scholarly journals Measuring Antibody Avidity To Plasmodium Falciparum Merozoite Antigens Using A Multiplex Immunoassay Approach

2020 ◽  
Author(s):  
Diane Wallace Taylor ◽  
Naveen Bobbili ◽  
Alexander K Kayatani ◽  
Samuel Tassi-Yunga ◽  
Rose FG Leke
Keyword(s):  
Plasmodium Falciparum ◽  
Clonal Selection ◽  
Ammonium Thiocyanate ◽  
Plasmodium Falciparum Malaria ◽  
Affinity Maturation ◽  
Binding Studies ◽  
Simple Method ◽  
Antibody Avidity ◽  
Merozoite Antigens ◽  
One Year

Abstract Background: Antibodies (Ab) play a significant role in immunity to Plasmodium falciparum malaria. Usually, following repeated exposure to pathogens, affinity maturation and clonal selection take place, resulting in increased antibody avidity. However, some studies suggest affinity maturation may not take place to malaria antigens in endemic areas. Information on development of antibody avidity is confusing and conflicting, in part, because different techniques have been used to measure avidity. Today, bead-based multiplex immunoassays (MIA) are routinely used to simultaneously quantitate antibody levels to multiple antigens. This study evaluated the feasibility of developing an avidity MIA with 5 merozoite antigens (AMA1, EBA-175, MSP1-42, MSP2, MSP3) that used a single chaotropic concentration. Methods: The most common ELISA protocols that used the chaotropic reagents guanidine HCl (GdHCl), urea, and ammonium thiocyanate (NH4SCN) were adapted to a multiplex MIA format. Then, different concentrations of chaotropes and incubation times were compared and results were expressed as an Avidity Index (AI), i.e., percentage of antibody remaining bound in the presence of chaotrope. Experiments were conducted to i) identify the assay with the widest range of AI (discriminatory power), ii) determine the amount of chaotrope needed to release 50% of bound Ab using plasma from adults and infants, and iii) evaluate assay repeatability. Results: Overall, 4M GdHCl and 8M urea were weaker chaotropes than 3M NH4SCN. For example, they failed to release significant amounts of Ab bound to MSP1-42 in adult plasma samples; whereas, a range of AI values was obtained with NH4SCN. Titration of NH4SCN revealed that 2M urea gave the widest range of AI for the 5 antigens. Binding studies using plasma from 40 adults and 57 one-year old infants in Cameroon showed that 2.1M ± 0.32 (mean ± SD) NH4SCN (adults) and 1.8M ± 0.23M released 50% of bound Ab from the merozoite antigens. The final avidity multiplex assay was highly repeatable. Conclusions. An avidity MIA is feasible for the 5 merozoite antigens that uses a single concentration (2M) of NH4SCN. The assay provides a simple method to quickly obtain information about Ab quantity and quality in the acquisition of immunity to malaria in endemic populations.

scholarly journals Measuring Antibody Avidity to Plasmodium Falciparum Merozoite Antigens Using a Multiplex Immunoassay Approach

2020 ◽  
Author(s):  
Diane Wallace Taylor ◽  
Naveen Bobbili ◽  
Alexander K Kayatani ◽  
Samuel Tassi-Yunga ◽  
Winifrida Kidima ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Clonal Selection ◽  
Ammonium Thiocyanate ◽  
Affinity Maturation ◽  
Binding Studies ◽  
Simple Method ◽  
Antibody Avidity ◽  
Merozoite Antigens ◽  
Adult Plasma ◽  
One Year

