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Over the past century, the field of antibody discovery has undergone significant evolution, excluding the current exploration stage of artificial intelligence‐based antibody generation and the often overlooked non‐animal sourced antibody discovery, which typically requires mature in vitro affinity and the selection of high‐quality antigen formulations. This journey has traversed various stages, from methods involving serum‐based antibody acquisition, the isolation of B cells capable of perpetual antibody production through hybridoma technology, to the in‐depth exploration of genetic material using the phage display system, and the current stage involving diverse single B cell screening techniques. Additionally, the emergence of machine learning has brought impressive scientific and technological breakthroughs across research domains, proving to be a powerful application in the field of antibody discovery. However, each technique comes with its limitations, such as variability and control challenges in serum‐based acquisition, lengthy and difficult hybridoma‐derived antibody development, potential limitations in sequence and epitope diversity due to immunization biases in phage display techniques, and costly single B cell screening. Protein mass spectrometry sequencing, with shorter acquisition time and lower costs, is seen as a shortcut by diagnostic companies, impacting traditional antibody development. In diagnostic antibody development, methodological differences in downstream assays and the impact of constant regions outside the Fv core are often neglected. This paper deeply analyzes challenges, proposing innovative strategies for the next generation of diagnostic antibody development. Aimed at moving closer to the gold standard of antibody discovery, these strategies enhance the competitiveness of diagnostic reagent products. [ABSTRACT FROM AUTHOR]

Introduction: Studies indicate that individuals with chronic conditions and specific baseline characteristics may not mount a robust humoral antibody response to SARS-CoV-2 vaccines. In this paper, we used data from the Texas Coronavirus Antibody REsponse Survey (Texas CARES), a longitudinal state-wide seroprevalence program that has enrolled more than 90,000 participants, to evaluate the role of chronic diseases as the potential risk factors of non-response to SARS-CoV-2 vaccines in a large epidemiologic cohort. Methods: A participant needed to complete an online survey and a blood draw to test for SARS-CoV-2 circulating plasma antibodies at four-time points spaced at least three months apart. Chronic disease predictors of vaccine non-response are evaluated using logistic regression with non-response as the outcome and each chronic disease + age as the predictors. Results: As of April 24, 2023, 18,240 participants met the inclusion criteria; 0.58% (N = 105) of these are non-responders. Adjusting for age, our results show that participants with self-reported immunocompromised status, kidney disease, cancer, and "other" non-specified comorbidity were 15.43, 5.11, 2.59, and 3.13 times more likely to fail to mount a complete response to a vaccine, respectively. Furthermore, having two or more chronic diseases doubled the prevalence of non-response. Conclusion: Consistent with smaller targeted studies, a large epidemiologic cohort bears the same conclusion and demonstrates immunocompromised, cancer, kidney disease, and the number of diseases are associated with vaccine non-response. This study suggests that those individuals, with chronic diseases with the potential to affect their immune system response, may need increased doses or repeated doses of COVID-19 vaccines to develop a protective antibody level. [ABSTRACT FROM AUTHOR]

In this paper, a fractional order HIV/HTLV co-infection model with HIV-specific antibody immune response is established. Two cases are considered: constant control and optimal control. For the constant control system, the existence and uniqueness of the positive solutions are proved, and then the sufficient conditions for the existence and stability of five equilibriums are obtained. For the second case, the Pontryagin's Maximum Principle is used to analyze the optimal control, and the formula of the optimal solution are derived. After that, some numerical simulations are performed to validate the theoretical prediction. Numerical simulations indicate that in the case of HIV/HTLV co-infection, the concentration of CD4+T cells is no longer suitable as an effective reference data for understanding the development process of the disease. On the contrary, the number of HIV virus particles should be used as an important indicator for reference. [ABSTRACT FROM AUTHOR]

Background: Resilience is the capacity of an animal to be minimally affected by disturbances or to rapidly return to its initial state before exposure to a disturbance. Resilient livestock are desired because of their improved health and increased economic profit. Genetic improvement of resilience may also lead to trade-offs with production traits. Recently, resilience indicators based on longitudinal data have been suggested, but they need further evaluation to determine whether they are indeed predictive of improved resilience, such as disease resilience. This study investigated different resilience indicators based on deviations between expected and observed egg production (EP) by exploring their genetic parameters, their possible trade-offs with production traits, and their relationships with antibody traits in chickens. Methods: Egg production in a nucleus breeding herd environment based on 1-week-, 2-week-, or 3-week-intervals of two purebred chicken lines, a white egg-laying (33,825 chickens) and a brown egg-laying line (34,397 chickens), were used to determine deviations between observed EP and expected average batch EP, and between observed EP and expected individual EP. These deviations were used to calculate three types of resilience indicators for two life periods of each individual: natural logarithm-transformed variance (ln(variance)), skewness, and lag-one autocorrelation (autocorrelation) of deviations from 25 to 83 weeks of age and from 83 weeks of age to end of life. Then, we estimated their genetic correlations with EP traits and with two antibody traits. Results: The most promising resilience indicators were those based on 1-week-intervals, as they had the highest heritability estimates (0.02–0.12) and high genetic correlations (above 0.60) with the same resilience indicators based on longer intervals. The three types of resilience indicators differed genetically from each other, which indicates that they possibly capture different aspects of resilience. Genetic correlations of the resilience indicator traits based on 1-week-intervals with EP traits were favorable or zero, which means that trade-off effects were marginal. The resilience indicator traits based on 1-week-intervals also showed no genetic correlations with the antibody traits, which suggests that they are not informative for improved immunity or vice versa in the nucleus environment. Conclusions: This paper gives direction towards the evaluation and implementation of resilience indicators, i.e. to further investigate resilience indicator traits based on 1-week-intervals, in breeding programs for selecting genetically more resilient layer chickens. [ABSTRACT FROM AUTHOR]

In the presence of uncertainty, the optimum obtained based on a nominal identified model can neither provide any performance guarantee nor ensure that critical constraints are satisfied, which is crucial for e.g., bioprocess applications characterized by a high degree of complexity combined with costly experiments. Hence, uncertainty should be considered in the optimization and, furthermore, experiments designed to reduce the uncertainty most important for optimization. Herein, we propose a general framework that combines model-based robust optimization with optimal experiment design. The proposed framework can take advantage of prior knowledge in the form of a mechanistic model structure, and the importance of this is demonstrated by comparing it to more standard black-box models typically employed in learning. Through optimal experiment design, we repeatedly reduce the uncertainty most relevant for optimization so as to maximize the potential for improving the worst-case performance by balancing between exploration and exploitation. This makes the proposed method an efficient model-based robust optimization framework, especially in cases with limited experimental resources. The main part of the paper focuses on the case with modeling uncertainty that can be reduced with the availability of more experimental data. Towards the end of the paper, we consider extending the method to also include inherent uncertainty, such as input uncertainty and unmeasured disturbances. The effectiveness of the method is illustrated through a realistic simulation case study of medium optimization of Chinese hamster ovary cell cultivation in continuous monoclonal antibody production, where the metabolic network consists of 23 extracellular metabolites and 126 reactions. [ABSTRACT FROM AUTHOR]

This study reports on the immunogenicity and biodistribution of H5 hemagglutinin (HA)-based self-amplifying (sa) mRNA vaccines in mice. Four sa-mRNA vaccines encoding either a secreted full-length HA, a secreted HA head domain, a secreted HA stalk domain, or a full-length membrane-anchored HA were investigated. All vaccines elicited an adaptive immune response. However, the full-length HA sa-RNA vaccines demonstrated superior performance compared to head and stalk domain vaccines. The antibody titers positively correlated with the vaccine dose. Cellular immune responses and antigen-specific IgA antibodies in the lungs were also observed. The comparison of the sa-mRNA vaccines encoding the secreted and membrane-anchored full-length HA revealed that anchoring of the HA to the membrane significantly enhanced the antibody and cellular responses. In addition to the injection site, the intramuscularly injected sa-mRNA-LNPs were also detected in the draining lymph nodes, spleen, and to a lesser extent, in the lung, kidney, liver, and heart. [ABSTRACT FROM AUTHOR]

Under the dual pressure of emerging zoonoses and the difficulty in eliminating conventional zoonoses, the strategic management of bird diseases through vaccination represents a highly efficacious approach to disrupting the transmission of zoonotic pathogens to humans. Immunization with a DNA vaccine yielded limited protection against avian pathogen infection. To improve its immunogenicity, the extracellular domain of duck-derived CD40L (designated as dusCD40L) was employed as a bio-adjuvant. Our findings unequivocally established the evolutionary conservation of dusCD40L across avian species. Notably, dusCD40L exhibited a compelling capacity to elicit robust immune responses from both B and T lymphocytes. Furthermore, when employed as an adjuvant, dusCD40L demonstrated a remarkable capacity to significantly augment the titers of neutralizing antibodies and the production of IFNγ elicited by a DNA vaccine encoding the prM-E region of an avian flavivirus, namely, the Tembusu virus (TMUV). Moreover, dusCD40L could strengthen virus clearance of the prM-E DNA vaccine in ducks post-TMUV challenge. This research study presents a highly effective adjuvant for advancing the development of DNA vaccines targeting TMUV in avian hosts. Additionally, it underscores the pivotal role of duCD40L as a potent adjuvant in the context of vaccines designed to combat zoonotic infections in avian species. [ABSTRACT FROM AUTHOR]

