Humoral Immune Responses Research Articles

The capicua-ataxin-1-like complex regulates Notch-driven marginal zone B cell development and sepsis progression

Follicular B (FOB) and marginal zone B (MZB) cells are pivotal in humoral immune responses against pathogenic infections. MZB cells can exacerbate endotoxic shock via interleukin-6 secretion. Here we show that the transcriptional repressor capicua (CIC) and its binding partner, ataxin-1-like (ATXN1L), play important roles in FOB and MZB cell development. CIC deficiency reduces the size of both FOB and MZB cell populations, whereas ATXN1L deficiency specifically affects MZB cells. B cell receptor signaling is impaired only in Cic-deficient FOB cells, whereas Notch signaling is disrupted in both Cic-deficient and Atxn1l-deficient MZB cells. Mechanistically, ETV4 de-repression leads to inhibition of Notch1 and Notch2 transcription, thereby inhibiting MZB cell development in B cell-specific Cic-deficient (Cicf/f;Cd19-Cre) and Atxn1l-deficient (Atxn1lf/f;Cd19-Cre) mice. In Cicf/f;Cd19-Cre and Atxn1lf/f; Cd19-Cre mice, humoral immune responses and lipopolysaccharide-induced sepsis progression are attenuated but are restored upon Etv4-deletion. These findings highlight the importance of the CIC-ATXN1L complex in MZB cell development and may provide proof of principle for therapeutic targeting in sepsis.

Immunogenicity and Safety According to Immunosuppressive Drugs and Different COVID-19 Vaccine Platforms in Immune-Mediated Disease: Data from SAFER Cohort

Background/Objectives: The effectiveness of COVID-19 vaccine in patients with immune-mediated inflammatory diseases (IMID) depends on the underlying disease, immunosuppression degree and the vaccine regimens. We evaluate the safety and immunogenicity of different COVID-19 vaccine schedules. Methods: The SAFER study: “Safety and effectiveness of the COVID-19 Vaccine in Rheumatic Disease”, is a Brazilian multicentric prospective observational phase IV study in the real-life. Data were analyzed after 2 or 3 doses of COVID-19 vaccines: adenoviral vectored vaccine (ChAdOx1 nCoV-19, Astrazeneca), mRNA vaccine (BNT162b2, Pfizer–BioNTech) or inactivated SARS-COV-2 vaccine (CoronaVac, Sinovac Biotech). IgG antibody against SARS-CoV-2 spike (IgG-S) receptor-binding domain level were quantified at baseline (T1) and 28 days after the first (T2), 2nd (T3) and 3rd (T4) doses by chemiluminescence (SARS-CoV-2-IgG-II Quant-assay, Abbott-Laboratories). Results: 721 patients with IMID were included in the analysis. The median titers of IgG-S (BAU/mL) increased progressively over the times: at baseline was 6.26 (5.41–7.24), T2: 73.01 (61.53–86.62), T3: 200.0 (174.36–229.41) and T4: 904.92 (800.49–1022.97). The multivariate linear regression showed that greater IgG-S titers were associated with pre-exposure to COVID-19 (p < 0.001) and BNT162b2 booster vaccine (p < 0.001). Rituximab and immunosuppressant drugs were independent factors for low titers (p = 0.002, p < 0.001, respectively). No serious adverse event was reported. Conclusions: All platforms were safe and induced an increase in IgG-S antibodies. COVID-19 pre-exposure and BNT162b2 booster regimens were predictors of higher humoral immune responses, which is relevant in immunosuppressed populations. Immunosuppressants (mainly rituximab) predicted the lowest antibodies.

A Self‐Adjuvanting Large Pore 2D Covalent Organic Framework as a Vaccine Platform

Vaccines are one of the greatest human achievements in public health, as they help prevent the spread of diseases, reduce illness and death rates, saving thousands of lives with few side effects. Traditional vaccine development is centered around using live attenuated or inactivated pathogens, which is expensive and has resulted in vaccine‐associated illnesses. Advancements have led to the development of safer subunit vaccines, which contain recombinant proteins isolated from pathogens. Their short half‐life and small size make most subunit vaccines less immunogenic. Here, we introduce a large pore 2D covalent organic framework (COF), PyCOFamide, as a promising solution for an effective subunit platform. Our study demonstrates that simple adsorption of a model antigen, ovalbumin (OVA), onto PyCOFamide (OVA@COF) significantly enhances humoral and cell‐mediated immune response compared to free OVA. OVA@COF exhibited heightened immune cell activation and acts as an antigen reservoir, facilitating antigen‐presenting cell trafficking to the draining lymph nodes, amplifying the humoral immune response. Additionally, the breakdown of the COF releases monomers that adjuvant the activation of immune cells vital to creating strong immunity. This platform offers a potential avenue for safer, more effective subunit vaccines.