Abstract Background: Antibodies (Ab) play a significant role in immunity to Plasmodium falciparum malaria. Usually, following repeated exposure to pathogens, affinity maturation and clonal selection take place, resulting in increased antibody avidity. However, some studies suggest affinity maturation may not occur to malaria antigens in endemic areas. Information on development of antibody avidity is confusing and conflicting, in part, because different techniques have been used to measure avidity. Today, bead-based multiplex immunoassays (MIA) are routinely used to simultaneously quantitate antibody levels to multiple antigens. This study evaluated the feasibility of developing an avidity MIA with 5 merozoite antigens (AMA1, EBA-175, MSP1-42, MSP2, MSP3) that uses a single chaotropic concentration.Methods: The most common ELISA protocols that used the chaotropic reagents guanidine HCl (GdHCl), urea, and ammonium thiocyanate (NH4SCN) were adapted to a multiplex MIA format. Then, different concentrations of chaotropes and incubation times were compared and results were expressed as an Avidity Index (AI), i.e., percentage of antibody remaining bound in the presence of chaotrope. Experiments were conducted to i) identify the assay with the widest range of AI (discriminatory power), ii) determine the amount of chaotrope needed to release 50% of bound Ab using plasma from adults and infants, and iii) evaluate assay repeatability. Results: Overall, 4M GdHCl and 8M urea were weaker chaotropes than 3M NH4SCN. For example, they failed to release significant amounts of Ab bound to MSP1-42 in adult plasma samples; whereas, a range of AI values was obtained with NH4SCN. Titration of NH4SCN revealed that 2M NH4SCN gave the widest range of AI for the 5 antigens. Binding studies using plasma from 40 adults and 57 one-year old infants in Cameroon showed that 2.1M ± 0.32 (mean ± SD) NH4SCN (adults) and 1.8M ± 0.23M (infants) released 50% of bound Ab from the merozoite antigens. Conclusions. An avidity MIA is feasible for the 5 merozoite antigens that uses a single concentration (2M) of NH4SCN. The assay provides a simple method to quickly obtain information about Ab quantity and quality in the acquisition of immunity to malaria in endemic populations.

scholarly journals Measuring Antibody Avidity to Plasmodium Falciparum Merozoite Antigens Using a Multiplex Immunoassay Approach

2020 ◽  
Author(s):  
Diane Wallace Taylor ◽  
Naveen Bobbili ◽  
Alexander K Kayatani ◽  
Samuel Tassi-Yunga ◽  
Winifrida Kidima ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Clonal Selection ◽  
Ammonium Thiocyanate ◽  
Affinity Maturation ◽  
Binding Studies ◽  
Simple Method ◽  
Antibody Avidity ◽  
Merozoite Antigens ◽  
Adult Plasma ◽  
One Year

Abstract Background: Antibodies (Ab) play a significant role in immunity to Plasmodium falciparum malaria. Usually, following repeated exposure to pathogens, affinity maturation and clonal selection take place, resulting in increased antibody avidity. However, some studies suggest affinity maturation may not occur to malaria antigens in endemic areas. Information on development of antibody avidity is confusing and conflicting, in part, because different techniques have been used to measure avidity. Today, bead-based multiplex immunoassays (MIA) are routinely used to simultaneously quantitate antibody levels to multiple antigens. This study evaluated the feasibility of developing an avidity MIA with 5 merozoite antigens (AMA1, EBA-175, MSP1-42, MSP2, MSP3) that uses a single chaotropic concentration.Methods: The most common ELISA protocols that used the chaotropic reagents guanidine HCl (GdHCl), urea, and ammonium thiocyanate (NH4SCN) were adapted to a multiplex MIA format. Then, different concentrations of chaotropes and incubation times were compared and results were expressed as an Avidity Index (AI), i.e., percentage of antibody remaining bound in the presence of chaotrope. Experiments were conducted to i) identify the assay with the widest range of AI (discriminatory power), ii) determine the amount of chaotrope needed to release 50% of bound Ab using plasma from adults and infants, and iii) evaluate assay repeatability. Results: Overall, 4M GdHCl and 8M urea were weaker chaotropes than 3M NH4SCN. For example, they failed to release significant amounts of Ab bound to MSP1-42 in adult plasma samples; whereas, a range of AI values was obtained with NH4SCN. Titration of NH4SCN revealed that 2M NH4SCN gave the widest range of AI for the 5 antigens. Binding studies using plasma from 40 adults and 57 one-year old infants in Cameroon showed that 2.1M ± 0.32 (mean ± SD) NH4SCN (adults) and 1.8M ± 0.23M (infants) released 50% of bound Ab from the merozoite antigens. Conclusions: An avidity MIA is feasible for the 5 merozoite antigens that uses a single concentration (2M) of NH4SCN. The assay provides a simple method to quickly obtain information about Ab quantity and quality in the acquisition of immunity to malaria in endemic populations.

