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Anti‐severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) seroprevalence is used to estimate the proportion of individuals within a population previously infected, to track viral transmission, and to monitor naturally and vaccine‐induced immune protection. However, in sub‐Saharan African settings, antibodies induced by higher exposure to pathogens may increase unspecific seroreactivity to SARS‐CoV‐2 antigens, resulting in false positive responses. To investigate the level and type of unspecific seroreactivitiy to SARS‐CoV‐2 in Africa, we measured immunoglobulin G (IgG), IgA, and IgM to a broad panel of antigens from different pathogens by Luminex in 602 plasma samples from African and European subjects differing in coronavirus disease 2019, malaria, and other exposures. Seroreactivity to SARS‐CoV‐2 antigens was higher in prepandemic African than in European samples and positively correlated with antibodies against human coronaviruses, helminths, protozoa, and especially Plasmodium falciparum. African subjects presented higher levels of autoantibodies, a surrogate of polyreactivity, which correlated with P. falciparum and SARS‐CoV‐2 antibodies. Finally, we found an improved sensitivity in the IgG assay in African samples when using urea as a chaotropic agent. In conclusion, our data suggest that polyreactive antibodies induced mostly by malaria are important mediators of the unspecific anti‐SARS‐CoV‐2 responses, and that the use of dissociating agents in immunoassays could be useful for more accurate estimates of SARS‐CoV‐2 seroprevalence in African settings. [ABSTRACT FROM AUTHOR]

Abstract Background Assessment of malaria endemicity at different altitudes and transmission intensities, in the era of dwindling vector densities in the highlands, will provide valuable information for malaria control and surveillance. Measurement of serum anti-malarial antibodies is a useful marker of malaria exposure that indicates long-term transmission potential. We studied the serologic evidence of malaria endemicity at two highland sites along a transmission intensity cline. An improved understanding of the micro-geographic variation in malaria exposure in the highland ecosystems will be relevant in planning effective malaria control. Methods Total IgG levels to Plasmodium falciparum MSP-119 were measured in an age-stratified cohort (< 5, 5-14 and ≥ 15 years) in 795 participants from an uphill and valley bottom residents during low and high malaria transmission seasons. Antibody prevalence and level was compared between different localities. Regression analysis was performed to examine the association between antibody prevalence and parasite prevalence. Age-specific MSP-119 seroprevalence data was fitted to a simple reversible catalytic model to investigate the relationship between parasite exposure and age. Results Higher MSP-119 seroprevalence and density were observed in the valley residents than in the uphill dwellers. Adults (> 15 years) recorded high and stable immune response in spite of changing seasons. Lower responses were observed in children (≤ 15 years), which, fluctuated with changing seasons particularly in the valley residents. In the uphill population, annual seroconversion rate (SCR) was 8.3% and reversion rate was 3.0%, with seroprevalence reaching a plateau of 73.3% by age of 20. Contrary, in the valley bottom population, the annual SCR was 35.8% and the annual seroreversion rate was 3.5%, and seroprevalence in the population had reached 91.2% by age 10. Conclusion The study reveals the micro-geographic variation in malaria endemicity in the highland eco-system; this validates the usefulness of sero-epidemiological tools in assessing malaria endemicity in the era of decreasing sensitivity of conventional tools.

Abstract Background The development of an asexual blood stage vaccine against Plasmodium falciparum malaria based on the major merozoite surface protein-1 (MSP1) antigen is founded on the protective efficacy observed in preclinical studies and induction of invasion and growth inhibitory antibody responses. The 42 kDa C-terminus of MSP1 has been developed as the recombinant protein vaccine antigen, and the 3D7 allotype, formulated with the Adjuvant System AS02A, has been evaluated extensively in human clinical trials. In preclinical rabbit studies, the FVO allele of MSP142 has been shown to have improved immunogenicity over the 3D7 allele, in terms of antibody titres as well as growth inhibitory activity of antibodies against both the heterologous 3D7 and homologous FVO parasites. Methods Two Phase 1 clinical studies were conducted to examine the safety, reactogenicity and immunogenicity of the FVO allele of MSP142 in the adjuvant system AS01 administered intramuscularly at 0-, 1-, and 2-months: one in the USA and, after evaluation of safety data results, one in Western Kenya. The US study was an open-label, dose escalation study of 10 and 50 μg doses of MSP142 in 26 adults, while the Kenya study, evaluating 30 volunteers, was a double-blind, randomized study of only the 50 μg dose with a rabies vaccine comparator. Results In these studies it was demonstrated that this vaccine formulation has an acceptable safety profile and is immunogenic in malaria-naïve and malaria-experienced populations. High titres of anti-MSP1 antibodies were induced in both study populations, although there was a limited number of volunteers whose serum demonstrated significant inhibition of blood-stage parasites as measured by growth inhibition assay. In the US volunteers, the antibodies generated exhibited better cross-reactivity to heterologous MSP1 alleles than a MSP1-based vaccine (3D7 allele) previously tested at both study sites. Conclusions Given that the primary effector mechanism for blood stage vaccine targets is humoral, the antibody responses demonstrated to this vaccine candidate, both quantitative (total antibody titres) and qualitative (functional antibodies inhibiting parasite growth) warrant further consideration of its application in endemic settings. Trial registrations Clinical Trials NCT00666380

