) and 24.8% by risk ratio (p = 0.10, 95%CI -4·8, 54·3). Antibody responses to PfCSP were poor; 28% of vaccinees sero-converted. Interpretation: PfSPZ-CVac (CQ) was well-tolerated. The tested dosing regimen failed to significantly protect against Pf infection in this very high transmission setting. Funding: U.S. National Institutes of Health, Sanaria. Registration number: ClinicalTrials.gov identifier (NCT number): NCT02996695.

Serologic and Cytokine Profiles of Children with Concurrent Cerebral Malaria and Severe Malarial Anemia Are Distinct from Other Subtypes of Severe Malaria.

We used a protein microarray featuring Plasmodium falciparum field variants of a merozoite surface antigen to examine malaria exposure in Malian children with different severe malaria syndromes. Unlike children with cerebral malaria alone or severe malarial anemia alone, those with concurrent cerebral malaria and severe malarial anemia had serologic responses demonstrating a broader prior parasite exposure pattern than matched controls with uncomplicated disease. Comparison of levels of malaria-related cytokines revealed that children with the concurrent phenotype had elevated levels of interleukin (IL)-6, IL-8, and IL-10. Our results suggest that the pathophysiology of this severe subtype is unique and merits further investigation.

[Epidemiological and etiological profile of vulvovaginal candidiasis in women in consultation in the gynecology-obstetrics department, CHU Gabriel Toure, Bamako - Mali]. / Profil epidemiologique et etiologique des candidoses vulvovaginales chez les femmes en consultation dans le service de gynecologie-obstetrique, CHU Gabriel Toure, Bamako - Mali.

Vulvovaginal candidiasis (VVC) is a fungal infection caused by yeasts of the genus Candida that leads to vulvar pruritus and vaginal discharge. METHOD: In order to evaluate the epidemiological and etiological Profile of vvc, we carried out a cross-sectional study among women in consultation in the gynecological department of the CHU-Gabriel Toure in Bamako. Two swabs were taken from each woman for mycological diagnosis. The presence of yeasts and pseudo-filaments was observed on direct examination. The culture was performed on Sabouraud-Chloramphenicol medium and at 37°C for 24 to 48 hours. Identification was based on the macroscopic and microscopic characteristics of the colonies, the germinative tube test and the Vitek•2 instrument. of the colonies, the germinative tube test and the VITEK® 2 instrument. RESULT: A total of 240 women were included with a mean age of 31.5 ± 3.15 [15-64 years]. Married women represented 91.67% (n=220) and 51.25% were housewises. Pruritus 49.17% (118/240) and dyspareunia 42.08% (101/240) were the most frequent clinical signs. Previous use of antifungals was demonstrated in 85.83% of women. Candida species were confirmed in 60.42% (145/240) of cases. C. albicans was the most frequent species followed by C. famata, C. dubliniensis, C. krusei.. This study allowed us to identify the most frequent cases of C. albicans, followed by C. famata, C. dubliniensis, and C. krusei..Further studies are still needed to characterize the antifungal susceptibility profile of the Candida species involved.

La candidose vulvo-vaginale (CVV) est une infection fongique causée par des levures du genre Candida provoquant du prurit vulvaire et des pertes vaginales. MÉTHODE: Afin d'évaluer le profil épidémiologique et étiologique des CVV, nous avons réalisé une étude transversale chez les femmes en consultation dans le service de gynécologie du CHU-Gabriel TOURE de Bamako. Deux écouvillons vaginaux ont été prélevés sur chaque femme pour le diagnostic mycologique. La présence de levures et de pseudo-filaments a été observée à l'examen direct. La culture a été réalisée sur milieu Sabouraud-Chloramphénicol et à 37°C pendant 24 à 48 heures. L'identification a été basée sur les caractéristiques macroscopiques et microscopiques des colonies, le test du tube germinatif et l'instrument VITEK® 2. RÉSULTAT: Un total de 240 femmes ont été incluses avec un âge moyen de 31,5 ans ± 3,15 [15-64 ans]. Les femmes mariées représentaient 91,67% et 51,25% étaient des menagères. Le prurit 49,17 % et la dyspareunie 42,08 % (101/240) étaient les signes cliniques les plus fréquents. La prise antérieure d'antifongiques a été retrouvée chez 85,83% des femmes. La présence des espèces de Candida a été confirmée dans 60,42 % des cas. L'espèce C. albicansétait lus fréquente suivies de C. famata, C. dubliniensis, C. krusei.. CONCLUSION: Les résultats de cette étude permettent d'élargir les connaissances sur l'épidémiologie du CVV au Mali. D'autres études restent nécessaires pour caractériser le profil de sensibilité aux antifongiques des espèces de Candida impliquées.