scholarly journals Memory B-Cell Responses Against Merozoite Antigens After Acute Plasmodium falciparum Malaria, Assessed Over One Year Using a Novel Multiplexed FluoroSpot Assay

2021 ◽  
Vol 11 ◽  
Author(s):  
Peter Jahnmatz ◽  
Christopher Sundling ◽  
Victor Yman ◽  
Linnea Widman ◽  
Muhammad Asghar ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
B Cell ◽  
Plasmodium Falciparum Malaria ◽  
Sub Saharan Africa ◽  
Single Infection ◽  
Cell Responses ◽  
Merozoite Antigens ◽  
Sub Saharan ◽  
One Year ◽  
Kinetics Of

Memory B cells (MBCs) are believed to be important for the maintenance of immunity to malaria, and these cells need to be explored in the context of different parasite antigens and their breadth and kinetics after natural infections. However, frequencies of antigen-specific MBCs are low in peripheral blood, limiting the number of antigens that can be studied, especially when small blood volumes are available. Here, we developed a multiplexed reversed B-cell FluoroSpot assay capable of simultaneously detecting MBCs specific for the four Plasmodium falciparum blood-stage antigens, MSP-119, MSP-2, MSP-3 and AMA-1. We used the assay to study the kinetics of the MBC response after an acute episode of malaria and up to one year following treatment in travelers returning to Sweden from sub-Saharan Africa. We show that the FluoroSpot assay can detect MBCs to all four merozoite antigens in the same well, and that the breadth and kinetics varied between individuals. We further found that individuals experiencing a primary infection could mount and maintain parasite-specific MBCs to a similar extent as previously exposed adults, already after a single infection. We conclude that the multiplexed B-cell FluoroSpot is a powerful tool for assessing antigen-specific MBC responses to several antigens simultaneously, and that the kinetics of MBC responses against merozoite surface antigens differ over the course of one year. These findings contribute to the understanding of acquisition and maintenance of immune responses to malaria.

scholarly journals Measuring antibody avidity to Plasmodium falciparum merozoite antigens using a multiplex immunoassay approach

2020 ◽  
Vol 19 (1) ◽  
Author(s):  
Diane Wallace Taylor ◽  
Naveen Bobbili ◽  
Alex Kayatani ◽  
Samuel Tassi Yunga ◽  
Winifrida Kidima ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Antibody Avidity ◽  
Multiplex Immunoassay ◽  
Falciparum Merozoite ◽  
Merozoite Antigens

scholarly journals Acquired Antibodies to Merozoite Antigens in Children from Uganda with Uncomplicated or Severe Plasmodium falciparum Malaria

2013 ◽  
Vol 20 (8) ◽  
pp. 1170-1180 ◽  
Author(s):  
Hodan Ahmed Ismail ◽  
Ulf Ribacke ◽  
Linda Reiling ◽  
Johan Normark ◽  
Tom Egwang ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Uncomplicated Malaria ◽  
Severe Disease ◽  
Plasmodium Falciparum Malaria ◽  
Antibody Responses ◽  
Enzyme Linked Immunosorbent Assay ◽  
Content Type ◽  
Apical Membrane Antigen 1 ◽  
Merozoite Antigens ◽  
Erythrocyte Binding

