High-avidity Antibodies Research Articles

A two-dose regimen of Qβ virus-like particle-based vaccines elicit protective antibodies against heroin and fentanyl.

Opioid overdoses and the growing rate of opioid use disorder (OUD) are major public health concerns, particularly in the United States. Current treatment approaches for OUD have failed to slow the growth of the opioid crisis. Opioid vaccines have shown pre-clinical success in targeting multiple different opioid drugs. However, the need for many immunizations can limit their clinical implementation. In this study, we investigate the development of novel opioid vaccines by independently targeting fentanyl and the active metabolites of heroin using a bacteriophage virus-like particle (VLP) vaccine platform. We establish the successful conjugation of haptens to bacteriophage Qβ VLPs and demonstrate immunogenicity of Qβ-fentanyl, Qβ-morphine, and Qβ-6-acetylmorphine in animal models after one or two immunizations. We show that in independently or in combination, these vaccines elicit high-titer, high-avidity, and durable antibody responses. Moreover, we reveal their protective capacities against heroin or fentanyl challenge after two immunizations. Overall, these findings establish Qβ-VLP conjugated vaccines for heroin and fentanyl as very promising opioid vaccine candidates.

Measles immunity status in Iranian infants and children and outbreak concerns: Time for reconsidering the vaccination schedule?

Introduction: Measles vaccination has greatly reduced the disease burden worldwide, but challenges remain due to variations in vaccine effectiveness across age groups. This study aimed to assess the serological profile of measles antibodies across different age groups, evaluate the impact of maternal immunity on antibody levels in infants under 12 months, and assess measles immunity in vaccinated individuals. Material and methods: This cross-sectional study was conducted from June 2022 to January 2023 at the Children’s Medical Center, a referral hospital in Iran. Serum samples were tested for measles-specific IgG and IgM antibodies using a commercial enzyme-linked immunosorbent assay (ELSA). An avidity assay was performed to assess measles virus-specific IgG antibodies on the samples that were positive and borderline for the measles IgG ELISA. Results: The study included 969 participants across various age groups. Among them, 23% (221 out of 953) tested positive for measles IgM ELISA, and 52% (504 out of 969) for measles IgG ELISA. Regarding the avidity assay for measles virus-specific IgG, the majority (418 out of 573, 73%) showed high-avidity antibodies. Measles-specific IgG levels varied significantly across different age groups, with infants below 6 months old showing a mean IgG level of 477 mIU/mL, declining to 230 mIU/mL between 6 and 12 months, and increasing significantly to 683 mIU/mL in the 12 to 18 month age group, reaching a peak at 938 mIU/mL among children aged 18–72 months. Conclusion: The increasing IgM positivity among young Iranians suggests a rising risk of measles outbreaks, possibly due to vaccination gaps. Inadequate antibody levels in infants raise concerns about vaccination effectiveness. Considering declining maternal antibodies, vaccinating infants at 6–9 months could be beneficial. Boosters for adolescents and women may further mitigate outbreak risks.

Determination of the immune response type in measles patients of different ages from a high-incidence area

According to the WHO data, the incidence of measles is now in the cyclic increase phase. The non-vaccinated subjects, like as persons who have previously received one or two doses of measles vaccine are involved in the epidemic process. The purpose of our study was to determine the immunological pattern in the groups of measles patients of different age in areas with high incidence of infection. To determine the immune response type in measles patients, qualitative and quantitative indices of IgM and IgG were used, detected by the following ELISA test-systems: VectoMeasles-IgM (JSC “Vector-Best”, Russia); Anti-Measles Viruses ELISA (IgG) and Avidity Anti-Measles Viruses ELISA IgG (Euroimmun, Germany). The serological study of 1893 patient allowed to determine the primary and secondary types of immune response. In 72.64% of the patients with primary immune response, the serum samples contained low-avidity IgG at a concentration of 0.45 (0.22- 0.74) IU/mL. In the group with a secondary immune response, high-avidity antibodies were detected at a concentration of 24.28 (21.59-27.4) IU/mL. The antibody levels in secondary type of response was 54-fold higher than IgG values in the first group (p 0.05). In the group with primary immune response, the ratio of children ( 1 to 14 years) and adults (18 to 70 years old) was almost the same (49.6% and 47.56%). Appropriate values for children and adult cohorts with a secondary immune response were 2.12% and 96.53%, respectively (p 0.05). Among 46 teenagers (15-17 years old), 84.8% responded with a primary immune response to the measles virus. Thus, in the area with high-incidence of measles among patients of different age, the primary-type immune response was determined in 72.64%, and secondary-type, in 27.36%. We have found that in the patients with a secondary type of immune response group, the proportion of children and teenagers was 27.8 times lower than among adults, thus indicating to high efficiency of vaccination in pediatric population. At the same time, the results of studies among measles patients with primary-type immune response, both children and adults, suggest some “gaps” in the vaccine prevention program.

