Memory CD8 + T cell compartment associated with delayed onset of Plasmodium falciparum infection and better parasite control in sickle‐cell trait children

Study of individuals with protection from Plasmodium falciparum (Pf) infection and clinical malaria, including individuals affected by the sickle-cell trait (HbAS), offers the potential to identify cellular targets that could be translated for therapeutic development. We previously reported the first involvement of cellular immunity in HbAS-associated relative protection and identified a novel subset of memory-activated NK cells that was enriched in HbAS children and associated with parasite control. We hypothesised that other memory cell subsets might distinguish the baseline profile of HbAS children and children with normal haemoglobin (HbAA). Subsets of memory T cells and NK cells were analysed by flow cytometry in paired samples collected from HbAS and HbAA children, at baseline and during the first malaria episode of the ensuing transmission season. Correlations between cell frequencies and features of HbAS-mediated protection from malaria were determined. HbAS children displayed significantly higher frequency of memory CD8+ T cells at baseline than HbAA children. Baseline frequency of memory CD8+ T cells correlated with features of HbAS-mediated protection from malaria. Exploration of memory CD8+ T cell subsets revealed that central memory CD8+ T cell frequency was higher in HbAS children than in HbAA children. This study shows that HbAS children develop a larger memory CD8+ T cell compartment than HbAA children, and associates this compartment with better control of subsequent onset of infection and parasite density. Our data suggest that central memory CD8+ T cells may play an important role in the relative protection against malaria experienced by HbAS individuals, and further work to investigate this is warranted

The Plasmodium falciparum-Specific Human Memory B Cell Compartment Expands Gradually with Repeated Malaria Infections

Immunity to Plasmodium falciparum (Pf) malaria is only acquired after years of repeated infections and wanes rapidly without ongoing parasite exposure. Antibodies are central to malaria immunity, yet little is known about the B-cell biology that underlies the inefficient acquisition of Pf-specific humoral immunity. This year-long prospective study in Mali of 185 individuals aged 2 to 25 years shows that Pf-specific memory B-cells and antibodies are acquired gradually in a stepwise fashion over years of repeated Pf exposure. Both Pf-specific memory B cells and antibody titers increased after acute malaria and then, after six months of decreased Pf exposure, contracted to a point slightly higher than pre-infection levels. This inefficient, stepwise expansion of both the Pf-specific memory B-cell and long-lived antibody compartments depends on Pf exposure rather than age, based on the comparator response to tetanus vaccination that was efficient and stable. These observations lend new insights into the cellular basis of the delayed acquisition of malaria immunity

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 dhesiveness of infected erythrocytes leads to a rapid increase in parasite burden, promoting higher parasitaemia and increased disease severity

Natural malaria infection elicits rare but potent neutralizing antibodies to the blood-stage antigen RH5

Plasmodium falciparum reticulocyte-binding protein homolog 5 (RH5) is the most advanced blood-stage malaria vaccine candidate and is being evaluated for efficacy in endemic regions, emphasizing the need to study the underlying antibody response to RH5 during natural infection, which could augment or counteract responses to vaccination. Here, we found that RH5-reactive B cells were rare, and circulating immunoglobulin G (IgG) responses to RH5 were short-lived in malaria-exposed Malian individuals, despite repeated infections over multiple years. RH5-specific monoclonal antibodies isolated from eight malaria-exposed individuals mostly targeted non-neutralizing epitopes, in contrast to antibodies isolated from five RH5-vaccinated, malaria-naive UK individuals. However, MAD8-151 and MAD8-502, isolated from two malaria-exposed Malian individuals, were among the most potent neutralizers out of 186 antibodies from both cohorts and targeted the same epitopes as the most potent vaccine-induced antibodies. These results suggest that natural malaria infection may boost RH5-vaccine-induced responses and provide a clear strategy for the development of next-generation RH5 vaccines

Increased circulation time of Plasmodium falciparum underlies persistent asymptomatic infection in the dry season