The aim of the present research was to evaluate the probiotic activity of Lactobacillus acidophilus in broilers of the ARBOR ACRE strain. The probiotic was administered in drinking water at a rate of 50 ml per 1000 chicks every 7 days during the breeding period. Growth performance, the effect on endogenous flora and on the structure of the gut and the bursa of Fabricius, as well as the immune response, were recorded at 20 and 37 days. We evaluated the effectiveness of Lactobacillus acidophilus added to broiler diets, a zootechnical study (mortality, growth rate, index of growth) and a clinical survey, homogeneity and antibody production. Zootechnical studies revealed that the best growth performances were achieved by the probiotic-treated chickens, with significant differences (p<0.05). which concluded with a positive effect on zootechnical performance compared with the controls. Similarly, the study of the organism's reaction showed a good response from the birds supplemented with Lactobacillus acidophilus in the animal's digestive tract, with interactions with the endogenous flora, compared with the control subjects (P<0.05). Antibody titration showed that the Lactobacillus acidophilus-based diet had a positive effect on antibody levels, resulting in a coverage rate higher than the average indicated by the baseline (1000-2500) and also higher than the control batch antibodies. [ABSTRACT FROM AUTHOR]

The emergence of SARS-CoV-2 in late 2019 initiated a global pandemic, which led to a need for effective therapeutics and diagnostic tools, including virus-specific antibodies. Here, we investigate different antigen preparations to produce SARS-CoV-2-specific and virus-neutralizing antibodies in chickens (n = 3/antigen) and rabbits (n = 2/antigen), exploring, in particular, egg yolk for large-scale production of immunoglobulin Y (IgY). Reactivity profiles of IgY preparations from chicken sera and yolk and rabbit sera were tested in parallel. We compared three types of antigens based on ancestral SARS-CoV-2: an inactivated whole-virus preparation, an S1 spike-protein subunit (S1 antigen) and a receptor-binding domain (RBD antigen, amino acids 319–519) coated on lumazine synthase (LS) particles using SpyCather/SpyTag technology. The RBD antigen proved to be the most efficient immunogen, and the resulting chicken IgY antibodies derived from serum or yolk, displayed strong reactivity with ELISA and indirect immunofluorescence and broad neutralizing activity against SARS-CoV-2 variants, including Omicron BA.1 and BA.5. Preliminary in vivo studies using RBD–lumazine synthase yolk preparations in a hamster model showed that local application was well tolerated and not harmful. However, despite the in vitro neutralizing capacity, this antibody preparation did not show protective effect. Further studies on galenic properties seem to be necessary. The RBD–lumazine antigen proved to be suitable for producing SARS-CoV-2 specific antibodies that can be applied to such therapeutic approaches and as reference reagents for SARS-CoV-2 diagnostics, including virus neutralization assays. [ABSTRACT FROM AUTHOR]

Equid alphaherpesviruses 1 (EHV-1) and 4 (EHV-4) are closely related and both endemic in horses worldwide. Both viruses replicate in the upper respiratory tract, but EHV-1 may additionally lead to abortion and equine herpesvirus myeloencephalopathy (EHM). We focused on antibody responses in horses against the receptor-binding glycoprotein D of EHV-1 (gD1), which shares a 77% amino acid identity with its counterpart in EHV-4 (gD4). Both antigens give rise to cross-reacting antibodies, including neutralizing antibodies. However, immunity against EHV-4 is not considered protective against EHM. While a diagnostic ELISA to discriminate between EHV-1 and EHV-4 infections is available based on type-specific fragments of glycoprotein G (gG1 and gG4, respectively), the type-specific antibody reaction against gD1 has not yet been sufficiently addressed. Starting from the N-terminus of gD1, we developed luciferase immunoprecipitation system (LIPS) assays, using gD1-fragments of increasing size as antigens, i.e. gD1_83 (comprising the first 83 amino acids), gD1_160, gD1_180, and gD1_402 (the full-length molecule). These assays were then used to analyse panels of horse sera from Switzerland (n = 60) and Iceland (n = 50), the latter of which is considered EHV-1 free. We detected only one true negative horse serum from Iceland, whereas all other sera in both panels were seropositive for both gG4 (ELISA) and gD1 (LIPS against gD1_402). In contrast, seropositivity against gG1 was rather rare (35% Swiss sera; 14% Icelandic sera). Therefore, a high percentage of antibodies against gD1 could be attributed to cross-reaction and due to EHV-4 infections. In contrast, the gD1_83 fragment was able to identify sera with type-specific antibodies against gD1. Interestingly, those sera stemmed almost exclusively from vaccinated horses. Although it is uncertain that the N-terminal epitopes of gD1 addressed in this communication are linked to better protection, we suggest that in future vaccine developments, type-common antigens should be avoided, while a broad range of type-specific antigens should be favored. [ABSTRACT FROM AUTHOR]

Arthropod-borne viruses represent a crucial public health threat. Current arboviral serology assays are either labor intensive or incapable of distinguishing closely related viruses, and many zoonotic arboviruses that may transition to humans lack any serologic assays. In this study, we present a programmable phage display platform, ArboScan, that evaluates antibody binding to overlapping peptides that represent the proteomes of 691 human and zoonotic arboviruses. We confirm that ArboScan provides detailed antibody binding information from animal sera, human sera, and an arthropod blood meal. ArboScan identifies distinguishing features of antibody responses based on exposure history in a Colombian cohort of Zika patients. Finally, ArboScan details epitope level information that rapidly identifies candidate epitopes with potential protective significance. ArboScan thus represents a resource for characterizing human and animal arbovirus antibody responses at cohort scale. Arboviruses are transmitted by arthropods and include a number of critical human and zoonotic pathogens. Here the authors report a phage display library, ArboScan, for evaluating antibody binding to the peptidome of 691 human and zoonotic arthropod borne viruses, and use it to profile anti-arbovirus antibodies from diverse samples. [ABSTRACT FROM AUTHOR]

Heterologous prime-boost has broken the protective immune response bottleneck of the COVID-19 vaccines. however, the underlying mechanisms have not been fully elucidated. Here, we investigated antibody responses and explored the response of germinal center (GC) to priming with inactivated vaccines and boosting with heterologous adenoviral-vectored vaccines or homologous inactivated vaccines in mice. Antibody responses were dramatically enhanced by both boosting regimens. Heterologous immunization induced more robust GC activation, characterized by increased Tfh cell populations and enhanced helper function. Additionally, increased B-cell activation and antibody production were observed in a heterologous regimen. Libra-seq was used to compare the differences of S1-, S2- and NTD-specific B cells between homologous and heterologous vaccination, respectively. S2- specific CD19+ B cells presented increased somatic hypermutations (SHMs), which were mainly enriched in plasma cells. Moreover, a heterologous booster dose promoted the clonal expansion of B cells specific to S2 and NTD regions. In conclusion, the functional role of Tfh and B cells following SARS-CoV-2 heterologous vaccination may be important for modulating antibody responses. These findings provide new insights for the development of SARS-CoV- 2 vaccines that induce more robust antibody response. [ABSTRACT FROM AUTHOR]

Notwithstanding prevalence gaps in micronutrients supporting immune functions, the significance of their deficits/supplementation for the efficacy of vaccines is underinvestigated. Thus, the influence of supplementation combining vitamins C and D, zinc, selenium, manganese, and N-acetyl cysteine on immune correlates/surrogates of protection conferred by a quadrivalent influenza vaccine (QIV) in mice was investigated. The supplementation starting 5 days before the first of two QIV injections given 28 days apart increased the serum titres of total and neutralizing IgG against each of four influenza strains from QIV. Accordingly, the frequencies of germinal center B cells, follicular CD4+ T helper (Th) cells, and IL-21-producing Th cells increased in secondary lymphoid organs (SLOs). Additionally, the supplementation improved already increased IgG response to the second QIV injection by augmenting not only neutralizing antibody production, but also IgG2a response, which is important for virus clearance, through favoring Th1 differentiation as indicated by Th1 (IFN-γ)/Th2 (IL-4) signature cytokine level ratio upon QIV restimulation in SLO cell cultures. This most likely partly reflected antioxidant action of the supplement as indicated by splenic redox status analyses. Thus, the study provides a solid scientific background for further research aimed at repurposing the use of this safe and inexpensive micronutrient combination to improve response to the influenza vaccine. [ABSTRACT FROM AUTHOR]

Patients with liver cirrhosis, due to their weakened innate and adaptive immunity, are more prone to frequent and severe vaccine-preventable infections. Moreover, impaired adaptive immunity results in a limited antibody response to vaccines. Despite this suboptimal antibody response, vaccines have proven to be very effective in reducing severe outcomes and deaths in these patients. In the Western world, regulatory authorities and scientific liver societies (e.g., AASLD and EASL) have recommended vaccinations for cirrhotic patients. However, despite these strong recommendations, vaccine coverage remains suboptimal. Improving vaccine effectiveness and safety information, providing comprehensive counseling to patients, fact-checking to combat fake news and disinformation and removing barriers to vaccination for disadvantaged individuals may help overcome the low coverage rate. In view of this, vaccines should be administered early in the course of chronic liver diseases, as their efficacy declines with the increasing severity of the disease. [ABSTRACT FROM AUTHOR]