Site-specific serology unveils cross-reactive monoclonal antibodies targeting influenza A hemagglutinin epitopes.

Efficient identification of human monoclonal antibodies targeting specific antigenic sites is pivotal for advancing vaccines and immunotherapies against infectious diseases and cancer. Existing screening techniques, however, limit our ability to discover monoclonal antibodies with desired specificity. In this study, we introduce a novel method, blocking of binding (BoB) enzyme-linked immunoassay (ELISA), enabling the detection of high-avidity human antibodies directed to defined epitopes. Leveraging BoB-ELISA, we analyzed the antibody response to known epitopes of influenza A hemagglutinin (HA) in the serum of vaccinated donors. Our findings revealed that serum antibodies targeting head epitopes were immunodominant, whereas antibodies against the stem epitope, although subdominant, were highly prevalent. Extending our analysis across multiple HA strains, we examined the cross-reactive antibody response targeting the stem epitope. Importantly, employing BoB-ELISA we identified donors harboring potent heterosubtypic antibodies targeting the HA stem. B-cell clonal analysis of these donors revealed three novel, genealogically independent monoclonal antibodies with broad cross-reactivity to multiple HAs. In summary, we demonstrated that BoB-ELISA is a sensitive technique for measuring B-cell epitope immunogenicity, enabling the identification of novel monoclonal antibodies with implications for enhanced vaccine development and immunotherapies.

Malaria transmission blocking activity of Anopheles stephensi alanyl aminopeptidase N antigen formulated with MPL, CpG, and QS21 adjuvants.

Malaria, a preventive and treatable disease, is still responsible for annual deaths reported in most tropical regions, principally in sub-Saharan Africa. Subunit recombinant transmission-blocking vaccines (TBVs) have been proposed as promising vaccines to succeed in malaria elimination and eradication. Here, a provisional study was designed to assess the immunogenicity and functional activity of alanyl aminopeptidase N (APN1) of Anopheles stephensi, as a TBV candidate, administered with MPL, CpG, and QS21 adjuvants in the murine model. The mouse groups were immunized with recombinant APN1 (rAPN1) alone or formulated with CpG, MPL, QS-21, or a combination of adjuvants (CMQ), and the elicited immune responses were evaluated after the third immunization. The standard membrane feeding assay (SMFA) measured the functional activity of antibodies against bacterial-expressed APN1 protein in adjuvanted vaccine groups on transmission of P. falciparum (NF54) to An. stephensi mosquitoes. Evaluation of mice vaccinated with rAPN1 formulated with distinct adjuvants manifested a significant increase in the high-avidity level of anti-APN1 IgG and IgG subclasses; however, rAPN1 induced the highest level of high-avidity anti-APN1 IgG1, IgG2a, and IgG2b antibodies in the immunized vaccine group 5 (APN1/CMQ). In addition, vaccine group 5 (receiving APN1/CMQ), had still the highest level of anti-APN1 IgG antibodies relative to other immunized groups after six months, on day 180. The SMFA data indicates a trend towards higher transmission-reducing activity in groups 2 and 5, which received the antigen formulated with CpG or a combination of three adjuvants. The results have shown the capability of admixture to stimulate high-affinity and long-lasting antibodies against the target antigen to hinder Plasmodium parasite development in the mid-gut of An. stephensi. The attained results authenticated APN1/CMQ and APN1/CpG as a potent APN1-based TBV formulation which will be helpful in designing a vaccine in the future.