The dry season is a major challenge for Plasmodium falciparum parasites in many malaria endemic regions, where water availability limits mosquito vectors to only part of the year. How P. falciparum bridges two transmission seasons months apart, without being cleared by the human host or compromising host survival, is poorly understood. Here we show that low levels of P. falciparum parasites persist in the blood of asymptomatic Malian individuals during the 5- to 6-month dry season, rarely causing symptoms and minimally affecting the host immune response. Parasites isolated during the dry season are transcriptionally distinct from those of individuals with febrile malaria in the transmission season, coinciding with longer circulation within each replicative cycle of parasitized erythrocytes without adhering to the vascular endothelium. Low parasite levels during the dry season are not due to impaired replication but rather to increased splenic clearance of longer-circulating infected erythrocytes, which likely maintain parasitemias below clinical and immunological radar. We propose that P. falciparum virulence in areas of seasonal malaria transmission is regulated so that the parasite decreases its endothelial binding capacity, allowing increased splenic clearance and enabling several months of subclinical parasite persistence

B cell analysis of ethnic groups in Mali with differential susceptibility to malaria

<p>Abstract</p> <p>Background</p> <p>Several studies indicate that people of the Fulani ethnic group are less susceptible to malaria compared to those of other ethnic groups living sympatrically in Africa, including the Dogon ethnic group. Although the mechanisms of this protection remain unclear, the Fulani are known to have higher levels of <it>Plasmodium falciparum</it>-specific antibodies of all Ig classes as compared to the Dogon. However, the proportions of B cell subsets in the Fulani and Dogon that may account for differences in the levels of Ig have not been characterized.</p> <p>Methods</p> <p>In this cross-sectional study, venous blood was collected from asymptomatic Fulani (n = 25) and Dogon (n = 25) adults in Mali during the malaria season, and from <it>P. falciparum</it>-naïve adults in the U.S. (n = 8). At the time of the blood collection, <it>P. falciparum</it> infection was detected by blood-smear in 16% of the Fulani and 36% of the Dogon volunteers. Thawed lymphocytes were analysed by flow cytometry to quantify B cell subsets, including immature and naïve B cells; plasma cells; and classical, activated, and atypical memory B cells (MBCs).</p> <p>Results</p> <p>The overall distribution of B cell subsets was similar between Fulani and Dogon adults, although the percentage of activated MBCs was higher in the Fulani group (Fulani: 11.07% [95% CI: 9.317 – 12.82]; Dogon: 8.31% [95% CI: 6.378 – 10.23]; P = 0.016). The percentage of atypical MBCs was similar between Fulani and Dogon adults (Fulani: 28.3% [95% CI: 22.73 – 34.88]; Dogon: 29.3% [95% CI: 25.06 – 33.55], but higher than U.S. adults (U.S.: 3.0% [95% CI: -0.21 - 6.164]; P < 0.001). <it>Plasmodium falciparum</it> infection was associated with a higher percentage of plasma cells among Fulani (Fulani infected: 3.3% [95% CI: 1.788 – 4.744]; Fulani uninfected: 1.71% [95% CI: 1.33 – 2.08]; P = 0.011), but not Dogon adults.</p> <p>Conclusion</p> <p>These data show that the malaria-resistant Fulani have a higher percentage of activated MBCs compared to the Dogon, and that <it>P. falciparum</it> infection is associated with a higher percentage of plasma cells in the Fulani compared to the Dogon, findings that may account for the higher levels of <it>P. falciparum</it> antibodies in the Fulani.</p

Susceptibility to febrile malaria is associated with an inflammatory gut microbiome

Malaria is a major public health problem, but many of the factors underlying the pathogenesis of this disease are not well understood. Here, we demonstrate in Malian children that susceptibility to febrile malaria following infection with Plasmodium falciparum is associated with the composition of the gut microbiome prior to the malaria season. Gnotobiotic mice colonized with the fecal samples of malaria-susceptible children had a significantly higher parasite burden following Plasmodium infection compared to gnotobiotic mice colonized with the fecal samples of malaria-resistant children. The fecal microbiome of the susceptible children was enriched for bacteria associated with inflammation, mucin degradation, gut permeability and inflammatory bowel disorders (e.g., Ruminococcus gauvreauii, Ruminococcus torques, Dorea formicigenerans, Dorea longicatena, Lachnoclostridium phocaeense and Lachnoclostridium sp. YL32). However, the susceptible children also had a greater abundance of bacteria known to produce anti-inflammatory short-chain fatty acids and those associated with favorable prognosis and remission following dysbiotic intestinal events (e.g., Anaerobutyricum hallii, Blautia producta and Sellimonas intestinalis). Metabolomics analysis of the human fecal samples corroborated the existence of inflammatory and recovery-associated features within the gut microbiome of the susceptible children. There was an enrichment of nitric oxide-derived DNA adducts (deoxyinosine and deoxyuridine) and long-chain fatty acids, the absorption of which has been shown to be inhibited by inflamed intestinal epithelial cells, and a decrease in the abundance of mucus phospholipids. Nevertheless, there were also increased levels of pseudouridine and hypoxanthine, which have been shown to be regulated in response to cellular stress and to promote recovery following injury or hypoxia. Overall, these results indicate that the gut microbiome may contribute malaria pathogenesis and suggest that therapies targeting intestinal inflammation could decrease malaria susceptibility