Evolution of SARS-CoV-2 alters the antigenicity of the immunodominant spike (S) receptor-binding domain and N-terminal domain, undermining the efficacy of vaccines and antibody therapies. To overcome this challenge, we set out to develop a vaccine focusing antibody responses on the highly conserved but metastable S2 subunit, which folds as a spring-loaded fusion machinery. We describe a strategy for prefusion-stabilization and high yield recombinant production of SARS-CoV-2 S2 trimers with native structure and antigenicity. We demonstrate that our design strategy is broadly generalizable to sarbecoviruses, as exemplified with the SARS-CoV-1 (clade 1a) and PRD-0038 (clade 3) S2 subunits. Immunization of mice with a prefusion-stabilized SARS-CoV-2 S2 trimer elicits broadly reactive sarbecovirus antibodies and neutralizing antibody titers of comparable magnitude against Wuhan-Hu-1 and the immune evasive XBB.1.5 variant. Vaccinated mice were protected from weight loss and disease upon challenge with XBB.1.5, providing proof-of-principle for fusion machinery sarbecovirus vaccines. Evolution of viruses undermines the efficacy of vaccines and antibody therapies. To overcome this challenge, authors describe a strategy for prefusion-stabilization to provide proof-of-principle for fusion-machinery sarbecovirus vaccines. [ABSTRACT FROM AUTHOR]

Vaccines generally require T lymphocytes for B-cell activation and immunoglobulin class switching in response to peptide or protein antigens. In the absence of T cells, limited IgG class switch takes place, germinal centers are short-lived, and the B cells lack memory. Here, immunization of mice with liposomes containing 15mer peptides and monophosphoryl lipid A (MPLA) as adjuvant, induced T-cell independent (TI) IgG class switch within three days, as well as germinal center formation. The antibody responses were long-lived, strictly dependent on Toll-like receptor 4 (TLR4) signaling, partly dependent on Bruton's tyrosine kinase (BTK) signal transmission, and independent of signaling through T-cell receptors, MHC class II and inflammasome. The antibody response showed characteristics of both TI type 1 and TI type 2. All IgG subclasses could be boosted months after primary immunization, and the biological function of the secreted antibodies was demonstrated in murine models of allergic anaphylaxis and of bacterial infection. Moreover, antibody responses after immunization with peptide- and MPLA-loaded liposomes could be triggered in neonatal mice and in mice receiving immune-suppressants. This study demonstrates T-cell independent endogenous B-cell memory and recall responses in vivo using a peptide antigen. The stimulation of these antibody responses required a correct and dense assembly and administration of peptide and adjuvant on the surface of liposomes. In the future, TI vaccines may prove beneficial in pathological conditions in which T-cell immunity is compromised through disease or medicines or when rapid, antibody-mediated immune protection is needed. [ABSTRACT FROM AUTHOR]

An appropriately designed pharmacokinetic (PK) assay that is sensitive for antidrug antibody (ADA) impact on relevant exposure is an alternative strategy to understand the neutralizing potential of ADAs. However, guidance on how to develop such PK assays and how to confirm the functional ADA impact on exposure is missing. Here, the PK assay of a T-cell-engaging bispecific antibody, cibisatamab, was developed based on its mechanism of action (MoA). Using critical monoclonal anti-idiotypic (anti-ID) antibody positive controls as ADA surrogates, the impact on exposure was evaluated pre-clinically. In a phase I clinical trial (NCT02324257), initial data suggest that the combination of ADA and PK assays for correlation of the ADA response with cibisatamab exposure. To understand the neutralizing potential of patient-derived ADAs on drug activity, advanced ADA characterization has been performed. Structural binding analysis of ADAs to antibody domains of the drug and its impact on targeting were assessed. For this purpose, relevant patient ADA binding features were identified and compared with the specific monoclonal anti-ID antibody-positive controls. Comparable results of target binding inhibition and similar impacts on exposure suggest that the observed reduction of Cmax and Ctrough levels in patients is caused by the neutralizing potential of ADAs and allows a correlation between ADA response and loss of exposure. Therefore, the described study provides important functional aspects for the development of an appropriately designed PK assay for bispecific antibodies as an alternative option towards understanding the neutralizing ADA impact on exposure. [ABSTRACT FROM AUTHOR]

SARS-CoV-2 infections elicit antibodies against the viral spike (S) and nucleocapsid (N) proteins; COVID-19 vaccines against the S-protein only. The BCG-Corona trial, initiated in March 2020 in SARS-CoV-2-naïve Dutch healthcare workers, captured several epidemic peaks and the introduction of COVID-19 vaccines during the one-year follow-up. We assessed determinants of systemic anti-S1 and anti-N immunoglobulin type G (IgG) responses using trial data. Participants were randomised to BCG or placebo vaccination, reported daily symptoms, SARS-CoV-2 test results, and COVID-19 vaccinations, and donated blood for SARS-CoV-2 serology at two time points. In the 970 participants, anti-S1 geometric mean antibody concentrations (GMCs) were much higher than anti-N GMCs. Anti-S1 GMCs significantly increased with increasing number of immune events (SARS-CoV-2 infection or COVID-19 vaccination): 104.7 international units (IU)/mL, 955.0 IU/mL, and 2290.9 IU/mL for one, two, and three immune events, respectively (p < 0.001). In adjusted multivariable linear regression models, anti-S1 and anti-N log10 concentrations were significantly associated with infection severity, and anti-S1 log10 concentration with COVID-19 vaccine type/dose. In univariable models, anti-N log10 concentration was also significantly associated with acute infection duration, and severity and duration of individual symptoms. Antibody concentrations were not associated with long COVID or long-term loss of smell/taste. [ABSTRACT FROM AUTHOR]

T help (Th), stimulation of toll-like receptors (pathogen-associated molecular patterns, PAMPs), and antigen organization and repetitiveness (pathogen-associated structural patterns, PASPs) were shown numerous times to be important in driving B-cell and antibody responses. In this study, we dissected the individual contributions of these parameters using newly developed "Immune-tag" technology. As model antigens, we used eGFP and the third domain of the dengue virus 1 envelope protein (DV1 EDIII), the major target of virus-neutralizing antibodies. The respective proteins were expressed alone or genetically fused to the N-terminal fragment of the cucumber mosaic virus (CMV) capsid protein—nCMV, rendering the antigens oligomeric. In a step-by-step manner, RNA was attached as a PAMP, and/or a universal Th-cell epitope was genetically added for additional Th. Finally, a PASP was added to the constructs by displaying the antigens highly organized and repetitively on the surface of CMV-derived virus-like particles (CuMV VLPs). Sera from immunized mice demonstrated that each component contributed stepwise to the immunogenicity of both proteins. All components combined in the CuMV VLP platform induced by far the highest antibody responses. In addition, the DV1 EDIII induced high levels of DENV-1-neutralizing antibodies only if displayed on VLPs. Thus, combining multiple cues typically associated with viruses results in optimal antibody responses. [ABSTRACT FROM AUTHOR]

We report on a highly significant, positive association between anthrax vaccination and occurrence of Gulf War Illness (GWI) in 111 Gulf War veterans (42 with GWI and 69 controls). GWI was diagnosed in 47.1% of vaccinated veterans but only in 17.2% of non-vaccinated veterans (Pearson χ2 = 7.08, p = 0.008; odds ratio = 3.947; relative risk = 2.617), with 1.6x higher GWI symptom severity in vaccinated veterans (p = 0.007, F-test in analysis of covariance). Next, we tested the hypothesis that the susceptibility to GWI following anthrax vaccination could be due to inability to make antibodies against the anthrax protective antigen (PA), the key protein contained in the vaccine. Since the first step in initiating antibody production would be the binding of PA peptide fragments (typically 15-amino acid long [15-mer]) to peptide-binding motifs of human leukocyte antigen (HLA) Class II molecules, we assessed the binding-motif affinities of such HLA specific molecules to all linear 15-mer peptide fragments of the anthrax PA. We identified a total of 58 HLA Class II alleles carried by the veterans in our sample and found that, of those, 18 (31%) were present in the vaccinated group that did not develop GWI but were absent from the vaccinated group who developed GWI. Remarkably, in silico analyses revealed very high binding affinities of peptide-binding motifs of those 18 HLA alleles with fragments of anthrax vaccine PA, leading to the successful production of anti-PA antibodies. Conversely, the absence of these protective HLA alleles points to a reduced ability to develop antibodies against PA, thus resulting in harmful PA persistence and development of GWI. [ABSTRACT FROM AUTHOR]