Antibody-mediated immunological memory correlates with long-term Lyme veterinary vaccine protection in mice

Lyme disease, caused by the bacterium Borrelia burgdorferi, is the most common tick-borne illness in the United States. Despite the rise in Lyme disease incidence, there is no vaccine against B. burgdorferi approved for human use. Little is known about the immune correlates of protection needed to prevent Lyme disease. In this work, a mouse model was used to characterize the immune response and compare the protection provided by two USDA-approved vaccines for use in canines: Duramune (bacterin vaccine) and Vanguard crLyme (subunit vaccine composed of two outer surface proteins, OspA and OspC). C3H/HeNCrl mice were immunized with two doses of either Duramune or Vanguard, and immune responses and protection against B. burgdorferi were assessed in short (35 days) and long-term (120 days) studies. Flow cytometry, ELISPOT detection of antibody-producing cells, and antibody affinity studies were performed to identify correlates of vaccine-mediated protection. Both vaccines induced humoral responses, with high IgG titers against B. burgdorferi. However, the levels of anti-B. burgdorferi antibodies decayed over time in Vanguard-vaccinated mice. While both vaccines triggered the production of antibodies against both OspA and OspC, antibody levels against these proteins were also lower in Vanguard-vaccinated mice 120 days post-vaccination. Both vaccines only provided partial protection against B. burgdorferi at the dose used in this model. The protection provided by Duramune was superior to Vanguard 120 days post-vaccination, and was characterized by higher antibody titers, higher abundance of long-lived plasma cells, and higher avidity antibodies than Vanguard. Overall, these studies provide insights into the importance of the humoral memory response to veterinary vaccines against Lyme disease and will help inform the development of future human vaccines.

Avidity maturation of humoral response following primary and booster doses of BNT162b2 mRNA vaccine among nursing home residents and healthcare workers

Infections, despite vaccination, can be clinically consequential for frail nursing home residents (NHR). Poor vaccine-induced antibody quality may add risk for such subsequent infections and more severe disease. We assessed antibody binding avidity, as a surrogate for antibody quality, among NHR and healthcare workers (HCW). We longitudinally sampled 112 NHR and 52 HCWs who received the BNT162b2 mRNA vaccine after each dose up to the Wuhan-BA.4/5-based Omicron bivalent boosters. We quantified anti-spike, anti-receptor binding domain (RBD), and avidity levels to the ancestral Wuhan, Delta, and Omicron BA.1 & 4/5 strains. The primary vaccination series produced substantial anti-spike and RBD levels which were low in avidity against all strains tested. Antibody avidity progressively increased in the 6–8 months that followed. Avidity significantly increased after the 1st booster but not for subsequent boosters. This study underscores the importance of booster vaccination among NHR and HCWs. The 1st booster dose increases avidity, increasing vaccine-induced functional antibody. The higher cross-reactivity of higher avidity antibodies to other SARS-CoV-2 strains should translate to better protection from ever-evolving strains. Higher avidities may help explain how the vaccine’s protective effects persist despite waning antibody titers after each vaccine dose.

Hybrid response to SARS-CoV-2 and Neisseria meningitidis C after an OMV-adjuvanted immunization in mice and their offspring

ABSTRACT COVID-19, caused by SARS-CoV-2, and meningococcal disease, caused by Neisseria meningitidis, are relevant infectious diseases, preventable through vaccination. Outer membrane vesicles (OMVs), released from Gram-negative bacteria, such as N. meningitidis, present adjuvant characteristics and may confer protection against meningococcal disease. Here, we evaluated in mice the humoral and cellular immune response to different doses of receptor binding domain (RBD) of SARS-CoV-2 adjuvanted by N. meningitidis C:2a:P1.5 OMVs and aluminum hydroxide, as a combined preparation for these pathogens. The immunization induced IgG antibodies of high avidity for RBD and OMVs, besides IgG that recognized the Omicron BA.2 variant of SARS-CoV-2 with intermediary avidity. Cellular immunity showed IFN-γ and IL-4 secretion in response to RBD and OMV stimuli, demonstrating immunologic memory and a mixed Th1/Th2 response. Offspring presented transferred IgG of similar levels and avidity as their mothers. Humoral immunity did not point to the superiority of any RBD dose, but the group immunized with a lower antigenic dose (0.5 μg) had the better cellular response. Overall, OMVs enhanced RBD immunogenicity and conferred an immune response directed to N. meningitidis too.