Abstract Background MSP1 is the major surface protein on merozoites and a prime candidate for a blood stage malaria vaccine. Preclinical and seroepidemiological studies have implicated antibodies to MSP1 in protection against blood stage parasitaemia and/or reduced parasite densities, respectively. Malaria endemic areas have multiple strains of Plasmodium falciparum circulating at any given time, giving rise to complex immune responses, an issue which is generally not addressed in clinical trials conducted in non-endemic areas. A lack of understanding of the effect of pre-existing immunity to heterologous parasite strains may significantly contribute to vaccine failure in the field. The purpose of this study was to model the effect of pre-existing immunity to MSP142 on the immunogenicity of blood-stage malaria vaccines based on alternative MSP1 alleles. Methods Inbred and outbred mice were immunized with various recombinant P. falciparum MSP142 proteins that represent the two major alleles of MSP142, MAD20 (3D7) and Wellcome (K1, FVO). Humoral immune responses were analysed by ELISA and LuminexTM, and functional activity of induced MSP142-specific antibodies was assessed by growth inhibition assays. T-cell responses were characterized using ex vivo ELISpot assays. Results Analysis of the immune responses induced by various immunization regimens demonstrated a strong allele-specific response at the T cell level in both inbred and outbred mice. The success of heterologous regimens depended on the degree of homology of the N-terminal p33 portion of the MSP142, likely due to the fact that most T cell epitopes reside in this part of the molecule. Analysis of humoral immune responses revealed a marked cross-reactivity between the alleles. Functional analyses showed that some of the heterologous regimens induced antibodies with improved growth inhibitory activities. Conclusion The development of a more broadly efficacious MSP1 based vaccine may be hindered by clonally imprinted p33 responses mainly restricted at the T cell level. In this study, the homology of the p33 sequence between the clonally imprinted response and the vaccine allele determines the magnitude of vaccine induced responses.

Abstract Background The immunological mechanisms responsible for protection against malaria infection vary among Plasmodium species, host species and the developmental stage of parasite, and are poorly understood. A challenge with live parasites is the most relevant approach to testing the efficacy of experimental malaria vaccines. Nevertheless, in the mouse models of Plasmodium berghei and Plasmodium yoelii, parasites are usually delivered by intravenous injection. This route is highly artificial and particularly in the P. berghei model produces inconsistent challenge results. The initial objective of this study was to compare an optimized intravenous (IV) delivery challenge model with an optimized single infectious mosquito bite challenge model. Finding shortcomings of both approaches, an alternative approach was explored, i.e., the subcutaneous challenge. Methods Mice were infected with P. berghei sporozoites by intravenous (tail vein) injection, single mosquito bite, or subcutaneous injection of isolated parasites into the subcutaneous pouch at the base of the hind leg. Infection was determined in blood smears 7 and 14 days later. To determine the usefulness of challenge models for vaccine testing, mice were immunized with circumsporozoite-based DNA vaccines by gene gun. Results Despite modifications that allowed infection with a much smaller than reported number of parasites, the IV challenge remained insufficiently reliable and reproducible. Variations in the virulence of the inoculum, if not properly monitored by the rigorous inclusion of sporozoite titration curves in each experiment, can lead to unacceptable variations in reported vaccine efficacies. In contrast, mice with different genetic backgrounds were consistently infected by a single mosquito bite, without overwhelming vaccine-induced protective immune responses. Because of the logistical challenges associated with the mosquito bite model, the subcutaneous challenge route was optimized. This approach, too, yields reliable challenge results, albeit requiring a relatively large inoculum. Conclusions Although a single bite by P. berghei infected Anopheles mosquitoes was superior to the IV challenge route, it is laborious. However, any conclusive evaluation of a pre-erythrocytic malaria vaccine candidate should require challenge through the natural anatomic target site of the parasite, the skin. The subcutaneous injection of isolated parasites represents an attractive compromise. Similar to the mosquito bite model, it allows vaccine-induced antibodies to exert their effect and is, therefore not as prone to the artifacts of the IV challenge.