Successful Profiling of Plasmodium falciparum var Gene Expression in Clinical Samples via a Custom Capture Array.

var genes encode Plasmodium falciparum erythrocyte membrane protein-1 (PfEMP1) antigens. These highly diverse antigens are displayed on the surface of infected erythrocytes and play a critical role in immune evasion and sequestration of infected erythrocytes. Studies of var expression using non-leukocyte-depleted blood are challenging because of the predominance of host genetic material and lack of conserved var segments. Our goal was to enrich for parasite RNA, allowing de novo assembly of var genes and detection of expressed novel variants. We used two overall approaches: (i) enriching for total mRNA in the sequencing library preparations and (ii) enriching for parasite RNA with a custom capture array based on Roche's SeqCap EZ enrichment system. The capture array was designed with probes based on the whole 3D7 reference genome and an additional >4,000 full-length var gene sequences from other P. falciparum strains. We tested each method on RNA samples from Malian children with severe or uncomplicated malaria infections. All reads mapping to the human genome were removed, the remaining reads were assembled de novo into transcripts, and from these, var-like transcripts were identified and annotated. The capture array produced the longest maximum length and largest numbers of var gene transcripts in each sample, particularly in samples with low parasitemia. Identifying the most-expressed var gene sequences in whole-blood clinical samples without the need for extensive processing or generating sample-specific reference genome data is critical for understanding the role of PfEMP1s in malaria pathogenesis. IMPORTANCE Malaria parasites display antigens on the surface of infected red blood cells in the human host that facilitate attachment to blood vessels, contributing to the severity of infection. These antigens are highly variable, allowing the parasite to evade the immune system. Identifying these expressed antigens is critical to understanding the development of severe malarial disease. However, clinical samples contain limited amounts of parasite genetic material, a challenge for sequencing efforts further compounded by the extreme diversity of the parasite surface antigens. We present a method that enriches for these antigen sequences in clinical samples using a custom capture array, requiring minimal processing in the field. While our results are focused on the malaria parasite Plasmodium falciparum, this approach has broad applicability to other highly diverse antigens from other parasites and pathogens such as those that cause giardiasis and leishmaniasis.

Immunoprofiles associated with controlled human malaria infection and naturally acquired immunity identify a shared IgA pre-erythrocytic immunoproteome.

Knowledge of the Plasmodium falciparum antigens that comprise the human liver stage immunoproteome is important for pre-erythrocytic vaccine development, but, compared with the erythrocytic stage immunoproteome, more challenging to classify. Previous studies of P. falciparum antibody responses report IgG and rarely IgA responses. We assessed IgG and IgA antibody responses in adult sera collected during two controlled human malaria infection (CHMI) studies in malaria-naïve volunteers and in 1- to 6-year-old malaria-exposed Malian children on a 251 P. falciparum antigen protein microarray. IgG profiles in the two CHMI groups were equivalent and differed from Malian children. IgA profiles were robust in the CHMI groups and a subset of Malian children. We describe immunoproteome differences in naïve vs. exposed individuals and report pre-erythrocytic proteins recognized by the immune system. IgA responses detected in this study expand the list of pre-erythrocytic antigens for further characterization as potential vaccine candidates.

Draft Genome Sequence of Bacillus velezensis Strain Marseille-Q1230, Isolated from a Stool Sample from a Severely Malnourished Child.

Bacillus velezensis, a species first described in 2005, has been mostly associated with plants and the environment. To date, there is no genome available for this species from human samples. In this announcement, we present the genome of Bacillus velezensis strain Marseille-Q1230, which was isolated from a stool sample from a child suffering from severe acute malnutrition. The genome assembled into 15 contigs and had a size of 3,861,152 bp, with a GC content of 46.6%. A total of 3,716 protein-coding genes, including 3 antibiotic resistance genes and 92 RNAs, were predicted.

Plasmodium falciparum transcription in different clinical presentations of malaria associates with circulation time of infected erythrocytes.

Following Plasmodium falciparum infection, individuals can remain asymptomatic, present with mild fever in uncomplicated malaria cases, or show one or more severe malaria symptoms. Several studies have investigated associations between parasite transcription and clinical severity, but no broad conclusions have yet been drawn. Here, we apply a series of bioinformatic approaches based on P. falciparum's tightly regulated transcriptional pattern during its ~48-hour intraerythrocytic developmental cycle (IDC) to publicly available transcriptomes of parasites obtained from malaria cases of differing clinical severity across multiple studies. Our analysis shows that within each IDC, the circulation time of infected erythrocytes without sequestering to endothelial cells decreases with increasing parasitaemia or disease severity. Accordingly, we find that the size of circulating infected erythrocytes is inversely related to parasite density and disease severity. We propose that enhanced adhesiveness of infected erythrocytes leads to a rapid increase in parasite burden, promoting higher parasitaemia and increased disease severity.