The continuous manufacturing of biologics offers significant advantages in terms of reducing manufacturing costs and increasing capacity, but it is not yet widely implemented by the industry due to major challenges in the automation, scheduling, process monitoring, continued process verification, and real-time control of multiple interconnected processing steps, which must be tightly controlled to produce a safe and efficacious product. The process produces a large amount of data from different sensors, analytical instruments, and offline analyses, requiring organization, storage, and analyses for process monitoring and control without compromising accuracy. We present a case study of a cyber–physical production system (CPPS) for the continuous manufacturing of mAbs that provides an automation infrastructure for data collection and storage in a data historian, along with data management tools that enable real-time analysis of the ongoing process using multivariate algorithms. The CPPS also facilitates process control and provides support in handling deviations at the process level by allowing the continuous train to re-adjust itself via a series of interconnected surge tanks and by recommending corrective actions to the operator. Successful steady-state operation is demonstrated for 55 h with end-to-end process automation and data collection via a range of in-line and at-line sensors. Following this, a series of deviations in the downstream unit operations, including affinity capture chromatography, cation exchange chromatography, and ultrafiltration, are monitored and tracked using multivariate approaches and in-process controls. The system is in line with Industry 4.0 and smart manufacturing concepts and is the first end-to-end CPPS for the continuous manufacturing of mAbs. [ABSTRACT FROM AUTHOR]

Serological testing can be a powerful complementary approach to achieve timely diagnosis of severe acute respiratory coronavirus 2 (SARS-CoV-2) infection, along with nucleic acid detection. Immunoglobulin (Ig) A antibodies are less frequently utilized to detect SARS-CoV-2 infection than IgM and IgG antibodies, even though IgA antibodies play an important role in protective immunity against SARS-CoV-2. This review discusses the differences in kinetics and assay performance between IgA and IgM antibodies and the factors influencing antibody responses. It highlights the potential usefulness of analyzing IgA antibodies for the early detection of SARS-CoV-2 infection. The early appearance of IgA and the high sensitivity of IgA-based immunoassays can aid in diagnosing coronavirus disease 2019. However, because of cross-reactivity, it is important to recognize the only moderate specificity of the early detection of SARS-CoV-2 IgA antibodies against spike antigens. Either the analysis of antibodies targeting the nucleocapsid antigen or a combination of antibodies against the nucleocapsid and spike antigens may strengthen the accuracy of serological evaluation. [ABSTRACT FROM AUTHOR]

Simple Summary: Improving the health of chickens is becoming of increasing importance as consumer expectations of cage-free and free-range housing result in more potential disease challenges while approved antibiotic treatments are decreasing. Antibodies are part of the immune response of chickens. Natural antibodies are present even though the bird has never been exposed to the specific antigen, and adaptive antibodies are produced after vaccination or exposure to a pathogen. This study looks for regions of the genome that influence both natural and adaptive antibody levels in three different breeds of commercially utilized egg production chickens. The genome regions that resulted in differential antibody-level production were identified with a high incidence of immune-related genes. Genetic influence (heritability) for antibody levels was also found. However, the genome regions and the level of heritability were different for different lines and different antigens, suggesting that there are not just a small number of genetic factors that influence overall antibody production. This study provides supporting evidence that overall health of chickens can be improved by genetic selection, though the exact genes and genomic regions still require further investigation, especially considering that the specific region can vary for different antigens and different chicken lines. Selection of livestock for disease resistance is challenging due to the difficulty in obtaining reliable phenotypes. Antibodies are immunological molecules that provide direct and indirect defenses against infection and link the activities of both the innate and adaptive compartments of the immune system. As a result, antibodies have been used as a trait in selection for immune defense. The goal of this study was to identify genomic regions associated with natural and induced antibodies in chickens using low-pass sequencing. Enzyme-linked immunosorbent assays were used to quantify innate (natural) antibodies binding KLH, OVA, and PHA and induced (adaptive) antibodies binding IBD, IBV, NDV, and REO. We collected plasma from four White Leghorn (WL), two White Plymouth Rock (WPR), and two Rhode Island Red (RIR) lines. Samples numbers ranged between 198 and 785 per breed. GWAS was performed within breed on data pre-adjusted for Line-Hatch-Sex effects using GCTA. A threshold of p = 10−6 was used to select genes for downstream annotation and enrichment analysis with SNPEff and Panther. Significant enrichment was found for the defense/immunity protein, immunoglobulin receptor superfamily, and the antimicrobial response protein in RIR; and the immunoglobulin receptor superfamily, defense/immunity protein, and protein modifying enzyme in WL. However, none were present in WPR, but some of the selected SNP were annotated in immune pathways. This study provides new insights regarding the genetics of the antibody response in layer chickens. [ABSTRACT FROM AUTHOR]

Current Influenza virus vaccines primarily induce antibody responses against variable epitopes in hemagglutinin (HA), necessitating frequent updates. However, antibodies against neuraminidase (NA) can also confer protection against influenza, making NA an attractive target for the development of novel vaccines. In this study, we aimed to enhance the immunogenicity of recombinant NA antigens by presenting them multivalently on a nanoparticle carrier. Soluble tetrameric NA antigens of the N1 and N2 subtypes, confirmed to be correctly folded by cryo-electron microscopy structural analysis, were conjugated to Mi3 self-assembling protein nanoparticles using the SpyTag-SpyCatcher system. Immunization of mice with NA-Mi3 nanoparticles induced higher titers of NA-binding and -inhibiting antibodies and improved protection against a lethal challenge compared to unconjugated NA. Additionally, we explored the co-presentation of N1 and N2 antigens on the same Mi3 particles to create a mosaic vaccine candidate. These mosaic nanoparticles elicited antibody titers that were similar or superior to the homotypic nanoparticles and effectively protected against H1N1 and H3N2 challenge viruses. The NA-Mi3 nanoparticles represent a promising vaccine candidate that could complement HA-directed approaches for enhanced potency and broadened protection against influenza A virus. [ABSTRACT FROM AUTHOR]

Background: Snakebite envenomation inflicts a high burden of mortality and morbidity in sub-Saharan Africa. Antivenoms are the mainstay in the therapy of envenomation, and there is an urgent need to develop antivenoms of broad neutralizing efficacy for this region. The venoms used as immunogens to manufacture snake antivenoms are normally selected considering their medical importance and availability. Additionally, their ability to induce antibody responses with high neutralizing capability should be considered, an issue that involves the immunization scheme and the animal species being immunized. Methodology/Principal findings: Using the lethality neutralization assay in mice, we compared the intrageneric neutralization scope of antisera generated by immunization of horses with monospecific, bispecific/monogeneric, and polyspecific/monogeneric immunogens formulated with venoms of Bitis spp., Echis spp., Dendroaspis spp., spitting Naja spp. or non-spitting Naja spp. It was found that the antisera raised by all the immunogens were able to neutralize the homologous venoms and, with a single exception, the heterologous congeneric venoms (considering spitting and non-spitting Naja separately). In general, the polyspecific antisera of Bitis spp, Echis spp, and Dendroaspis spp gave the best neutralization profile against venoms of these genera. For spitting Naja venoms, there were no significant differences in the neutralizing ability between monospecific, bispecific and polyspecific antisera. A similar result was obtained in the case of non-spitting Naja venoms, except that polyspecific antiserum was more effective against the venoms of N. melanoleuca and N. nivea as compared to the monospecific antiserum. Conclusions/Significance: The use of polyspecific immunogens is the best alternative to produce monogeneric antivenoms with wide neutralizing coverage against venoms of sub-Saharan African snakes of the Bitis, Echis, Naja (non-spitting) and Dendroaspis genera. On the other hand, a monospecific immunogen composed of venom of Naja nigricollis is suitable to produce a monogeneric antivenom with wide neutralizing coverage against venoms of spitting Naja spp. These findings can be used in the design of antivenoms of wide neutralizing scope for sub-Saharan Africa. Author summary: Parenteral administration of antivenoms is the core of the current treatment of snakebite envenomations, and there is an urgent need to produce antivenoms of wide neutralizing efficacy for sub-Saharan Africa. The active substance of antivenoms are antibodies (or antibody fragments) purified from plasma of horses or sheep immunized by the repeated injection of snake venoms. Generally, these antibodies can neutralize the venoms used as immunogens and other related venoms. Normally, the venoms used as immunogens are selected considering their medical importance and availability. To complement these criteria with information regarding the immunogenicity of venoms, we compared monospecific, bispecific/monogeneric, and polyspecific/monogeneric antisera towards venoms of Bitis spp., Echis spp., Dendroaspis spp., spitting Naja spp. or non-spitting Naja spp, regarding their intrageneric neutralization scope, evaluated by the lethality neutralization assay in mice. We found that the polyspecific antisera against venoms of Bitis spp, Echis spp, Dendroaspis spp, or non-spitting Naja gave the best neutralization profile. On the other hand, the monospecific, bispecific and polyspecific antisera towards venoms of spitting Naja venoms showed a similar performance. This information suggests that polyspecific immunogens could be the best alternative to produce antivenoms with the widest neutralizing coverage against sub-Saharan African snake venoms. [ABSTRACT FROM AUTHOR]