Abstract Background Antibodies are the main effector molecules in the defense against blood stages of the malaria parasite Plasmodium falciparum. Understanding the mechanisms by which vaccine-induced anti-blood stage antibodies work in protecting against malaria is essential for vaccine design and testing. Methods The effects of MSP-1p42-specific antibodies on the development of blood stage parasites were studied using microscopy, flow cytometry and the pLDH assay. To determine allele-specific effects, if present, allele-specific antibodies and the various parasite clones representative of these alleles of MSP-1 were employed. Results The mode of action of anti-MSP-1p42 antibodies differs among the parasite clones tested: anti-MSP-1p42 sera act mainly through invasion-inhibitory mechanisms against FVO parasites, by either preventing schizonts from rupturing or agglutinating merozoites upon their release. The same antibodies do not prevent the rupture of 3D7 schizonts; instead they agglutinate merozoites and arrest the development of young parasites at the early trophozoite stage, thus acting through both invasion- and growth inhibitory mechanisms. The second key finding is that antibodies have access to the intra-erythrocytic parasite, as evidenced by the labeling of developing merozoites with fluorochrome-conjugated anti-MSP-1p42 antibodies. Access to the parasite through this route likely allows antibodies to exert their inhibitory activities on the maturing schizonts leading to their inability to rupture and be released as infectious merozoites. Conclusion The identification of various modes of action by which anti-MSP-1 antibodies function against the parasite during erythrocytic development emphasizes the importance of functional assays for evaluating malaria vaccines and may also open new avenues for immunotherapy and vaccine development.

Abstract Background Antibodies are the main effectors against malaria blood-stage parasites. Evaluation of functional activities in immune sera from Phase 2a/b vaccine trials may provide invaluable information in the search for immune correlates of protection. However, the presence of anti-malarial-drugs, improper collection/storage conditions or concomitant immune responses against other pathogens can contribute to non-specific anti-parasite activities when the sera/plasma are tested in vitro. Purification of immunoglobulin is a standard approach for reducing such non-specific background activities, but the purification method itself can alter the quality and yield of recovered Ag-specific antibodies. Methods To address this concern, various immunoglobulin (Ig) purification methods (protein G Sepharose, protein A/G Sepharose, polyethylene glycol and caprylic acid-ammonium sulphate precipitation) were evaluated for their impact on the quality, quantity and functional activity of purified rabbit and human Igs. The recovered Igs were analysed for yield and purity by SDS-PAGE, for quality by Ag-specific ELISAs (determining changes in titer, avidity and isotype distribution) and for functional activity by in vitro parasite growth inhibition assay (GIA). Results This comparison demonstrated that overall polyethylene glycol purification of human serum/plasma samples and protein G Sepharose purification of rabbit sera are optimal for recovering functional Ag-specific antibodies. Conclusion Consequently, critical consideration of the purification method is required to avoid selecting non-representative populations of recovered Ig, which could influence interpretations of vaccine efficacy, or affect the search for immune correlates of protection.

Transplacental transfer of antibodies is essential for conferring protection in newborns against infectious diseases. We assessed the impact of different factors, including gestational age and maternal infections such as HIV and malaria, on the efficiency of cord blood levels and placental transfer of IgG subclasses. We measured total IgG and IgG subclasses by quantitative suspension array technology against 14 pathogens and vaccine antigens, including targets of maternal immunization, in 341 delivering HIV-uninfected and HIV-infected mother-infant pairs from southern Mozambique. We analyzed the association of maternal HIV infection, Plasmodium falciparum exposure, maternal variables and pregnancy outcomes on cord antibody levels and transplacental transfer. Our results show that maternal antibody levels were the main determinant of cord antibody levels. Univariable and multivariable analysis showed that HIV reduced the placental transfer and cord levels of IgG and IgG1 principally, but also IgG2 to half of the antigens tested. P. falciparum exposure and prematurity were negatively associated with cord antibody levels and placental transfer, but this was antigen-subclass dependent. Our findings suggest that lower maternally transferred antibodies may underlie increased susceptibility to infections of HIV-exposed infants. This could affect efficacy of maternal vaccination, especially in sub-Saharan Africa, where there is a high prevalence of HIV, malaria and unfavorable environmental factors. [ABSTRACT FROM AUTHOR]