Seroprevalence and risk factors of Toxoplasma gondii in goats (Capra hircus) in the state of Espírito Santo, Southeast Brazil

Toxoplasma gondii is described as a potential cause of abortion in goats and as a threat to public health. To estimate the prevalence of goats infected by T. gondii, in different cities in the Espírito Santo State, and to identify possible risk factors for infection a serological study was conducted. A total of 146 goat serum samples from the cities of Cariacica, Serra and Vila Velha were analyzed. The presence of IgG Class Immunoglobulins was serologically evaluated by Immunofluorescence antibody test (IFAT) and by Enzyme-linked Immunosorbent Assay (ELISA). The seroprevalence of anti-T. gondii was 46.6% (68/146) in both techniques and the same samples got the same results in both techniques. Among the analyzed sera, 70.6% (48/68) exhibited high-avidity IgG antibodies, and 29.4% (20/68) exhibited low-avidity IgG antibodies, suggesting that the infection was chronic in the infected animals. Female sex, age group over two years old, water from the public supply system, storage of food and supplies in an open and unprotected place, and the presence of a domestic cat on the property were identified as risk factors for T. gondii infection in goats. The state of Espirito Santo has a high frequency of infected goats, and this is the first research on caprine toxoplasmosis seroepidemiology in that region.

Epidemiological and laboratory examination of pregnant women for toxoplasmosis in a polyclinic of the Arkhangelsk region

The article presents the results of examinations of 41 pregnant women by an infectious disease specialist in a clinic for the period from September 22, 2017 to November 19, 2023. The patients were tested for avidity for toxoplasmosis, as well as underwent standard laboratory tests. The results of the epidemic assessment of the situation for each pregnant woman are presented. As a result, it was found that each woman had high-avidity antibodies to toxoplasmosis. These women were not prescribed treatment.

A bacteriophage virus-like particle vaccine against oxycodone elicits high-titer and long-lasting antibodies that sequester drug in the blood

Opioid use disorder (OUD) and opioid overdoses are public health emergencies. In 2021, 80,000 opioid overdose associated deaths were reported in the United States. Despite the availability of treatment strategies, including medications for opioid use disorder (MOUD) and naloxone, opioid overdoses continue to increase at an alarming rate. Opioid vaccines are a novel approach to combat the growing crisis with several candidates recently entering human clinical trials. In this study, we investigated Qβ bacteriophage virus-like particles (VLPs) as a vaccine platform for immunogenic display of oxycodone. A derivative of oxycodone was conjugated to pre-formed Qβ VLPs using a sulfhydryl-amine reactive heterobifunctional crosslinker with high loading of oxycodone. In mice, intramuscular immunization with Qβ-oxycodone elicited high-titer, high-avidity and long-lasting antibody responses. Qβ-oxycodone was also immunogenic after storage at ambient room temperature for over two weeks, demonstrating that the vaccine is highly thermostable. In mice, immunization with Qβ-oxycodone elicited antibodies that sequester oxycodone in the serum, an important mechanism for preventing the adverse effects of opioid activity. Finally, Qβ-oxycodone is immunogenic in nonhuman primates, eliciting serum oxycodone antibodies after intramuscular immunization of rhesus macaques. These data establish Qβ-oxycodone as a promising opioid vaccine candidate.

2373. "Longitudinal Investigation of Avidity Maturation following Multiple Doses of BNT162b2 mRNA Vaccine Including Bivalent Boosters among Nursing Home Residents and Healthcare Workers"