Introduction: Diversity in malarial antigens is an immune evasion mechanism that gives malaria parasites an edge over the host. Immune responses against one variant of a polymorphic antigen are usually not fully effective against other variants due to altered epitopes. This study aimed to evaluate diversity in the Plasmodium falciparum antigens apical membrane antigen 1 (PfAMA1) and circumsporozoite protein (PfCSP) from circulating parasites in a malaria-endemic community in southern Ghana and to determine the effects of polymorphisms on antibody response specificity. Methods: The study involved 300 subjects, whose P. falciparum infection status was determined by microscopy and PCR. Diversity within the two antigens was evaluated by msp2 gene typing and molecular gene sequencing, while the host plasma levels of antibodies against PfAMA1, PfCSP, and two synthetic 24mer peptides from the conserved central repeat region of PfCSP, were measured by ELISA. Results: Of the 300 subjects, 171 (57%) had P. falciparum infection, with 165 of the 171 (96.5%) being positive for either or both of the msp2 allelic families. Gene sequencing of DNA from 55 clonally infected samples identified a total of 56 non-synonymous single nucleotide polymorphisms (SNPs) for the Pfama1 gene and these resulted in 44 polymorphic positions, including two novel positions (363 and 365). Sequencing of the Pfcsp gene from 69 clonal DNA samples identified 50 non-synonymous SNPs that resulted in 42 polymorphic positions, with half (21) of these polymorphic positions being novel. Of the measured antibodies, only anti-PfCSP antibodies varied considerably between PCR parasite-positive and parasite-negative persons. Discussion: These data confirm the presence of a considerable amount of unique, previously unreported amino acid changes, especially within PfCSP. Drivers for this diversity in the Pfcsp gene do not immediately seem apparent, as immune pressure will be expected to drive a similar level of diversity in the Pfamal gene. [ABSTRACT FROM AUTHOR]

Introduction: Antibodies against the SARS-CoV-2 spike protein are a critical immune determinant for protection against the virus. While virus neutralization is a key function of spike-specific antibodies, antibodies also mediate Fcdependent activities that can play a role in protection or pathogenesis. Methods: This study characterized serum antibody responses elicited after two doses of heterologous adenovirus-vectored (Ad26/Ad5) vaccines. Results: Vaccine-induced antibody binding titers and Fc-mediated functions decreased over six months, while neutralization titers remained stable. Comparison of antibody isotypes elicited after Ad26/Ad5 vs. LNP-mRNA vaccination and after infection showed that anti-spike IgG1 were dominant and produced to high levels in all groups. The Ad26/Ad5 vaccines also induced IgG4 but not IgG2 and IgG3, whereas the LNP-mRNA vaccines elicited a full Ig spectrum (IgM, IgG1-4, IgA1-2). Convalescent COVID-19 patients had mainly IgM and IgA1 alongside IgG1. Despite these differences, the neutralization potencies against early variants were similar. However, both vaccine groups had antibodies with greater Fc potencies of binding complement and Fcg receptors than the COVID-19 group. The Ad26/Ad5 group also displayed a greater potency of RBD-specific antibody-mediated cellular phagocytosis. Discussion: Antibodies with distinctive quality were induced by different vaccines and infection. The data imply the utility of different vaccine platforms to elicit antibody responses with fine-tuned Fc activities. [ABSTRACT FROM AUTHOR]

Coxiella burnetii, the causative agent of Q fever, is a zoonotic bacteria of global public health significance. The organism has a complex, diverse, and relatively poorly understood animal reservoir but there is increasing evidence that macropods play some part in the epidemiology of Q fever in Australia. The aim of this cross-sectional survey was to estimate the animal- and tissue-level prevalence of coxiellosis amongst eastern grey (Macropus giganteus) and red (Osphranter rufus) kangaroos co-grazing with domestic cattle in a Q fever endemic area in Queensland. Serum, faeces and tissue samples from a range of organs were collected from 50 kangaroos. A total of 537 tissue samples were tested by real-time PCR, of which 99 specimens from 42 kangaroos (84% of animals, 95% confidence interval [CI], 71% to 93%) were positive for the C. burnetii IS1111 gene when tested in duplicate. Twenty of these specimens from 16 kangaroos (32%, 95% CI 20% to 47%) were also positive for the com1 or htpAB genes. Serum antibodies were present in 24 (57%, 95% CI 41% to 72%) of the PCR positive animals. There was no statistically significant difference in PCR positivity between organs and no single sample type consistently identified C. burnetii positive kangaroos. The results from this study identify a high apparent prevalence of C. burnetii amongst macropods in the study area, albeit seemingly with an inconsistent distribution within tissues and in relatively small quantities, often verging on the limits of detection. We recommend Q fever surveillance in macropods should involve a combination of serosurveys and molecular testing to increase chances of detection in a population, noting that a range of tissues would likely need to be sampled to confirm the diagnosis in a suspect positive animal. [ABSTRACT FROM AUTHOR]

A dynamic of virus adaptation and a mass vaccination campaign could significantly reduce the severity of clinical manifestations of COVID-19 and transmission. Hence, COVID-19 may become an endemic disease globally. Moreover, mass infection as the COVID-19 pandemic progressed affected the serology of the patients as a result of virus mutation and vaccination. Therefore, a need exists to acquire accurate serological testing to monitor the emergence of new outbreaks of COVID-19 to promptly prevent and control the disease spreading. In this study, the anti-Orf8 antibodies among samples collected in Thailand's first, fourth, and fifth waves of COVID-19 outbreaks compared with pre-epidemic sera were determined by indirect ELISA. The diagnostic sensitivity and specificity of the anti-Orf8 IgG ELISA for COVID-19 samples from the first, fourth, and fifth waves of outbreaks was found to be 100% compared with pre-epidemic sera. However, the diagnostic sensitivity and specificity of the anti-Orf8 IgG ELISA for a larger number of patient samples and controls from the fifth wave of outbreaks which were collected on day 7 and 14 after an RT-PCR positive result were 58.79 and 58.44% and 89.19 and 58.44%, respectively. Our data indicated that some of the controls might have antibodies from natural past infections. Our study highlighted the potential utility of anti-Orf8 IgG antibody testing for seroprevalence surveys but still warrants further investigations. [ABSTRACT FROM AUTHOR]

Plasma RNAemia, delayed antibody responses and inflammation predict COVID-19 outcomes, but the mechanisms underlying these immunovirological patterns are poorly understood. We profile 782 longitudinal plasma samples from 318 hospitalized patients with COVID-19. Integrated analysis using k-means reveals four patient clusters in a discovery cohort: mechanically ventilated critically-ill cases are subdivided into good prognosis and high-fatality clusters (reproduced in a validation cohort), while non-critical survivors segregate into high and low early antibody responders. Only the high-fatality cluster is enriched for transcriptomic signatures associated with COVID-19 severity, and each cluster has distinct RBD-specific antibody elicitation kinetics. Both critical and non-critical clusters with delayed antibody responses exhibit sustained IFN signatures, which negatively correlate with contemporaneous RBD-specific IgG levels and absolute SARS-CoV-2-specific B and CD4+ T cell frequencies. These data suggest that the "Interferon paradox" previously described in murine LCMV models is operative in COVID-19, with excessive IFN signaling delaying development of adaptive virus-specific immunity. The role of IFN signaling in SARS-CoV-2 infection and outcome is still debated. Here, the authors longitudinally profiled plasma samples from hospitalized patients and show that a persistent inflammatory response is linked to delayed generation of adaptive immunity and increased risk of death when coupled with severe infection. [ABSTRACT FROM AUTHOR]

Public antibody responses have been found against many infectious agents. Structural convergence of public antibodies is usually determined by immunoglobulin V genes. Recently, a human antibody public class against SARS-CoV-2 was reported, where the D gene (IGHD3-22) encodes a common YYDxxG motif in heavy-chain complementarity-determining region 3 (CDR H3), which determines specificity for the receptor-binding domain (RBD). In this review, we discuss the isolation, structural characterization, and genetic analyses of this class of antibodies, which have been isolated from various cohorts of COVID-19 convalescents and vaccinees. All eleven YYDxxG antibodies with available structures target the SARS-CoV-2 RBD in a similar binding mode, where the CDR H3 dominates the interaction with antigen. The antibodies target a conserved site on the RBD that does not overlap with the receptor-binding site, but their particular angle of approach results in direct steric hindrance to receptor binding, which enables both neutralization potency and breadth. We also review the properties of CDR H3-dominant antibodies that target other human viruses. Overall, unlike most public antibodies, which are identified by their V gene usage, this newly discovered public class of YYDxxG antibodies is dominated by a D-gene-encoded motif and uncovers further opportunities for germline-targeting vaccine design. [ABSTRACT FROM AUTHOR]

A natural monkeypox virus infection may not induce sufficient neutralizing antibody responses in a subset of healthy individuals. The aim of this study was to evaluate monkeypox virus-neutralizing antibodies six months after infection and to assess the virological factors predictive of a poor immunological response. Antibodies were assessed using a plaque reduction neutralization test at six months from mpox infection; mpox cutaneous, oropharyngeal, and anal swabs, semen, and plasma samples were tested during infection. Overall, 95 people were included in the study; all developed detectable antibodies. People who were positive for the monkeypox virus for more days had higher levels of antibodies when considering all tested samples (p = 0.029) and all swabs (p = 0.005). Mpox cycle threshold values were not predictive of antibody titers. This study found that the overall days of monkeypox virus detection in the body, irrespective of the viral loads, were directly correlated with monkeypox virus neutralizing antibodies at six months after infection. [ABSTRACT FROM AUTHOR]