Background: The impact of preexisting immunity on the efficacy of artemisinin combination therapy must be examined to monitor resistance, and for implementation of new treatment strategies.Methods: Serum samples obtained from a clinical trial in Western Kenya randomized to receive artemether-lumefantrine (AL) or artesunate-mefloquine (ASMQ) were screened for total immunoglobulin G against preerythrocytic and erythrocytic antigens. The association and correlation between different variables, and impact of preexisting immunity on parasite slope half-life (t½) was determined.Results: There was no significant difference in t½, but the number of individuals with lag phase was significantly higher in the AL than in the ASMQ arm (29 vs 13, respectively; P < .01). Circumsporozoite protein-specific antibodies correlate positively with t½ (AL, P = .03; ASMQ, P = .09), but negatively with clearance rate in both study arms (AL, P = .16; ASMQ, P = .02). The t½ correlated negatively with age in ASMQ group. When stratified based on t½, the antibody titers against circumsporozoite protein and merozoite surface protein 1 were significantly higher in participants who cleared parasites rapidly in the AL group (P = .01 and P = .02, respectively).Conclusion: Data presented here define immunoprofiles associated with distinct responses to 2 different antimalarial drugs, revealing impact of preexisting immunity on the efficacy of artemisinin combination therapy regimens in a malaria-holoendemic area.Clinical Trials Registration: NCT01976780. [ABSTRACT FROM AUTHOR]

Naturally acquired immunity (NAI) to Plasmodium falciparum malaria is mainly mediated by IgG antibodies but the subclasses, epitope targets and effector functions have not been unequivocally defined. Dissecting the type and specificity of antibody responses mediating NAI is a key step toward developing more effective vaccines to control the disease. We investigated the role of IgG subclasses to malaria antigens in protection against disease and the factors that affect their levels, including vaccination with RTS,S/AS01E. We analyzed plasma and serum samples at baseline and 1 month after primary vaccination with RTS,S or comparator in African children and infants participating in a phase 3 trial in two sites of different malaria transmission intensity: Kintampo in Ghana and Manhiça in Mozambique. We used quantitative suspension array technology (qSAT) to measure IgG1−4 responses to 35 P. falciparum pre-erythrocytic and blood stage antigens. Our results show that the pattern of IgG response is predominantly IgG1 or IgG3, with lower levels of IgG2 and IgG4. Age, site and RTS,S vaccination significantly affected antibody subclass levels to different antigens and susceptibility to clinical malaria. Univariable and multivariable analysis showed associations with protection mainly for cytophilic IgG3 levels to selected antigens, followed by IgG1 levels and, unexpectedly, also with IgG4 levels, mainly to antigens that increased upon RTS,S vaccination such as MSP5 and MSP1 block 2, among others. In contrast, IgG2 was associated with malaria risk. Stratified analysis in RTS,S vaccinees pointed to novel associations of IgG4 responses with immunity mainly involving pre-erythrocytic antigens upon RTS,S vaccination. Multi-marker analysis revealed a significant contribution of IgG3 responses to malaria protection and IgG2 responses to malaria risk. We propose that the pattern of cytophilic and non-cytophilic IgG antibodies is antigen-dependent and more complex than initially thought, and that mechanisms of both types of subclasses could be involved in protection. Our data also suggests that RTS,S efficacy is significantly affected by NAI, and indicates that RTS,S vaccination significantly alters NAI. [ABSTRACT FROM AUTHOR]

Reducing variability of quantitative suspension array assays is key for multi-center and large sero-epidemiological studies. To maximize precision and robustness of an in-house IgG multiplex assay, we analyzed the effect of several conditions on variability to find the best combination. The following assay conditions were studied through a fractional factorial design: antigen-bead coupling (stock vs. several), sample predilution (stock vs. daily), temperature of incubation of sample with antigen-bead (22°C vs. 37°C), plate washing (manual vs. automatic) and operator expertise (expert vs. apprentice). IgG levels against seven P. falciparum antigens with heterogeneous immunogenicities were measured in test samples, in a positive control and in blanks. We assessed the variability and MFI quantification range associated to each combination of conditions, and their interactions, and evaluated the minimum number of samples and blank replicates to achieve good replicability. Results showed that antigen immunogenicity and sample seroreactivity defined the optimal dilution to assess the effect of assay conditions on variability. We found that a unique antigen-bead coupling, samples prediluted daily, incubation at 22°C, and automatic washing, had lower variability. However, variability increased when performing several couplings and incubating at 22°C vs. 37°C. In addition, no effect of temperature was seen with a unique coupling. The expertise of the operator had no effect on assay variability but reduced the MFI quantification range. Finally, differences between sample replicates were minimal, and two blanks were sufficient to capture assay variability, as suggested by the constant Intraclass Correlation Coefficient of three and two blanks. To conclude, a single coupling was the variable that most consistently reduced assay variability, being clearly advisable. In addition, we suggest having more sample dilutions instead of replicates to increase the likelihood of sample MFIs falling in the linear part of the antigen-specific curve, thus increasing precision. [ABSTRACT FROM AUTHOR]