Malian adults maintain serologic responses to virulent PfEMP1s amid seasonal patterns of fluctuation.

Plasmodium falciparum erythrocyte membrane protein-1s (PfEMP1s), diverse malaria proteins expressed on the infected erythrocyte surface, play an important role in pathogenesis, mediating adhesion to host vascular endothelium. Antibodies to particular non-CD36-binding PfEMP1s are associated with protection against severe disease. We hypothesized that given lifelong P. falciparum exposure, Malian adults would have broad PfEMP1 serorecognition and high seroreactivity levels during follow-up, particularly to non-CD36-binding PfEMP1s such as those that attach to endothelial protein C receptor (EPCR) and intercellular adhesion molecule-1 (ICAM-1). Using a protein microarray, we determined serologic responses to 166 reference PfEMP1 fragments during a dry and subsequent malaria transmission season in Malian adults. Malian adult sera had PfEMP1 serologic responses throughout the year, with decreased reactivity to a small subset of PfEMP1 fragments during the dry season and increases in reactivity to a different subset of PfEMP1 fragments during the subsequent peak malaria transmission season, especially for intracellular PfEMP1 domains. For some individuals, PfEMP1 serologic responses increased after the dry season, suggesting antigenic switching during asymptomatic infection. Adults were more likely to experience variable serorecognition of CD36-binding PfEMP1s than non-CD36-binding PfEMP1s that bind EPCR or ICAM-1, which remained serorecognized throughout the year. Sustained seroreactivity to non-CD36-binding PfEMP1s throughout adulthood amid seasonal fluctuation patterns may reflect underlying protective severe malaria immunity and merits further investigation.

High-throughput detection of eukaryotic parasites and arboviruses in mosquitoes.

Vector-borne pathogens cause many human infectious diseases and are responsible for high mortality and morbidity throughout the world. They can also cause livestock epidemics with dramatic social and economic consequences. Due to its high costs, vector-borne disease surveillance is often limited to current threats, and the investigation of emerging pathogens typically occurs after the reports of clinical cases. Here, we use high-throughput sequencing to detect and identify a wide range of parasites and viruses carried by mosquitoes from Cambodia, Guinea, Mali and the USA. We apply this approach to individual Anopheles mosquitoes as well as pools of mosquitoes captured in traps; and compare the outcomes of this assay when applied to DNA or RNA. We identified known human and animal pathogens and mosquito parasites belonging to a wide range of taxa, as well as DNA sequences from previously uncharacterized organisms. Our results also revealed that analysis of the content of an entire trap could be an efficient approach to monitor and identify rare vector-borne pathogens in large surveillance studies. Overall, we describe a high-throughput and easy-to-customize assay to screen for a wide range of pathogens and efficiently complement current vector-borne disease surveillance approaches.

A Decline and Age Shift in Malaria Incidence in Rural Mali following Implementation of Seasonal Malaria Chemoprevention and Indoor Residual Spraying.

Many African countries have reported declines in malaria incidence, attributed to the implementation of control strategies. In Mali, artemisinin-based combination therapy (ACT) was introduced in 2004, and long-lasting insecticide-treated nets (LLINs) have been partially distributed free of charge since 2007. In the Malian town of Bandiagara, a study conducted from 2009 to 2013 showed a stable incidence of malaria compared with 1999, despite the implementation of ACTs and LLINs. Since 2016, seasonal malaria chemoprevention has been scaled up across the country. In addition to these strategies, the population of Bandiagara benefited from indoor residual spray implementation in 2017 and 2018 and continued universal bed net coverage. This study aimed to measure the incidence of malaria in Bandiagara, given this recent scaling up of control strategies. A cohort of 300 children aged 6 months to 15 years was followed up from October 2017 to December 2018. We performed monthly cross-sectional surveys to measure anemia and the prevalence of malaria infection by microscopy. The overall incidence of symptomatic malaria was 0.5 episodes/person-year. Malaria incidence in children up to 5 years old significantly declined since 2012 and since 1999 (incidence rate ratio estimates: 6.7 [95% CI: 4.2-11.4] and 13.5 [95% CI: 8.4-22.7]), respectively. The average prevalence of malaria parasitemia was 6.7%. Malaria incidence was higher in children older than 5 years than in those younger than 5 years, highlighting the need to extend malaria control efforts to these older children.