Background: This study aimed to assess the changes in anti‐spike protein IgG antibody titer over time following mRNA vaccination (BNT162b2) against severe acute respiratory syndrome coronavirus 2. Methods: We monitored IgG levels in 23 medical care workers (MCWs) for up to 3 months after administering the third dose of BNT162b2. Blood samples were periodically collected from all participants. Results: Following the third dose, the median antibody titer increased to 252 and 327% compared with antibody levels at 1 and 3 months after the second dose, respectively. Additionally, compared with 1 month after the second dose, the median antibody titer decreased to 30.2 and 9.8% at 3 and 6 months, respectively, and to 39.1% at 3 months, compared with 1 month after the third dose. Conclusion: Antibody levels declined quickly after the second dose but declined more slowly after the third dose, showing a booster effect. This study provides insights into the immunogenicity of booster doses and time intervals for booster vaccination strategies. [ABSTRACT FROM AUTHOR]

The objective of the current investigation was to evaluate the induction of heat shock proteins (HSPs) in SP2/0 transgenic cells and the effect of these proteins on the production of monoclonal antibodies (mAbs). The SP2/0 cell line expressing the PSG-026 antibody, a biosimilar candidate of golimumab, the culture parameters, and the target protein expression were not justified for industrial production and were used for the experiments. Paracetamol and heat shock were used as chemical and physical inducers of HSPs, respectively. The results showed that paracetamol and heat shock increased the expression of HSP70 and HSP27 at the mRNA and protein levels. The expression of HSPs was greater in paracetamol-treated cells than in heat shock-treated cells. Paracetamol treatment at concentrations above 0.5 mM significantly reduced cell viability and mAb expression. However, treatment with 0.25 mM paracetamol results in delayed cell death and increased mAb production. Heat shock treatment at 45°C for 30 minutes after enhanced mAb expression was applied after pre-treatment with paracetamol. In bioreactor cultures, pretreatment of cells with paracetamol improved cell viability and shortened the lag phase, resulting in increased cell density. The production of mAbs in paracetamol-treated cultures was markedly greater than that in the control. Analysis of protein quality and charge variants revealed no significant differences between paracetamol-treated and control cultures, indicating that the induction of HSPs did not affect protein aggregation or charge variants. These findings suggest that inducing and manipulating HSP expression can be a valuable strategy for improving recombinant protein production in biopharmaceutical processes. [ABSTRACT FROM AUTHOR]

Background: Priming with ChAdOx1 followed by heterologous boosting is considered in several countries. Nevertheless, analyses comparing the immunogenicity of heterologous booster to homologous primary vaccination regimens and natural infection are lacking. In this study, we aimed to conduct a comparative assessment of the immunogenicity between homologous primary vaccination regimens and heterologous prime‐boost vaccination using BNT162b2 or mRNA‐1273. Methods: We matched vaccinated naïve (VN) individuals (n = 673) with partial vaccination (n = 64), primary vaccination (n = 590), and primary series plus mRNA vaccine heterologous booster (n = 19) with unvaccinated naturally infected (NI) individuals with a documented primary SARS‐CoV‐2 infection (n = 206). We measured the levels of neutralizing total antibodies (NTAbs), total antibodies (TAbs), anti‐S‐RBD IgG, and anti‐S1 IgA titers. Results: Homologous primary vaccination with ChAdOx1 not only showed less potent NTAb, TAb, anti‐S‐RBD IgG, and anti‐S1 IgA immune responses compared to primary BNT162b2 or mRNA‐1273 vaccination regimens (p < 0.05) but also showed ~3‐fold less anti‐S1 IgA response compared to infection‐induced immunity (p < 0.001). Nevertheless, a heterologous booster led to an increase of ~12 times in the immune response when compared to two consecutive homologous ChAdOx1 immunizations. Furthermore, correlation analyses revealed that both anti‐S‐RBD IgG and anti‐S1 IgA significantly contributed to virus neutralization among NI individuals, particularly in symptomatic and pauci‐symptomatic individuals, whereas among VN individuals, anti‐S‐RBD IgG was the main contributor to virus neutralization. Conclusion: The results emphasize the potential benefit of using heterologous mRNA boosters to increase antibody levels and neutralizing capacity particularly in patients who received primary vaccination with ChAdOx1. [ABSTRACT FROM AUTHOR]

The immune escape of Omicron variants significantly subsides by the third dose of an mRNA vaccine. However, it is unclear how Omicron variant-neutralizing antibodies develop under repeated vaccination. We analyze blood samples from 41 BNT162b2 vaccinees following the course of three injections and analyze their B-cell receptor (BCR) repertoires at six time points in total. The concomitant reactivity to both ancestral and Omicron receptor-binding domain (RBD) is achieved by a limited number of BCR clonotypes depending on the accumulation of somatic hypermutation (SHM) after the third dose. Our findings suggest that SHM accumulation in the BCR space to broaden its specificity for unseen antigens is a counterprotective mechanism against virus variant immune escape. Repeat vaccination with COVID-19 mRNA vaccines has been shown to increase breadth of the antibody response. Here the authors demonstrate that B cell clones induced by the ancestral COVID-19 vaccine develop into daughter clones with different reactivity to individual SARS-CoV-2 variants through the accumulation of somatic hypermutations. [ABSTRACT FROM AUTHOR]

While poly(ethylene glycol) (PEG) hydrogels are generally regarded as biologically inert blank slates, concerns over PEG immunogenicity are growing, and the implications for tissue engineering are unknown. Here, we investigate these implications by immunizing mice against PEG to stimulate anti-PEG antibody production and evaluating bone defect regeneration after treatment with bone morphogenetic protein-2-loaded PEG hydrogels. Quantitative analysis reveals that PEG sensitization increases bone formation compared to naive controls, whereas histological analysis shows that PEG sensitization induces an abnormally porous bone morphology at the defect site, particularly in males. Furthermore, immune cell recruitment is higher in PEG-sensitized mice administered the PEG-based treatment than their naive counterparts. Interestingly, naive controls that were administered a PEG-based treatment also develop anti-PEG antibodies. Sex differences in bone formation and immune cell recruitment are also apparent. Overall, these findings indicate that anti-PEG immune responses can impact tissue engineering efficacy and highlight the need for further investigation. Concerns over the immunogenicity of poly(ethylene glycol) (PEG) are growing, and the implications for tissue engineering are unknown. Here the authors evaluate the impact of anti-PEG antibodies and PEG immunogenicity on the efficacy of a PEG hydrogel-based tissue engineering therapy. [ABSTRACT FROM AUTHOR]

Efficient precision vaccines against several highly pathogenic zoonotic viruses are currently lacking. Proteolytic activation is instrumental for a number of these viruses to gain host-cell entry and develop infectivity. For SARS-CoV-2, this process is enhanced by the insertion of a furin cleavage site at the junction of the spike protein S1/S2 subunits upstream of the metalloprotease TMPRSS2 common proteolytic site. Here, we describe a new approach based on specific epitopes selection from the region involved in proteolytic activation and infectivity for the engineering of precision candidate vaccinating antigens. This approach was developed through its application to the design of SARS-CoV-2 cross-variant candidates vaccinating antigens. It includes an in silico structural analysis of the viral region involved in infectivity, the identification of conserved immunogenic epitopes and the selection of those eliciting specific immune responses in infected people. The following step consists of engineering vaccinating antigens that carry the selected epitopes and mimic their 3D native structure. Using this approach, we demonstrated through a Covid-19 patient-centered study of a 500 patients' cohort, that the epitopes selected from SARS-CoV-2 protein S1/S2 junction elicited a neutralizing antibody response significantly associated with mild and asymptomatic COVID-19 (p<0.001), which strongly suggests protective immunity. Engineered antigens containing the SARS-CoV-2 selected epitopes and mimicking the native epitopes 3D structure generated neutralizing antibody response in mice. Our data show the potential of this combined computational and experimental approach for designing precision vaccines against viruses whose pathogenicity is contingent upon proteolytic activation. [ABSTRACT FROM AUTHOR]

Background: The exposure to parasites may influence the immune response to vaccines in endemic African countries. In this study, we aimed to assess the association between helminth exposure to the most prevalent parasitic infections, schistosomiasis, soil transmitted helminths infection and filariasis, and the Ebola virus glycoprotein (EBOV GP) antibody concentration in response to vaccination with the Ad26.ZEBOV, MVA-BN-Filo vaccine regimen in African and European participants using samples obtained from three international clinical trials. Methods/Principal findings: We conducted a study in a subset of participants in the EBL2001, EBL2002 and EBL3001 clinical trials that evaluated the Ad26.ZEBOV, MVA-BN-Filo vaccine regimen against EVD in children, adolescents and adults from the United Kingdom, France, Burkina Faso, Cote d'Ivoire, Kenya, Uganda and Sierra Leone. Immune markers of helminth exposure at baseline were evaluated by ELISA with three commercial kits which detect IgG antibodies against schistosome, filarial and Strongyloides antigens. Luminex technology was used to measure inflammatory and activation markers, and Th1/Th2/Th17 cytokines at baseline. The association between binding IgG antibodies specific to EBOV GP (measured on day 21 post-dose 2 and on Day 365 after the first dose respectively), and helminth exposure at baseline was evaluated using a multivariable linear regression model adjusted for age and study group. Seventy-eight (21.3%) of the 367 participants included in the study had at least one helminth positive ELISA test at baseline, with differences of prevalence between studies and an increased prevalence with age. The most frequently detected antibodies were those to Schistosoma mansoni (10.9%), followed by Acanthocheilonema viteae (9%) and then Strongyloides ratti (7.9%). Among the 41 immunological analytes tested, five were significantly (p <.003) lower in participants with at least one positive helminth ELISA test result: CCL2/MCP1, FGFbasic, IL-7, IL-13 and CCL11/Eotaxin compared to participants with negative helminth ELISA tests. No significant association was found with EBOV-GP specific antibody concentration at 21 days post-dose 2, or at 365 days post-dose 1, adjusted for age group, study, and the presence of any helminth antibodies at baseline. Conclusions/Significance: No clear association was found between immune markers of helminth exposure as measured by ELISA and post-vaccination response to the Ebola Ad26.ZEBOV/ MVA-BN-Filo vaccine regimen. Trial registration: NCT02416453, NCT02564523, NCT02509494. ClinicalTrials.gov. Author summary: Recurrent exposure to parasites may influence the immune response to vaccines, especially in endemic African countries. In this study we aimed to assess the association between immune markers of helminth exposure and the immune response post-vaccination with the Ad26.ZEBOV, MVA-BN-Filo Ebola vaccine regimen in African and European participants who participated in three international clinical trials. Seventy-eight (21.3%) of the 367 participants included in the study, had at least one helminth ELISA positive test at baseline with differences of prevalence between studies and an increased prevalence with increasing age. After adjustment for confounding factors, the study did not show a clear association between immunological markers of helminth exposure and the antibody concentration in response to the Ebola vaccine regimen. [ABSTRACT FROM AUTHOR]