In Vitro Growth-Inhibitory Activity and Malaria Risk in a Cohort Study in Mali▿ †

Immunity to the asexual blood stage of Plasmodium falciparum is complex and likely involves several effector mechanisms. Antibodies are thought to play a critical role in malaria immunity, and a corresponding in vitro correlate of antibody-mediated immunity has long been sought to facilitate malaria vaccine development. The growth inhibition assay (GIA) measures the capacity of antibodies to limit red blood cell (RBC) invasion and/or growth of P. falciparum in vitro. In humans, naturally acquired and vaccine-induced P. falciparum-specific antibodies have growth-inhibitory activity, but it is unclear if growth-inhibitory activity correlates with protection from clinical disease. In a longitudinal study in Mali, purified IgGs, obtained from plasmas collected before the malaria season from 220 individuals aged 2 to 10 and 18 to 25 years, were assayed for growth-inhibitory activity. Malaria episodes were recorded by passive surveillance over the subsequent 6-month malaria season. Logistic regression showed that greater age (odds ratio [OR], 0.78; 95% confidence interval [95% CI], 0.63 to 0.95; P = 0.02) and growth-inhibitory activity (OR, 0.50; 95% CI, 0.30 to 0.85; P = 0.01) were significantly associated with decreased malaria risk in children. A growth-inhibitory activity level of 40% was determined to be the optimal cutoff for discriminating malaria-immune and susceptible individuals in this cohort, with a sensitivity of 97.0%, but a low specificity of 24.3%, which limited the assay's ability to accurately predict protective immunity and to serve as an in vitro correlate of antibody-mediated immunity. These data suggest that antibodies which block merozoite invasion of RBC and/or inhibit the intra-RBC growth of the parasite contribute to but are not sufficient for the acquisition of malaria immunity

Co-infection of long-term carriers of Plasmodium falciparum with Schistosoma haematobium enhances protection from febrile malaria: a prospective cohort study in Mali.

Malaria and schistosomiasis often overlap in tropical and subtropical countries and impose tremendous disease burdens; however, the extent to which schistosomiasis modifies the risk of febrile malaria remains unclear.We evaluated the effect of baseline S. haematobium mono-infection, baseline P. falciparum mono-infection, and co-infection with both parasites on the risk of febrile malaria in a prospective cohort study of 616 children and adults living in Kalifabougou, Mali. Individuals with S. haematobium were treated with praziquantel within 6 weeks of enrollment. Malaria episodes were detected by weekly physical examination and self-referral for 7 months. The primary outcome was time to first or only malaria episode defined as fever (≥ 37.5 °C) and parasitemia (≥ 2500 asexual parasites/µl). Secondary definitions of malaria using different parasite densities were also explored.After adjusting for age, anemia status, sickle cell trait, distance from home to river, residence within a cluster of high S. haematobium transmission, and housing type, baseline P. falciparum mono-infection (n = 254) and co-infection (n = 39) were significantly associated with protection from febrile malaria by Cox regression (hazard ratios 0.71 and 0.44; P = 0.01 and 0.02; reference group: uninfected at baseline). Baseline S. haematobium mono-infection (n = 23) did not associate with malaria protection in the adjusted analysis, but this may be due to lack of statistical power. Anemia significantly interacted with co-infection (P = 0.009), and the malaria-protective effect of co-infection was strongest in non-anemic individuals. Co-infection was an independent negative predictor of lower parasite density at the first febrile malaria episode.Co-infection with S. haematobium and P. falciparum is significantly associated with reduced risk of febrile malaria in long-term asymptomatic carriers of P. falciparum. Future studies are needed to determine whether co-infection induces immunomodulatory mechanisms that protect against febrile malaria or whether genetic, behavioral, or environmental factors not accounted for here explain these findings