Abstract Background Increased breakthrough infections among nursing home residents (NHR) are clinically consequential despite good vaccine coverage and substantial antibody titers. While vaccination blunted severe outcomes, breakthrough infections may arise from lower vaccine-induced antibody quality. We aimed to assess vaccine-induced antibody quality among NHR and younger healthcare workers (HCW) using antibody avidity. Methods We longitudinally sampled 50 NHR and 30 HCWs, with or without prior history of COVID-19, who received the BNT162b2 mRNA vaccine at 0-14 days before, 2 weeks, and 4-6 months after each vaccine dose up to the bivalent (BV) boosters. Primary outcomes included anti-spike, anti-Receptor Binding Domain (RBD), and avidity levels to the ancestral Wuhan, Delta, and Omicron BA.1 strains determined by bead-multiplex immunoassay and ability to bind after 6M urea vs. saline incubation. Results After the primary series, anti-spike and RBD titers increased significantly to all 3 strains in both NHR and HCW. Avidity levels progressively increased across all time points up to the 3rd dose. Interestingly, this increase continued 6-8 months after vaccination suggesting that affinity maturation continues to occur for many months after the primary vaccination series. NHRs without prior infection have the poorest avidity levels to Delta and Omicron, especially before the 3rd dose. The 3rd dose raised avidity levels substantially in all groups and for all strains. Avidity levels after boosting, across the 3rd and 4th doses, demonstrate evidence of durability. Preliminary findings after BV boost show a continuation of avidity maturation with a larger impact on prior infected individuals. Wuhan Spike Avidity Avidity index of the Wuhan spike antibody across different time points among naive and prior infected individuals. Prevax- before primary series; Postvax- 2 weeks after primary series; M6- 6 months after primary series; Postboost- 2 weeks after 1st monovalent booster; M3M6postboost- 3-6months post booster; Postboost 2- 2 weeks after 2nd monovalent booster. NH- Nursing home; HCW- Healthcare Workers Omicron BA.1 spike Avidity Avidity index of the BA.1 spike antibody across different time points among naive and prior infected individuals. Prevax- before primary series; Postvax- 2 weeks after primary series; M6- 6 months after primary series; Postboost- 2 weeks after 1st monovalent booster; M3M6postboost- 3-6months post booster; Postboost 2- 2 weeks after 2nd monovalent booster. NH- Nursing home; HCW- Healthcare Workers Conclusion This study underscores the importance of booster vaccination among NHR and HCWs. The booster dose increases avidity, adding to vaccine-induced antibody functional ability. Higher avidity antibodies have higher cross-reactivity to other SARS-CoV-2 strains, as Wuhan boosting improves Delta and Omicron avidity, and should improve protection from ever-evolving strains. Higher avidities may help explain how the vaccine’s protective effects persist even while antibody titers fade between vaccine doses. Disclosures Stefan Gravenstein, MD, MPH, CDC: Grant/Research Support|Genentech: Advisor/Consultant|Genentech: Grant/Research Support|GSK: Advisor/Consultant|GSK: Honoraria|Janssen: Advisor/Consultant|Janssen: Honoraria|NIH: Grant/Research Support|Pfizer: Grant/Research Support|Pfizer: Honoraria|Sanofi: Advisor/Consultant|Sanofi: Grant/Research Support|Sanofi: Honoraria|Seqirus: Grant/Research Support|Seqirus: Honoraria David Canaday, MD, Pfizer: Grant/Research Support

1726. Characterization of humoral immune responses to COVID-19 vaccination in children in Canada: interim analysis from the Severe acute respiratory syndrome-related coronavirus 2 PRevalence In children and youNG adults in British Columbia (SPRING) Study

Abstract Background A vaccine series consisting of two doses of coronavirus disease 2019 (COVID-19) mRNA vaccines plus one booster dose is currently recommended for healthy children aged 5-11 years in Canada. Additional studies are needed to understand the longitudinal immunity to COVID-19 vaccination in the pediatric population. The Severe acute respiratory syndrome-related coronavirus 2 PRevalence In children and youNG adults in British Columbia (SPRING) sub-study aimed to specifically investigate antibody responses to COVID-19 vaccination in children aged 5-11 years. Methods Fifty-four children without immunocompromising conditions or taking immunosuppressive medications were enrolled prior to dose one or dose two of their COVID-19 vaccine. Responses were quantified via: anti-index virus spike protein specific IgG (S-IgG) geometric mean concentrations (GMCs) (binding antibody units, BAU/mL) and S-IgG avidity reported as total relative IgG avidity index (TRAI) (avidity units, AU) at pre-dose two, as well as one and six months post-dose two. A Welch’s t-test compared responses between timepoints. Results The cohort consisted of 55% male and 45% female with a median age of 8 years. The interval between first and second doses ranged from 7 – 18 (median 11) weeks. Concentrations of S-IgG increased from pre-dose two to one month post-dose two (222 vs. 1152 BAU/mL, p < 0.0001). GMCs then decreased by six months post-dose two (1152 vs. 343 BAU/mL, p < 0.0001) to levels similar to pre-dose two (p = 0.0696). TRAI increased between time points. By six months post-dose two, S-IgG TRAI (68 AU) remained higher than pre-dose two (46 AU, p < 0.0001) and one month post-dose two (65 AU, p = 0.0495). This increase in avidity was due to higher concentrations of medium high, high and very high avidity S-IgG as well as a decrease in very low avidity S-IgG at six months post-dose two. Conclusion Concentrations of S-IgG waned significantly by six months post-dose two. The results suggest that due to repeated exposure to the S-protein, affinity maturation has resulted in the production of higher avidity antibodies post-dose two. This suggests that both S-IgG concentrations and avidity should be considered when determining protection against COVID-19 and need to be further explored to support future booster recommendations. Disclosures Sofia R. Bartlett, PhD, Abbvie: Advisor/Consultant|Abbvie: Grant/Research Support|Cepheid: Advisor/Consultant|Gilead: Advisor/Consultant|Gilead: Grant/Research Support Manish Sadarangani, BM BCh, FRCPC, DPhil, GlaxoSmithKline: Grant/Research Support|Merck: Grant/Research Support|Moderna: Grant/Research Support|Pfizer: Grant/Research Support|Sanofi Pasteur: Grant/Research Support|Seqirus: Grant/Research Support|Symvivo: Grant/Research Support|VBI Vaccines: Grant/Research Support

Kinetics of foot-and-mouth disease vaccine-induced antibody responses in buffaloes (Bubalus bubalis): avidity ELISA as an alternative to the virus neutralization test.