Bacteriophage therapy is a promising approach to address antimicrobial infections though questions remain regarding the impact of the immune response on clinical effectiveness. Here, we develop a mouse model to assess phage treatment using a cocktail of five phages from the Myoviridae and Siphoviridae families that target Vancomycin-Resistant Enterococcus gut colonization. Phage treatment significantly reduces fecal bacterial loads of Vancomycin-Resistant Enterococcus. We also characterize immune responses elicited following administration of the phage cocktail. While minimal innate responses are observed after phage administration, two rounds of treatment induces phage-specific neutralizing antibodies and accelerate phage clearance from tissues. Interestingly, the myophages in our cocktail induce a more robust neutralizing antibody response than the siphophages. This anti-phage immunity reduces the effectiveness of the phage cocktail in our murine model. Collectively, this study shows phage-specific immune responses may be an important consideration in the development of phage cocktails for therapeutic use. Bacteriophage can be used to target bacterial infection and used as a therapeutic approach for antibiotic resistant bacteria. Here the authors show that the use of bacteriophage to target antibiotic resistant bacteria in a mouse model of infection can induce phage specific immune responses and may impair their therapeutic efficacy. [ABSTRACT FROM AUTHOR]

Introduction: Following the coronavirus disease 19 (COVID-19), caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection, vaccination became the main strategy against disease severity and even death. Healthcare workers were considered high-risk for infection and, thus, were prioritised for vaccination. Methods: A follow-up to a SARS-CoV-2 seroprevalence study among clinical and non-clinical HCWs at the Aga Khan University Hospital, Nairobi, we assessed how vaccination influenced SARS-CoV-2 anti-spike IgG antibody responses and kinetics. Blood samples were drawn at two points spanning 6 to 18 months post-vaccination, and SARS-CoV-2 spike antibody levels were determined by enzyme-linked immunosorbent assay. Results: Almost all participants, 98% (961/981), received a second vaccine dose, and only 8.5% (83/981) received a third dose. SARS-CoV-2 spike IgG antibodies were detected in 100% (961/961) and 92.7% (707/762) of participants who received two vaccine doses, with the first and second post-vaccine test, respectively, and in 100% (83/83) and 91.4% (64/70) of those who received three vaccine doses at the first and second post-vaccine test, respectively. Seventy-six participants developed mild infections, not requiring hospitalisation even after receiving primary vaccination. Receiving three vaccine doses influenced the anti-spike S/Co at both the first (p<0.001) and second post-vaccination testing (p<0.001). Of those who tested SARS-CoV-2 positive, the anti-spike S/Co ratio was significantly higher than those who were seronegative at the first post-vaccine test (p = 0.001). Side effects were reported by almost half of those who received the first dose, 47.3% (464/981), 28.9% (278/961) and 25.3% (21/83) of those who received the second and third vaccine doses, respectively. Discussion and conclusion: Following the second dose of primary vaccination, all participants had detectable anti-spike antibodies. The observed mild breakthrough infections may have been due to emerging SARS-CoV-2 variants. Findings suggest that although protective antibodies are induced, vaccination protected against COVID-19 disease severity and not necessarily infection. [ABSTRACT FROM AUTHOR]

Post vaccine immunity following COVID-19 mRNA vaccination may be driven by extrinsic, or controllable and intrinsic, or inherent health factors. Thus, we investigated the effects of extrinsic and intrinsic on the peak antibody response following COVID-19 primary vaccination and on the trajectory of peak antibody magnitude and durability over time. Participants in a longitudinal cohort attended visits every 3 months for up to 2 years following enrollment. At baseline, participants provided information on their demographics, recreational behaviors, and comorbid health conditions which guided our model selection process. Blood samples were collected for serum processing and spike antibody testing at each visit. Cross-sectional and longitudinal models (linear-mixed effects models) were generated to assess the relationship between selected intrinsic and extrinsic health factors on peak antibody following vaccination and to determine the influence of these predictors on antibody over time. Following cross-sectional analysis, we observed higher peak antibody titers after primary vaccination in females, those who reported recreational drug use, younger age, and prior COVID-19 history. Following booster vaccination, females and Hispanics had higher peak titers after the 3rd and 4th doses, respectively. Longitudinal models demonstrated that Moderna mRNA-1273 recipients, females, and those previously vaccinated had increased peak titers over time. Moreover, drug users and half-dose Moderna mRNA-1273 recipients had higher peak antibody titers over time following the first booster, while no predictive factors significantly affected post-second booster antibody responses. Overall, both intrinsic and extrinsic health factors play a significant role in shaping humoral immunogenicity after initial vaccination and the first booster. The absence of predictive factors for second booster immunogenicity suggests a more robust and consistent immune response after the second booster vaccine administration. [ABSTRACT FROM AUTHOR]

Chitosan, a natural polysaccharide derived from chitin, possesses biocompatibility, biodegradability, and mucoadhesive characteristics, making it an attractive material for the delivery of mRNA payloads to the nasal mucosa and promoting their uptake by target cells such as epithelial and immune cells (e.g., dendritic cells and macrophages). In this project, we aimed at developing novel lipid-based nanoformulations for mRNA delivery to counteract the pandemic caused by SARS-CoV-2 virus. The formulations achieved a mRNA encapsulation efficiency of ~80.2% with chitosan-lipid nanoparticles, as measured by the RiboGreen assay. Furthermore, the evaluation of SARS-CoV-2 Spike (S) receptor-binding domain (RBD) expression via ELISA for our vaccine formulations showed transfection levels in human embryonic kidney cells (HEK 293), lung carcinoma cells (A549), and dendritic cells (DC 2.4) equal to 9.9 ± 0.1 ng/mL (174.7 ± 1.1 fold change from untreated cells (UT)), 7.0 ± 0.2 ng/mL (128.1 ± 4.9 fold change from UT), and 0.9 ± 0.0 ng/mL (18.0 ± 0.1 fold change from UT), respectively. Our most promising vaccine formulation was also demonstrated to be amenable to lyophilization with minimal degradation of loaded mRNA, paving the way towards a more accessible and stable vaccine. Preliminary in vivo studies in mice were performed to assess the systemic and local immune responses. Nasal bronchoalveolar lavage fluid (BALF) wash showed that utilizing the optimized formulation resulted in local antibody concentrations and did not trigger any systemic antibody response. However, if further improved and developed, it could potentially contribute to the management of COVID-19 through nasopharyngeal immunization strategies. [ABSTRACT FROM AUTHOR]

Knowledge of the antibody response to the third dose of inactivated SARS-CoV-2 vaccines is crucial because it is the subject of one of the largest global vaccination programs. This study integrated microsampling with optical biosensors to profile neutralizing antibodies (NAbs) in fifteen vaccinated healthy donors, followed by the application of machine learning to predict antibody response at given timepoints. Over a nine-month duration, microsampling and venipuncture were conducted at seven individual timepoints. A refined iteration of a fiber optic biolayer interferometry (FO-BLI) biosensor was designed, enabling rapid multiplexed biosensing of the NAbs of both wild-type and Omicron SARS-CoV-2 variants in minutes. Findings revealed a strong correlation (Pearson r of 0.919, specificity of 100%) between wild-type variant NAb levels in microsamples and sera. Following the third dose, sera NAb levels of the wild-type variant increased 2.9-fold after seven days and 3.3-fold within a month, subsequently waning and becoming undetectable after three months. Considerable but incomplete evasion of the latest Omicron subvariants from booster vaccine-elicited NAbs was confirmed, although a higher number of binding antibodies (BAbs) was identified by another rapid FO-BLI biosensor in minutes. Significantly, FO-BLI highly correlated with a pseudovirus neutralization assay in identifying neutralizing capacities (Pearson r of 0.983). Additionally, machine learning demonstrated exceptional accuracy in predicting antibody levels, with an error level of <5% for both NAbs and BAbs across multiple timepoints. Microsample-driven biosensing enables individuals to access their results within hours of self-collection, while precise models could guide personalized vaccination strategies. The technology's innate adaptability means it has the potential for effective translation in disease prevention and vaccine development. [ABSTRACT FROM AUTHOR]