ABSTRACTMalaria can present itself as an uncomplicated or severe disease. We have here studied the quantity and quality of antibody responses against merozoite antigens, as well as multiplicity of infection (MOI), in children from Uganda. We found higher levels of IgG antibodies toward erythrocyte-binding antigen EBA181, MSP2 ofPlasmodium falciparum3D7 and FC27 (MSP2-3D7/FC27), and apical membrane antigen 1 (AMA1) in patients with uncomplicated malaria by enzyme-linked immunosorbent assay (ELISA) but no differences against EBA140, EBA175, MSP1, and reticulocyte-binding protein homologues Rh2 and Rh4 or for IgM against MSP2-3D7/FC27.Patients with uncomplicated malaria were also shown to have higher antibody affinities for AMA1 by surface plasmon resonance (SPR). Decreased invasion of two clinicalP. falciparumisolates in the presence of patient plasma correlated with lower initial parasitemia in the patients, in contrast to comparisons of parasitemia to ELISA values or antibody affinities, which did not show any correlations. Analysis of the heterogeneity of the infections revealed a higher MOI in patients with uncomplicated disease, with theP. falciparumK1 MSP1 (MSP1-K1) and MSP2-3D7 being the most discriminative allelic markers. Higher MOIs also correlated positively with higher antibody levels in several of the ELISAs. In conclusion, certain antibody responses and MOIs were associated with differences between uncomplicated and severe malaria. When different assays were combined, some antibodies, like those against AMA1, seemed particularly discriminative. However, only decreased invasion correlated with initial parasitemia in the patient, signaling the importance of functional assays in understanding development of immunity against malaria and in evaluating vaccine candidates.

scholarly journals Acquisition and decay of IgM and IgG responses to merozoite antigens after Plasmodium falciparum malaria in Ghanaian children

PLoS ONE ◽  
2020 ◽  
Vol 15 (12) ◽  
pp. e0243943
Author(s):  
Melanie R. Walker ◽  
Anne S. Knudsen ◽  
Frederica D. Partey ◽  
Maria R. Bassi ◽  
Asger M. Frank ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Falciparum Malaria ◽  
Longitudinal Analysis ◽  
Malaria Vaccine ◽  
Second Generation ◽  
Plasmodium Falciparum Malaria ◽  
Initial Diagnosis ◽  
Serum Samples ◽  
Growth Inhibitory ◽  
Merozoite Antigens

Developing a vaccine against Plasmodium falciparum malaria has been challenging, primarily due to high levels of antigen polymorphism and a complex parasite lifecycle. Immunization with the P. falciparum merozoite antigens PfMSRP5, PfSERA9, PfRAMA, PfCyRPA and PfRH5 has been shown to give rise to growth inhibitory and synergistic antisera. Therefore, these five merozoite proteins are considered to be promising candidates for a second-generation multivalent malaria vaccine. Nevertheless, little is known about IgG and IgM responses to these antigens in populations that are naturally exposed to P. falciparum. In this study, serum samples from clinically immune adults and malaria exposed children from Ghana were studied to compare levels of IgG and IgM specific for PfMSRP5, PfSERA9, PfRAMA, PfCyRPA and PfRH5. All five antigens were found to be specifically recognized by both IgM and IgG in serum from clinically immune adults and from children with malaria. Longitudinal analysis of the latter group showed an early, transient IgM response that was followed by IgG, which peaked 14 days after the initial diagnosis. IgG levels and parasitemia did not correlate, whereas parasitemia was weakly positively correlated with IgM levels. These findings show that IgG and IgM specific for merozoite antigens PfMSRP5, PfSERA9, PfRAMA, PfCyRPA and PfRH5 are high in children during P. falciparum malaria, but that the IgM induction and decline occurs earlier in infection than that of IgG.

scholarly journals Delayed clinical presentation of Plasmodium falciparum malaria after leaving an endemic area: a tale of two patients

2020 ◽  
Vol 27 (6) ◽  
Author(s):  
Kaylin Pennington ◽  
Samuel T Ives ◽  
Anne E P Frosch ◽  
Megan K Shaughnessy
Keyword(s):  
Plasmodium Falciparum ◽  
Falciparum Malaria ◽  
Incubation Period ◽  
Endemic Area ◽  
Clinical Presentation ◽  
Plasmodium Falciparum Malaria ◽  
International Airport ◽  
Source Of Infection ◽  
One Year

Malaria due to Plasmodium falciparum (Pf) may be missed if patients present with symptoms outside of the expected incubation period. We describe two patients who developed Pf malaria more than one year after visiting malaria-endemic countries. Both worked at an international airport, but no source of infection was identified.

scholarly journals Does Antibody Avidity to Plasmodium falciparum Merozoite Antigens increase with Age in Individuals living in Malaria-Endemic Areas?