Epstein-Barr virus (EBV) is a necessary cause of endemic Burkitt lymphoma (eBL), while the role of Plasmodium falciparum in eBL remains uncertain. This study aimed to generate new hypotheses on the interplay between both infections in the development of eBL by investigating the IgG and IgM profiles against several EBV and P. falciparum antigens. Serum samples collected in a childhood study in Malawi (2005-2006) from 442 HIV-seronegative children (271 eBL cases and 171 controls) between 1.4 and 15 years old were tested by quantitative suspension array technology against a newly developed multiplex panel combining 4 EBV antigens [Z Epstein-Barr replication activator protein (ZEBRA), early antigen-diffuse component (EA-D), EBV nuclear antigen 1, and viral capsid antigen p18 subunit (VCA-p18)] and 15 P. falciparum antigens selected for their immunogenicity, role in malaria pathogenesis, and presence in different parasite stages. Principal component analyses, multivariate logistic models, and elastic-net regressions were used. As expected, elevated levels of EBV IgG (especially against the lytic antigens ZEBRA, EA-D, and VCA-p18) were strongly associated with eBL [high vs low tertile odds ratio (OR) = 8.67, 95% confidence interval (CI) = 4.81-15.64]. Higher IgG responses to the merozoite surface protein 3 were observed in children with eBL compared with controls (OR = 1.29, 95% CI = 1.02-1.64), showing an additive interaction with EBV IgGs (OR = 10.6, 95% CI = 5.1-22.2, P = 0.05). Using elastic-net regression models, eBL serological profile was further characterized by lower IgM levels against P. falciparum preerythrocytic-stage antigen CelTOS and EBV lytic antigen VCA-p18 compared with controls. In a secondary analysis, abdominal Burkitt lymphoma had lower IgM to EBV and higher IgG to EA-D levels than cases with head involvement. Overall, this exploratory study confirmed the strong role of EBV in eBL and identified differential IgG and IgM patterns to erythrocytic vs preerythrocytic P. falciparum antigens that suggest a more persistent/chronic malaria exposure and a weaker IgM immune response in children with eBL compared with controls. Future studies should continue exploring how the malaria infection status and the immune response to P. falciparum interact with EBV infection in the development of eBL. [ABSTRACT FROM AUTHOR]

The blood-stage malaria vaccine FMP2.1/AS02A, comprised of recombinant Plasmodium falciparum apical membrane antigen 1 (AMA1) and the adjuvant system AS02A, had strain-specific efficacy against clinical malaria caused by P. falciparum with the vaccine strain 3D7 AMA1 sequence. To evaluate a potential correlate of protection, we measured the ability of participant sera to inhibit growth of 3D7 and FVO strains in vitro using high-throughput growth inhibition assay (GIA) testing. Sera from 400 children randomized to receive either malaria vaccine or a control rabies vaccine were assessed at baseline and over two annual malaria transmission seasons after immunization. Baseline GIA against vaccine strain 3D7 and FVO strain was similar in both groups, but more children in the malaria vaccine group than in the control group had 3D7 and FVO GIA activity ≥15% 30 days after the last vaccination (day 90) (49% vs. 16%, p<0.0001; and 71.8% vs. 60.4%, p = 0.02). From baseline to day 90, 3D7 GIA in the vaccine group was 7.4 times the mean increase in the control group (p<0.0001). In AMA1 vaccinees, 3D7 GIA activity subsequently returned to baseline one year after vaccination (day 364) and did not correlate with efficacy in the extended efficacy time period to day 730. In Cox proportional hazards regression models with time-varying covariates, there was a slight suggestion of an association between 3D7 GIA activity and increased risk of clinical malaria between day 90 and day 240. We conclude that vaccination with this AMA1-based malaria vaccine increased inhibition of parasite growth, but this increase was not associated with allele-specific efficacy in the first malaria season. These results provide a framework for testing functional immune correlates of protection against clinical malaria in field trials, and will help to guide similar analyses for next-generation malaria vaccines. Clinical trials registry: This clinical trial was registered on clinicaltrials.gov, registry number . [ABSTRACT FROM AUTHOR]