The role of water buffaloes in foot-and-mouth disease (FMD) epidemiology as one of the major hosts of the virus that can develop persistent asymptomatic infection highlights the importance of sustaining surveillance on the antibody response elicited by vaccination in these animals. There is gap in the knowledge on how serological assays that measure antibodies against capsid proteins perform with buffalo samples and which would be the most reliable test to substitute the virus neutralization test (VNT) a cumbersome and low-throughput tool for field surveillance. Alternatively, the liquid-phase blocking sandwich ELISA (LPBE) is commonly used. Previous data from our laboratory demonstrated that the vaccine-induced antibodies assessed by the LPBE yielded low specificity with buffaloes' samples. In contrast, a single-dilution avidity ELISA (AE) aimed to detect high-avidity antibodies against exposed epitopes, combined with an indirect ELISA (IE) to assess IgG levels, produced more reliable results. Here we analyzed for the first time the kinetics of the antibodies induced by vaccination in two different buffalo herds (n = 91) over 120 days using AE, IE, LPBE, and the VNT. Kinetics were similar in the different assays, with an increase of antibodies between 0- and 14-days post-vaccination (dpv) which were maintained thereafter. VNT and AE results were concordant (Kappa value = 0.76), and both assays revealed a decay in the antibody response in calves with maternal antibodies at 90 and 120 dpv, which was not evidenced by the LPBE. These results show that kinetics of antibody responses to FMD vaccination are similar in buffalo and cattle, and support the use of indirect ELISA assays, in particular Avidity ELISA, as alternatives to the VNT for vaccine-immunity monitoring irrespectively of the animal's passive or active immune status.

Influence of antigen density and TLR ligands on preclinical efficacy of a VLP-based vaccine against peanut allergy.

Virus-like particle (VLP) Peanut is a novel immunotherapeutic vaccine candidate for the treatment of peanut allergy. The active pharmaceutical ingredient represents cucumber mosaic VLPs (CuMVTT -VLPs) that are genetically fused with one of the major peanut allergens, Ara h 2 (CuMVTT -Ara h 2). We previously demonstrated the immunogenicity and the protective capacity of VLP Peanut-based immunization in a murine model for peanut allergy. Moreover, a Phase I clinical trial has been initiated using VLP Peanut material manufactured following a GMP-compliant manufacturing process. Key product characterization studies were undertaken here to understand the role and contribution of critical quality attributes that translate as predictive markers of immunogenicity and protective efficacy for clinical vaccine development. The role of prokaryotic RNA encapsulated within VLP Peanut on vaccine immunogenicity was assessed by producing a VLP Peanut batch with a reduced RNA content (VLP Peanut low RNA). Immunogenicity and peanut allergen challenge studies were conducted with VLP Peanut low RNA, as well as with VLP Peanut in WT and TLR 7 KO mice. Furthermore, mass spectrometry and SDS-PAGE based methods were used to determine Ara h 2 antigen density on the surface of VLP Peanut particles. This methodology was subsequently applied to investigate the relationship between Ara h 2 antigen density and immunogenicity of VLP Peanut. A TLR 7 dependent formation of Ara h 2 specific high-avidity IgG antibodies, as well as a TLR 7 dependent change in the dominant IgG subclass, was observed following VLP Peanut vaccination, while total allergen-specific IgG remained relatively unaffected. Consistently, a missing TLR 7 signal caused only a weak decrease in allergen tolerability after vaccination. In contrast, a reduced RNA content for VLP Peanut resulted in diminished total Ara h 2 specific IgG responses, followed by a significant impairment in peanut allergen tolerability. The discrepant effect on allergen tolerance caused by an absent TLR 7 signal versus a reduced RNA content is explained by the observation that VLP Peanut-derived RNA not only stimulates TLR 7 but also TLR 3. Additionally, a strong correlation was observed between the number of Ara h 2 antigens displayed on the surface of VLP Peanut particles and the vaccine's immunogenicity and protective capacity. Our findings demonstrate that prokaryotic RNA encapsulated within VLP Peanut, including antigen density of Ara h 2 on viral particles, are key contributors to the immunogenicity and protective capacity of the vaccine. Thus, antigenicity and RNA content are two critical quality attributes that need to be determined at the stage of manufacturing, providing robust information regarding the immunogenicity and protective capacity of VLP Peanut in the mouse which has translational relevance to the human setting.

