Breadth of Fc-mediated effector function correlates with clinical immunity following human malaria challenge.

Malaria is a life-threatening disease of global health importance, particularly in sub-Saharan Africa. The growth inhibition assay (GIA) is routinely used to evaluate, prioritize, and quantify the efficacy of malaria blood-stage vaccine candidates but does not reliably predict either naturally acquired or vaccine-induced protection. Controlled human malaria challenge studies in semi-immune volunteers provide an unparalleled opportunity to robustly identify mechanistic correlates of protection. We leveraged this platform to undertake a head-to-head comparison of seven functional antibody assays that are relevant to immunity against the erythrocytic merozoite stage of Plasmodium falciparum. Fc-mediated effector functions were strongly associated with protection from clinical symptoms of malaria and exponential parasite multiplication, while the gold standard GIA was not. The breadth of Fc-mediated effector function discriminated clinical immunity following the challenge. These findings present a shift in the understanding of the mechanisms that underpin immunity to malaria and have important implications for vaccine development.

A retrospective analysis of P. falciparum drug resistance markers detects an early (2016/17) high prevalence of the k13 C469Y mutation in asymptomatic infections in Northern Uganda.

The emergence of drug-resistant Plasmodium falciparum parasites in sub-Saharan Africa will substantially challenge malaria control. Here, we evaluated the frequency of common drug resistance markers among adolescents from Northern Uganda with asymptomatic infections. We used an established amplicon deep sequencing strategy to screen dried blood spot samples collected from 2016 to 2017 during a reported malaria epidemic within the districts of Kitgum and Pader in Northern Uganda. We screened single-nucleotide polymorphisms within: kelch13 (Pfk13), dihydropteroate synthase (Pfdhps), multidrug resistance-1 (Pfmdr1), dihydrofolate reductase (Pfdhfr), and apical membrane antigen (Pfama1) genes. Within the study population, the median age was 15 years (14.3-15.0, 95% CI), and 54.9% (78/142) were Plasmodium positive by 18S rRNA qPCR, which were subsequently targeted for sequencing analysis. We observed a high frequency of resistance markers particularly for sulfadoxine-pyrimethamine (SP), with no wild-type-only parasites observed for Pfdhfr (N51I, C59R, and S108N) and Pfdhps (A437G and K540E) mutations. Within Pfmdr1, mixed infections were common for NF/NY (98.5%). While for artemisinin resistance, in kelch13, there was a high frequency of C469Y (34%). Using the pattern for Pfama1, we found a high level of polygenomic infections with all individuals presenting with complexity of infection greater than 2 with a median of 6.9. The high frequency of the quintuple SP drug-resistant parasites and the C469Y artemisinin resistance-associated mutation in asymptomatic individuals suggests an earlier high prevalence than previously reported from symptomatic malaria surveillance studies (in 2016/2017). Our data demonstrate the urgency for routine genomic surveillance programs throughout Africa and the value of deep sequencing.

Bi-isotype immunoglobulins enhance antibody-mediated neutrophil activity against Plasmodium falciparum parasites.

Background: Malaria remains a major global health priority, and monoclonal antibodies (mAbs) are emerging as potential new tools to support efforts to control the disease. Recent data suggest that Fc-dependent mechanisms of immunity are important mediators of protection against the blood stages of the infection, but few studies have investigated this in the context of mAbs. We aimed to isolate mAbs agnostic to cognate antigens that target whole merozoites and simultaneously induce potent neutrophil activity measured by the level of reactive oxygen species (ROS) production using an antibody-dependent respiratory burst (ADRB) assay. Methods: We used samples from semi-immune adults living in coastal Kenya to isolate mAbs that induce merozoite-specific ADRB activity. We then tested whether modifying the expressed IgG1 isotype to an IgG-IgA Fc region chimera would enhance the level of ADRB activity. Results: We isolated a panel of nine mAbs with specificity to whole merozoites. mAb J31 induced ADRB activity in a dose-dependent fashion. Compared to IgG1, our modified antibody IgG-IgA bi-isotype induced higher ADRB activity across all concentrations tested. Further, we observed a negative hook effect at high IgG1 mAb concentrations (i.e., >200 µg/mL), but this was reversed by Fc modification. We identified MSP3.5 as the potential cognate target of mAb J31. Conclusions: We demonstrate an approach to engineer mAbs with enhanced ADRB potency against blood-stage parasites.

Antibody-Dependent Respiratory Burst against Plasmodium falciparum Merozoites in Individuals Living in an Area with Declining Malaria Transmission.

Malaria transmission intensity affects the development of naturally acquired immunity to malaria. An absolute correlate measure of protection against malaria is lacking. However, antibody-mediated functions against Plasmodium falciparum correlate with protection against malaria. In children, antibody-mediated functions against P. falciparum decline with reduced exposure. It is unclear whether adults maintain antibody-mediated functions as malaria transmission declines. This study assessed antibody-dependent respiratory burst (ADRB) in individuals from an area with declining malaria transmission. In an age-matched analysis, we compare ADRB activity during high versus low malaria transmission periods. Age significantly predicted higher ADRB activity in the high (p < 0.001) and low (p < 0.001) malaria transmission periods. ADRB activity was higher during the high compared to the low malaria transmission period in older children and adults. Only older adults during the high malaria transmission period had their median ADRB activity above the ADRB cut-off. Ongoing P. falciparum infection influenced ADRB activity during the low (p = 0.01) but not the high (p = 0.29) malaria transmission period. These findings propose that naturally acquired immunity to P. falciparum is affected in children and adults as malaria transmission declines, implying that vaccines will be necessary to induce and maintain protection against malaria.

Full-length MSP1 is a major target of protective immunity after controlled human malaria infection.

The merozoite surface protein 1 (MSP1) is the most abundant protein on the surface of the invasive merozoite stages of Plasmodium falciparum and has long been considered a key target of protective immunity. We used samples from a single controlled human malaria challenge study to test whether the full-length version of MSP1 (MSP1FL) induced antibodies that mediated Fc-IgG functional activity in five independent assays. We found that anti-MSP1FL antibodies induced complement fixation via C1q, monocyte-mediated phagocytosis, neutrophil respiratory burst, and natural killer cell degranulation as well as IFNγ production. Activity in each of these assays was strongly associated with protection. The breadth of MSP1-specific Fc-mediated effector functions was more strongly associated with protection than the individual measures and closely mirrored what we have previously reported using the same assays against merozoites. Our findings suggest that MSP1FL is an important target of functional antibodies that contribute to a protective immune response against malaria.