Whole malaria sporozoite vaccine regimens are promising new strategies, and some candidates have demonstrated high rates of durable clinical protection associated with memory T cell responses. Little is known about the anatomical distribution of memory T cells following whole sporozoite vaccines, and immunization of nonhuman primates can be used as a relevant model for humans. We conducted a chemoprophylaxis with sporozoite (CPS) immunization in P. knowlesi rhesus monkeys and challenged via mosquito bites. Half of CPS immunized animals developed complete protection, with a marked delay in parasitemia demonstrated in the other half. Antibody responses to whole sporozoites, CSP, and AMA1, but not CelTOS were detected. Peripheral blood T cell responses to whole sporozoites, but not CSP and AMA1 peptides were observed. Unlike peripheral blood, there was a high frequency of sporozoite-specific memory T cells observed in the liver and bone marrow. Interestingly, sporozoite-specific CD4+ and CD8+ memory T cells in the liver highly expressed chemokine receptors CCR5 and CXCR6, both of which are known for liver sinusoid homing. The majority of liver sporozoite-specific memory T cells expressed CD69, a phenotypic marker of tissue-resident memory (TRM) cells, which are well positioned to rapidly control liver-stage infection. Vaccine strategies that aim to elicit large number of liver TRM cells may efficiently increase the efficacy and durability of response against pre-erythrocytic parasites. [ABSTRACT FROM AUTHOR]

Introduction: As an increasing number of malaria-endemic countries approach the disease elimination phase, sustenance of control efforts and effective monitoring are necessary to ensure success. Mathematical models that estimate anti-parasite antibody seroconversion rates are gaining relevance as more sensitive transmission intensity estimation tools. Models however estimate yearly seroconversion and seroreversion rates and usually predict long term changes in transmission, occurring years before the time of sampling. Another challenge is the identification of appropriate antigen targets since specific antibody levels must directly reflect changes in transmission patterns. We therefore investigated the potential of antibodies to sporozoite and blood stage antigens for detecting short term differences in malaria transmission in two communities in Northern Ghana with marked, seasonal transmission. Methods: Cross-sectional surveys were conducted during the rainy and dry seasons in two communities, one in close proximity to an irrigation dam and the other at least 20 Km away from the dam. Antibodies against the sporozoite-specific antigens circumsporozoite protein (CSP) and Cell traversal for ookinetes and sporozoites (CelTOS) and the classical blood stage antigen apical membrane antigen 1 (AMA1) were measured by indirect ELISA. Antibody levels and seroprevalence were compared between surveys and between study communities. Antibody seroprevalence data were fitted to a modified reversible catalytic model to estimate the seroconversion and seroreversion rates. Results: Changes in sporozoite-specific antibody levels and seroprevalence directly reflected differences in parasite prevalence between the rainy and dry seasons and hence the extent of malaria transmission. Seroconversion rate estimates from modelled seroprevalence data did not however support the above observation. Conclusions: The data confirms the potential utility of sporozoite-specific antigens as useful markers for monitoring short term/seasonal changes in malaria transmission. It may however be essential to update models to allow for assessment of seasonal changes in malaria transmission, which usually occur within four to six months. [ABSTRACT FROM AUTHOR]

Background: In addition to evidence for a protective role of antibodies to the malaria blood stage antigen merozoite surface protein 1 (MSP1), MSP1 antibodies are also considered as a marker of past malaria exposure in sero-epidemiological studies. Methods: In order to better assess the potential use of MSP1 serology in malaria chemoprophylaxis trials in endemic areas, an analysis for the prevalence of antibodies to both Plasmodium falciparum and Plasmodium vivax MSP142 in healthy Cambodian adults was conducted at two sites as part of an active, observational cohort evaluating the efficacy of dihydroartemisinin-piperaquine (DP) for uncomplicated malaria (ClinicalTrials.gov identifier NCT01280162). Results: Rates of baseline sero-positivity were high (59 and 73 % for PfMSP142 and PvMSP142, respectively), and titers higher in those who lived in a higher transmission area, although there was little correlation in titers between the two species. Those volunteers who subsequently went on to develop malaria had higher baseline MSP142 titers than those who did not for both species. Titers to both antigens remained largely stable over the course of the 4-6 month study, except in those infected with P. falciparum who had multiple recurrences. Conclusion: These findings illuminate the difficulties in using MSP142 serology as either a screening criterion and/or biomarker of exposure in chemoprophylaxis studies. Further work remains to identify useful markers of malarial infection and/or immunity. [ABSTRACT FROM AUTHOR]