Prolonging the delivery of influenza virus vaccine improves the quantity and quality of the induced immune responses in mice.

Influenza vaccines play a vital role in protecting individuals from influenza virus infection and severe illness. However, current influenza vaccines have suboptimal efficacy, which is further reduced in cases where the vaccine strains do not match the circulating strains. One strategy to enhance the efficacy of influenza vaccines is by extended antigen delivery, thereby mimicking the antigen kinetics of a natural infection. Prolonging antigen availability was shown to quantitatively enhance influenza virus-specific immune responses but how it affects the quality of the induced immune response is unknown. Therefore, the current study aimed to investigate whether prolongation of the delivery of influenza vaccine improves the quality of the induced immune responses over that induced by prime-boost immunization. Mice were given daily doses of whole inactivated influenza virus vaccine for periods of 14, 21, or 28 days; the control group received prime-boost immunization with a 28 days interval. Our data show that the highest levels of cellular and humoral immune responses were induced by 28 days of extended antigen delivery, followed by 21, and 14 days of delivery, and prime-boost immunization. Moreover, prolonging vaccine delivery also improved the quality of the induced antibody response, as indicated by higher level of high avidity antibodies, a balanced IgG subclass profile, and a higher level of cross-reactive antibodies. Our findings contribute to a better understanding of the immune response to influenza vaccination and have important implications for the design and development of future slow-release influenza vaccines.

Humoral and cellular immune responses in fully vaccinated individuals with or without SARS-CoV-2 breakthrough infection: Results from the CoV-ADAPT cohort.

Despite recent advances in prophylactic vaccination, SARS-CoV-2 infections continue to cause significant morbidity. A better understanding of immune response differences between vaccinated individuals with and without later SARS-CoV-2 breakthrough infection is urgently needed. CoV-ADAPT is a prospective long-term study comparing humoral (anti-spike-RBD-IgG, neutralization capacity, avidity) and cellular (spike-induced T-cell interferon-γ [IFN-γ] release) immune responses in individuals vaccinated against SARS-CoV-2 at four different time points (three before and one after third vaccination). In this cohort study, 62 fully vaccinated individuals presented with SARS-CoV-2 breakthrough infections vs 151 without infection 3-7 months following third vaccination. Breakthrough infections significantly increased anti-spike-RBD-IgG (p < 0.01), but not spike-directed T-cell IFN-γ release (TC) or antibody avidity. Despite comparable surrogate neutralization indices, the functional neutralization capacity against SARS-CoV-2-assessed via a tissue culture-based assay-was significantly higher following breakthrough vs no breakthrough infection. Anti-spike-RBD-IgG and antibody avidity decreased with age (p < 0.01) and females showed higher anti-spike-RBD-IgG (p < 0.01), and a tendency towards higher antibody avidity (p = 0.051). The association between humoral and cellular immune responses previously reported at various time points was lost in subjects after breakthrough infections (p = 0.807). Finally, a machine-learning approach based on our large immunological dataset (a total of 49 variables) from different time points was unable to predict breakthrough infections (area under the curve: 0.55). In conclusion, distinct differences in humoral vscellular immune responses in fully vaccinated individuals with or without breakthrough infection could be demonstrated. Breakthrough infections predominantly drive the humoral response without boosting the cellular component. Breakthrough infections could not be predicted based on immunological data, which indicates a superior role of environmental factors (e.g., virus exposure) in individualized risk assessment.

Are high avidity antibodies to Plasmodium falciparum antigens preferentially transferred across the placenta of premature and term babies?