2021 ◽  
Author(s):  
Samuel Tassi Yunga ◽  
Naveen Bobbili ◽  
Yukie M. Lloyd ◽  
Jovikka Antallan ◽  
Masako Matsunaga ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Affinity Maturation ◽  
High Avidity ◽  
Plasma Samples ◽  
Avidity Index ◽  
Hyperendemic Area ◽  
Age Related ◽  
Falciparum Merozoite ◽  
Merozoite Antigens ◽  
Strength Of Association

Introduction: High avidity antibodies (Abs) are acquired after a few Plasmodium falciparum infections in low transmission areas, but it remains unclear if Ab avidity to different merozoite antigens increases with age in individuals with persistent antigenemia and if so, when a fully mature Ab response occurs. Methods: The study used plasma samples collected between 1996 and 1998 from 566 individuals aged 4-84 years in Simbok, Cameroon where residents received an estimated 1.6 infectious mosquito bites/person/night. Plasma samples were examined for Ab levels (median fluorescence intensity, MFI) and Ab avidity index (AI = [MFI after treatment with 2M NH4SCN/MFI without salt] x 100) using a bead-based multiplex immunoassay for recombinant AMA1, EBA-175, MSP1-42 (3D7, FVO), MSP2 (3D7, Fc27), and MSP3. Results: Blood-smear positivity for P. falciparum declined with age from 54.3% at 4-5 years to 18% at 16-40 years and <11% at >40 years of age, although most individuals had submicroscopic parasitemia. Ab affinity maturation, based on age-related patterns of median AI, percent of individuals with AI ≥50 and strength of association between MFI and AI, occurred at different rates among the antigens: developing rapidly before age 4 years for AMA1, increasing gradually with age for EBA-175 and MSP1 until ∼16-25 years, but occurring negligibly for MSP2 and MSP3. Conclusion: In a hyperendemic area with perennial transmission, affinity maturation resulting in an increase in the proportion of high avidity Abs occurred for some merozoite antigens, in parallel with a decline in malaria slide passivity, but not for others.

scholarly journals Targets and Mechanisms Associated with Protection from Severe Plasmodium falciparum Malaria in Kenyan Children

2016 ◽  
Vol 84 (4) ◽  
pp. 950-963 ◽  
Author(s):  
Linda M. Murungi ◽  
Klara Sondén ◽  
David Llewellyn ◽  
Josea Rono ◽  
Fatuma Guleid ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Rank Correlation ◽  
Plasmodium Falciparum Malaria ◽  
Cell Surface Antigens ◽  
Immunological Parameters ◽  
Content Type ◽  
Merozoite Antigens ◽  
Life Threatening ◽  
Matched Case ◽  
Functional Mechanisms

Severe malaria (SM) is a life-threatening complication of infection withPlasmodium falciparum. Epidemiological observations have long indicated that immunity against SM is acquired relatively rapidly, but prospective studies to investigate its immunological basis are logistically challenging and have rarely been undertaken. We investigated the merozoite targets and antibody-mediated mechanisms associated with protection against SM in Kenyan children aged 0 to 2 years. We designed a unique prospective matched case-control study of well-characterized SM clinical phenotypes nested within a longitudinal birth cohort of children (n= 5,949) monitored over the first 2 years of life. We quantified immunological parameters in sera collected before the SM event in cases and their individually matched controls to evaluate the prospective odds of developing SM in the first 2 years of life. Anti-AMA1 antibodies were associated with a significant reduction in the odds of developing SM (odds ratio [OR] = 0.37; 95% confidence interval [CI] = 0.15 to 0.90;P= 0.029) after adjustment for responses to all other merozoite antigens tested, while those against MSP-2, MSP-3,Plasmodium falciparumRh2 [PfRh2], MSP-119, and the infected red blood cell surface antigens were not. The combined ability of total IgG to inhibit parasite growth and mediate the release of reactive oxygen species from neutrophils was associated with a marked reduction in the odds of developing SM (OR = 0.07; 95% CI = 0.006 to 0.82;P= 0.03). Assays of these two functional mechanisms were poorly correlated (Spearman rank correlation coefficient [rs] = 0.12;P= 0.07). Our data provide epidemiological evidence that multiple antibody-dependent mechanisms contribute to protective immunity via distinct targets whose identification could accelerate the development of vaccines to protect against SM.

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