Background: As a nation reduces the burden of falciparum malaria, identifying areas of transmission becomes increasingly difficult. Over the past decade, the field of utilizing malaria serological assays to measure exposure has grown rapidly, and a variety of serological methods for data acquisition and analysis of human IgG against falciparum antigens are available. Here, different immunoassays and statistical methods are utilized to analyse samples from a low transmission setting and directly compare the estimates generated. Methods: A subset of samples (n = 580) from a 2012 Haitian nationwide malaria survey was employed as sample population of low falciparum endemicity. In addition to the Haitian samples, samples from 247 US residents were used as a reference population of 'true seronegatives'. Data acquisition was performed through standard ELISA and bead-based multiplex assays assaying for IgG antibodies to the Plasmodium falciparum antigens MSP-1p19, MSP- 1p42(D), MSP-1p42(F), and AMA-1. Appropriate parametric distributions and seropositivity cutoff values were determined by statistical measures. Results: Data from both assays showed a strong positive skew, and the lognormal distribution was found to be an appropriate statistical fit to the Haitian and American populations. The American samples served as a good serological true negative population for the multiplex assay, but not for ELISA-based data. Mixture model approaches to determine seronegative and seropositive populations from the Haitian data showed a high degree of distribution overlap- likely due to the historical low falciparum transmission in this nation. Different fittings to the reversible catalytic model resulted depending upon the immunoassay utilized and seropositivity cutoff method employed. Data were also analysed through fitting to penalized B-splines, presenting another possible analytical tool for the analysis of malaria serological data. Conclusions: Standardization of serological techniques and analyses may prove difficult as some tools can prove to be more useful depending on the area and parasite in question, making clear interpretation a vital pursuit. The presented analysis in the low-endemic nation of Haiti found malaria-naive US residents to be an appropriate seronegative reference population for the multiplex assay, and this assay providing consistent estimates between MSP-1 and AMA-1 antigens of percent seropositives for this low-endemic population. [ABSTRACT FROM AUTHOR]

Correction to: I npj Vaccines i https://doi.org/10.1038/s41541-020-0192-7, published online 04 June 2020 In the original version of this Article, the Data Availability statement was missing the link to the datasets generated during the study. The Data Availability statement has now been updated in both the HTML and PDF versions of the Article. [Extracted from the article]

Background: Reported malaria cases continue to decline globally, and this has been attributed to strategic implementation of multiple malaria control tools. Gains made would however need to be sustained through continuous monitoring to ensure malaria elimination and eradication. Entomological inoculation rate (EIR) is currently the standard tool for transmission monitoring but this is not sensitive enough, especially in areas of very low transmission. Transmission estimation models based on seroconversion rates (λ) of antibodies to Plasmodium falciparum blood stage antigens are gaining relevance. Estimates of λ, which is the measure of transmission intensity, correlate with EIR but are limited by long-term persistence of antibodies to blood stage antigens. Seroprevalence of antibodies to sporozoite antigens may be better alternatives since these antigens usually have shorter immune exposure times. The aim of this study was to develop transmission estimation models based on the seroprevalence of antibodies to two P. falciparum sporozoite antigens (CSP, CelTOS) and compare with models based on the classical blood stage antigen AMA1. Methods: Antibody levels in archived plasma from three cross-sectional surveys conducted in 2009 in a low transmission area of Southern Ghana were assessed by indirect ELISA. Seroprevalence of antibodies against CSP, CelTOS and AMA1 were fitted to reversible catalytic models to estimate λ and corresponding seroreversion rates (ρ) for each antibody. Results: Of the three models developed, the anti-CSP model predicted a 13-fold decrease in λ four years prior to the time of sampling (2009). Anti-AMA1 antibodies formed at a four-fold greater rate compared to that of anti-CelTOS antibodies, and anti-CSP antibodies during the period of decreased λ. In contrast, anti-AMA1 antibodies decayed at a five-fold slower rate relative to that of anti-CSP antibodies while anti-AMA1 and anti-CelTOS antibody decay rates were not significantly different. Anti-CSP antibodies were relatively short-lived as they formed at an 11.6-fold slower rate relative to their decay during the period of decreased λ. Conclusions: These features of anti-CSP antibodies can be exploited for the development of models for predicting seasonal, short-term changes in transmission intensity in malaria-endemic areas, especially as the elimination phase of malaria control is approached. [ABSTRACT FROM AUTHOR]