IntroductionTransplacental transport of maternal IgG via the neonatal Fc receptor (FcRn) provides babies with passive immunity. Several factors are reported to influence transport, including the avidity of antibodies (Abs) for their cognate antigens. Unfortunately, information on the role of antibody (Ab) avidity is limited. This study investigated if i) antibodies (Abs) with high avidity for 6 Plasmodium falciparum antigens and tetanus toxoid (TTx) were preferentially transferred to premature and term Cameroonian babies and ii) if Ab avidity was increased in babies whose mothers had placental malaria (PM), implicating the involvement of immune complexes. MethodsTotal IgG (mg/ml) and Abs to malarial antigens (AMA1, EBA-175, MSP1-42, MSP2, MSP3, DBL5 of VAR2CSA) and TTx were measured in paired mother-cord samples obtained from premature and term deliveries in Cameroon. Half the women had PM at delivery. Avidity Indices (AIs) were determined by treating antigen-bound-Abs with different molar concentrations of NH4SCN and calculating 50% endpoints. ResultsTotal IgG and antigen-specific Abs increased in cord blood with gestational age; however, AIs did not. AIs in paired maternal-cord blood samples were strongly associated for all antigens (r = 0.77–0.96). However, no significant different in AIs was found between paired mother-cord blood samples for any of the antigens (p values > 0.05). Similarly, Ab avidity was not increased in cord blood of babies whose mothers had PM or hypergammaglobulinemia. DiscussionOverall, there was no evidence that higher avidity Abs to any of the malarial antigens or TTx were preferentially transferred to Cameroonian babies.

Delayed generation of functional virus-specific circulating T follicular helper cells correlates with severe COVID-19

Effective humoral immune responses require well-orchestrated B and T follicular helper (Tfh) cell interactions. Whether these interactions are impaired and associated with COVID-19 disease severity is unclear. Here, longitudinal blood samples across COVID-19 disease severity are analysed. We find that during acute infection SARS-CoV-2-specific circulating Tfh (cTfh) cells expand with disease severity. SARS-CoV-2-specific cTfh cell frequencies correlate with plasmablast frequencies and SARS-CoV-2 antibody titers, avidity and neutralization. Furthermore, cTfh cells but not other memory CD4 T cells, from severe patients better induce plasmablast differentiation and antibody production compared to cTfh cells from mild patients. However, virus-specific cTfh cell development is delayed in patients that display or later develop severe disease compared to those with mild disease, which correlates with delayed induction of high-avidity neutralizing antibodies. Our study suggests that impaired generation of functional virus-specific cTfh cells delays high-quality antibody production at an early stage, potentially enabling progression to severe disease.

Dynamics in maturation of SARS-CoV-2 RBD-specific IgG antibody avidity depending on immunization timeframe and type

The aim is to examine dynamics of avidity maturation of IgG antibodies against SARS-CoV-2 RBD depending on the type of immunization (vaccination or infection), as well as on the duration and frequency of immunization. Materials and methods. The study was performed on two sample cohorts collected at two time points during COVID-19 pandemic. The first cohort (group No. 1) consisted of 87 samples of blood sera obtained from COVID-19 convalescents in the period from March to September 2020. The second cohort included 204 samples obtained in September 2021 from two patient groups. Group No. 2 (n = 64) patients immunized with a full course of Gam-Covid-Vac, group No. 3 (n = 140) COVID-19 convalescent patients and subjects vaccinated with Gam-Covid-Vac (hybrid immunity). Results and conclusion. The dynamics of avidity maturation for SARS-CoV-2 RBD IgG antibodies depending on the method and frequency of immunization, showed that the most effective immunity was formed in COVID-19 convalescent patients and subjects vaccinated with a full course of Gam-Covid-Vac. The hybrid immunity showed not only a significantly higher (compared with groups No. 1 and No. 2) level of IgG antibodies (median 228 BAU/ml vs 75 or 119 BAU/ml, p 0.001), but also a higher level of avidity (IA 90.5% vs 54.5 and 76.6, respectively, p 0.001, 4M urea). In the test for assessing the avidity index with the denaturing agent 8M urea in patients with hybrid immunity, the median level of IA was 25% versus 14.8% and 16% in COVID-19 convalescents and vaccinated subjects (p 0.001), only in 8 patients IA was higher than 50%. While comparing a single infection of COVID-19 with a full course of Gam-Covid-Vac, it was shown that vaccination leads to higher IgG levels (median values in groups 119 and 75 BAU/ml, p 0.001) and to a higher avidity index (median 76.6% vs 54.5%). Thus, the more rapid induction of high-avidity antibodies was in vaccinated individuals at early stages of immunization (up to 4 months), during the period when IgG avidity maturation has not yet been completed. Our results showed that during this period vaccination leads to production of antibodies with avidity index at median level of 82% versus 36% in COVID-19 convalescents at similar time point.