Studying the immunogenicity of insulin is crucial for managing and treating diabetes and other medical conditions. Although insulin therapy is proven effective for diabetic patients, the presence of insulin antibodies can diminish its effectiveness and lead to adverse immune reactions. Understanding the processes and factors affecting insulin immunogenicity is important to improve treatment outcomes and minimize complications. Research indicates that insulin's immunogenicity is influenced by various structural characteristics such as size, charge, and hydrophobicity. Additionally, factors like insulin dosage, frequency of administration, patient age, gender, genetic background, and mode of insulin delivery contribute to the likelihood of antibody formation. Therefore, optimizing insulin therapy for diabetic patients requires a comprehensive knowledge of insulin immunogenicity and effective insulin antibody detection assays. The development of advanced point-of-care immunoassays has enabled efficient screening for anti-insulin antibodies and assessment of insulin immunogenicity. [ABSTRACT FROM AUTHOR]

Under pressure from neutralising antibodies induced by vaccination or infection the SARS-CoV-2 spike gene has become a hotspot for evolutionary change, leading to the failure of all mAbs developed for clinical use. Most potent antibodies bind to the receptor binding domain which has become heavily mutated. Here we study responses to a conserved epitope in sub-domain-1 (SD1) of spike which have become more prominent because of mutational escape from antibodies directed to the receptor binding domain. Some SD1 reactive mAbs show potent and broad neutralization of SARS-CoV-2 variants. We structurally map the dominant SD1 epitope and provide a mechanism of action by blocking interaction with ACE2. Mutations in SD1 have not been sustained to date, but one, E554K, leads to escape from mAbs. This mutation has now emerged in several sublineages including BA.2.86, reflecting selection pressure on the virus exerted by the increasing prominence of the anti-SD1 response. Due to the focus of vaccination on the SARS CoV-2 spike protein, spike has been associated with high levels of viral mutation and subsequent immune escape. Here the authors study a conserved epitope in SARS CoV-2 sub-domain-1 and characterise the neutralising antibody response and evasion in contemporary SARS COV-2 viral strains. [ABSTRACT FROM AUTHOR]

Background: Babesiosis is a worldwide emerging protozoan infection that is associated with a spectrum of disease severity from asymptomatic infection to severe organ damage and death. While effective treatment strategies are available, some immunocompromised patients experience severe acute and prolonged/relapsing illness due in part to an impaired host antibody response. Intravenous immunoglobulin (IVIG) has been used as an adjunctive therapy in some immunocompromised babesiosis patients, but its therapeutic effect is uncertain. We evaluated the presence of Babesia microti antibodies in commercial samples of IVIG. Methods/principle findings: The presence of B. microti antibodies in commercial samples of IVIG were tested using an immunofluorescence assay. A subset of samples was then tested for B. microti antibodies using an enzyme linked immunosorbent assay. Out of 57 commercial IVIG samples tested using IFA, and 52 samples tested using ELISA, none were positive for B. microti antibodies. Conclusions: Commercially available IVIG may not be of therapeutic benefit for babesiosis patients. Additional sampling of IVIG for B. microti antibody and a clinical trial of babesiosis patients given IVIG compared with controls would provide further insight into the use of IVIG for the treatment of babesiosis. Author summary: Human babesiosis is usually a mild to moderate illness that clears with a standard course of atovaquone and azithromycin. The disease is often severe in immunocompromised patients, including those over 50 years of age, those with asplenia, malignancy, HIV/AIDS, or on immunosuppressive therapy. Patients with multiple immunosuppressive conditions that usually result in low or absent antibody production may experience severe acute disease followed by persistent relapsing disease that can last for months or even years. Intravenous immunoglobulin (IVIG) has been used as adjunct therapy in patients with severe babesiosis, although controlled therapeutic trials have not been carried out. This prompted us to evaluate Babesia microti antibody concentrations in IVIG samples from different commercial sources. We tested commercially available IVIG for antibodies against B. microti using IFA and ELISA assays. None of the IVIG samples contained detectable antibody against B. microti. Our results suggest that commercially available IVIG may be of limited therapeutic benefit for babesiosis patients. [ABSTRACT FROM AUTHOR]

Sustained Notch2 signals induce trans-differentiation of Follicular B (FoB) cells into Marginal Zone B (MZB) cells in mice, but the physiology underlying this differentiation pathway is still elusive. Here, we demonstrate that most B cells receive a basal Notch signal, which is intensified in pre-MZB and MZB cells. Ablation or constitutive activation of Notch2 upon T-cell-dependent immunization reveals an interplay between antigen-induced activation and Notch2 signaling, in which FoB cells that turn off Notch2 signaling enter germinal centers (GC), while high Notch2 signaling leads to generation of MZB cells or to initiation of plasmablast differentiation. Notch2 signaling is dispensable for GC dynamics but appears to be re-induced in some centrocytes to govern expansion of IgG1+ GCB cells. Mathematical modelling suggests that antigen-activated FoB cells make a Notch2 dependent binary fate-decision to differentiate into either GCB or MZB cells. This bifurcation might serve as a mechanism to archive antigen-specific clones into functionally and spatially diverse B cell states to generate robust antibody and memory responses. Sustained exogenous Notch2 signaling prompts Follicular B cells to trans-differentiate into Marginal Zone B cells. This study reveals that under physiological conditions, Notch2 signalling regulates a fate choice in antigen activated Follicular B cells, dictating whether they develop into Germinal Center B cells or Marginal Zone B cells. [ABSTRACT FROM AUTHOR]

Severe heterogeneity within glioblastoma has spurred the notion that disrupting the interplay between multiple elements on immunosuppression is at the core of meaningful anti-tumor responses. T cell immunoreceptor with Ig and ITIM domains (TIGIT) and its glioblastoma-associated antigen, CD155, form a highly immunosuppressive axis in glioblastoma and other solid tumors, yet targeting of TIGIT, a functionally heterogeneous receptor on tumor-infiltrating immune cells, has largely been ineffective as monotherapy, suggesting that disruption of its inhibitory network might be necessary for measurable responses. It is within this context that we show that the usurpation of the TIGIT − CD155 axis via engineered synNotch-mediated activation of induced pluripotent stem cell-derived natural killer (NK) cells promotes transcription factor-mediated activation of a downstream signaling cascade that results in the controlled, localized blockade of CD73 to disrupt purinergic activity otherwise resulting in the production and accumulation of immunosuppressive extracellular adenosine. Such "decoy" receptor engages CD155 binding to TIGIT, but tilts inhibitory TIGIT/CD155 interactions toward activation via downstream synNotch signaling. Usurping activities of TIGIT and CD73 promotes the function of adoptively transferred NK cells into intracranial patient-derived models of glioblastoma and enhances their natural cytolytic functions against this tumor to result in complete tumor eradication. In addition, targeting both receptors, in turn, reprograms the glioblastoma microenvironment via the recruitment of T cells and the downregulation of M2 macrophages. This study demonstrates that TIGIT/CD155 and CD73 are targetable receptor partners in glioblastoma. Our data show that synNotch-engineered pluripotent stem cell-derived NK cells are not only effective mediators of anti-glioblastoma responses within the setting of CD73 and TIGIT/CD155 co-targeting, but represent a powerful allogeneic treatment option for this tumor. Given its immunosuppressive effect in glioblastoma (GBM), targeting the TIGIT-CD155 axis presents an attractive therapeutic strategy. Here, the authors develop an adoptive natural killer (iNK) cells therapy with anti-CD155 synNotch-inducible CD73 antibody production to reverse the effect of TIGIT-CD155 signaling for the treatment of GBM. [ABSTRACT FROM AUTHOR]

Few data are available on antibody response for some SARS-CoV-2 vaccines, and there is a lack of ability to compare vaccine responses in the same population. This cross-sectional study conducted in Nicaragua examines the SARS-CoV-2 antibody responses in individuals, previously exposed to high infection rates who have received various vaccines. The vaccines under comparison include well-known ones like Pfizer (BNT162b2) and AstraZeneca (ChAdOx1-S), alongside less-studied vaccines including Soberana (Soberana 02), Abdala (CIGB-66), and Sputnik V/Sputnik Light. Overall, 3195 individuals participated, with 2862 vaccinated and 333 unvaccinated. We found that 95% of the unvaccinated were seropositive, with much lower titers than the vaccinated. Among the vaccinated, we found that Soberana recipients mounted the highest anti-spike response (mean difference (MD) = 36,498.8 [20,312.2, 52,685.5]), followed by Abdala (MD = 25,889.9 [10,884.1, 40,895.7]), BNT162b2 (MD = 12,967.2 [7543.7, 18,390.8]) and Sputnik with AstraZeneca as the reference group, adjusting for age, sex, vaccine status, days after last dose, and self-reported COVID-19. In addition, we found that subjects with complete vaccination series had higher antibody magnitude than those with incomplete series. Overall, we found no evidence of waning in the antibody magnitude across vaccines. Our study supports the conclusion that populations with high infection rates still benefit substantially from vaccination. [ABSTRACT FROM AUTHOR]