Vaccine development efforts will be guided by algorithms that predict immunogenic epitopes. Such prediction methods rely on classification-based algorithms that are trained against curated data sets of known B and T cell epitopes. It is unclear whether this empirical approach can be applied prospectively to predict epitopes associated with protective immunity for novel antigens. We present a comprehensive comparison of in silico B and T cell epitope predictions with in vivo validation using an previously uncharacterized malaria antigen, CelTOS. CelTOS has no known conserved structural elements with any known proteins, and thus is not represented in any epitope databases used to train prediction algorithms. This analysis represents a blind assessment of this approach in the context of a novel, immunologically relevant antigen. The limited accuracy of the tested algorithms to predict the in vivo immune responses emphasizes the need to improve their predictive capabilities for use as tools in vaccine design. [ABSTRACT FROM AUTHOR]

Genetically inactivated, Gram-negative bacteria that express malaria vaccine candidates represent a promising novel self-adjuvanting vaccine approach. Antigens expressed on particulate bacterial carriers not only target directly to antigen-presenting cells but also provide a strong danger signal thus circumventing the requirement for potent extraneous adjuvants. E. coli expressing malarial antigens resulted in the induction of eitherTh1 orTh2 biased responses that were dependent on both antigen and sub-cellular localization. Some of these constructs induced higher quality humoral responses compared to recombinant protein and most importantly they were able to induce sterile protection against sporo zoite challenge in a murine model of malaria. In light of these encouraging results, two major Plasmodium falciparum pre-erythrocytic malaria vaccine targets, the Cell-Traversal protein for Ookinetes and Sporozoites (CelTOS) fused to the Maltose-binding protein in the periplasmic space and the Circumsporozoite Protein (CSP) fused to the Outer membrane (OM) protein A in the OM were expressed in a clinically relevant, attenuated Shigella strain (Shigella flexneri 2a). This type of live-attenuated vector has previously undergone clinical investigations as a vaccine against shigellosis. Using this novel delivery platform for malaria, we find that vaccination with the whole-organism represents an effective vaccination alter native that induces protective efficacy against sporozoite challenge. Shigella GeMI-Vax expressing malaria targets warrant further evaluation to determine their full potential as a dual disease, multivalent, self-adjuvanting vaccine system, against both shigellosis, and malaria. [ABSTRACT FROM AUTHOR]

Background: The development of an asexual blood stage vaccine against Plasmodium falciparum malaria based on the major merozoite surface protein-1 (MSP1) antigen is founded on the protective efficacy observed in preclinical studies and induction of invasion and growth inhibitory antibody responses. The 42 kDa C-terminus of MSP1 has been developed as the recombinant protein vaccine antigen, and the 3D7 allotype, formulated with the Adjuvant System AS02A, has been evaluated extensively in human clinical trials. In preclinical rabbit studies, the FVO allele of MSP142 has been shown to have improved immunogenicity over the 3D7 allele, in terms of antibody titres as well as growth inhibitory activity of antibodies against both the heterologous 3D7 and homologous FVO parasites. Methods: Two Phase 1 clinical studies were conducted to examine the safety, reactogenicity and immunogenicity of the FVO allele of MSP142 in the adjuvant system AS01 administered intramuscularly at 0-, 1-, and 2-months: one in the USA and, after evaluation of safety data results, one in Western Kenya. The US study was an open-label, dose escalation study of 10 and 50 μg doses of MSP142 in 26 adults, while the Kenya study, evaluating 30 volunteers, was a double-blind, randomized study of only the 50 μg dose with a rabies vaccine comparator. Results: In these studies it was demonstrated that this vaccine formulation has an acceptable safety profile and is immunogenic in malaria-naïve and malaria-experienced populations. High titres of anti-MSP1 antibodies were induced in both study populations, although there was a limited number of volunteers whose serum demonstrated significant inhibition of blood-stage parasites as measured by growth inhibition assay. In the US volunteers, the antibodies generated exhibited better cross-reactivity to heterologous MSP1 alleles than a MSP1-based vaccine (3D7 allele) previously tested at both study sites. Conclusions: Given that the primary effector mechanism for blood stage vaccine targets is humoral, the antibody responses demonstrated to this vaccine candidate, both quantitative (total antibody titres) and qualitative (functional antibodies inhibiting parasite growth) warrant further consideration of its application in endemic settings. [ABSTRACT FROM AUTHOR]