Spatial transcriptomics and in situ immune cell profiling of the host ectocervical landscape of HIV infected Kenyan sex working women

IntroductionChronic immune activation is a hallmark of human immunodeficiency virus (HIV) infection that significantly impacts disease pathogenesis. However, in-depth studies characterizing the immunological landscape of the ectocervix during chronic HIV infection remain scarce despite the importance of this tissue site for HIV transmission.MethodsEctocervical tissue samples were obtained from antiretroviral-naïve HIV-seropositive and -seronegative Kenyan female sex workers. These samples were assessed by spatial transcriptomics and Gene Set Enrichment Analysis. We further performed multi-epitope ligand cartography (MELC) using an in situ staining panel that included 17 markers of primarily T cell–mediated immune responses.ResultsSpatial transcriptomics revealed tissue-wide immune activation encompassing immune responses associated with chronic HIV infection. First, both the epithelial and submucosal compartments showed diverse but significant upregulation of humoral immune responses, as indicated by the expression of several antibody-related genes. Second, an antiviral state–associated cellular immunity was also observed in the HIV-seropositive group, characterized by upregulation of genes involved in interferon signaling across the mucosal tissue and a more spatially restricted mucosal expression of genes related to T cell activity and effector functions relative to the HIV-seronegative group. Additionally, HIV associated structural alterations were evident within both compartments. Downregulated genes across the epithelium were mainly linked to epithelial integrity, with the outer layer involved in terminal differentiation and the inner layer associated with epithelial structure. MELC analysis further revealed a significantly increased ectocervical leukocyte population in HIV-seropositive participants, primarily driven by an increase in CD8+ T cells while the CD4+ T cell population remained stable. Consistent with our spatial transcriptomics data, T cells from HIV-seropositive participants showed an increased effector phenotype, defined by elevated expression of various granzymes.ConclusionBy combining spatial transcriptomics and MELC, we identified significant HIV-associated cervical immune activity driven by induction of both T and B cell activity, together with a general antiviral state characterized by sustained interferon induction. These findings underscore that chronic HIV infection is associated with an altered ectocervical mucosal immune landscape years after primary infection. This sheds light on HIV pathogenesis at distant local sites and complements current knowledge on HIV-associated systemic immune activation.

Personalized Membrane Protein Vaccine Based on a Lipid Nanoparticle Delivery System Prevents Postoperative Recurrence in Colorectal Cancer Models

While accessing tumor neoantigens and developing effective delivery systems have posed significant challenges in therapeutic oncology vaccines, this study introduces a cost- and time-efficient personalized tumor vaccine demonstrating potent anti-tumor effects in a mouse xenograft model. This vaccine utilizes a lipid nanoparticle (C5 LNP) system loaded with membrane protein antigens (mAg) derived from surgically excised tumor tissue. Its safety and efficacy were validated in a B16-OVA murine model. C5/OVA exhibited significant uptake by dendritic cells (DCs), leading to cross-presentation, maturation, and subsequent initiation of robust cell-mediated and humoral immune responses. This resulted in clear tumor growth suppression and extended survival in the B16-OVA model. Furthermore, the personalized C5/mAg vaccine effectively inhibited tumor growth in a colorectal carcinoma model. When combined with anti-PD-1 therapy, it notably increased complete remission (CR) rates in the CT26 xenograft model. Vaccinated mice demonstrated 100% resistance to tumor rechallenge, underscoring the vaccine's ability to induce long-term immune memory. This study presents a promising personalized cancer vaccine delivery system with potential for both treatment and prevention of carcinoma in clinical applications. Statement of significanceIn this paper, we present a scheme for manufacturing personalized tumor vaccines. The vaccine component consists of lipid nanoparticles (LNPs) designated C5 and membrane protein antigens (mAg) derived from autologous tumor tissue surgically resected from the patient. Our study demonstrates that the personalized mAg-C5 vaccine for colorectal carcinoma significantly inhibits tumor growth. Furthermore, conjugation with anti-PD-1 therapy demonstrably increased the complete remission (CR) rate in the murine CT26 xenograft tumor model. Additionally, mice treated with the C5/mAg vaccine exhibited 100% resistance to tumor growth in a colon carcinoma rechallenge model, indicating the induction of immune memory by the vaccine.Our research results suggest that the mAg-LNP vaccine is a simple, cost-effective treatment that clinicians can easily and quickly integrate into routine practice.

Safety and immunogenicity of the yellow fever vaccine for patients with end-stage renal disease.

In December 2016, an outbreak of sylvatic yellow fever (YF) occurred in the non-endemic areas of the south-eastern region of Brazil. The immune response to the yellow fever vaccine and its safety in individuals with chronic kidney disease (CKD) living in YF-endemic regions are not thoroughly understood. The objective of this study is to assess the incidence of adverse events and the serological response after primary vaccination with the 17DD-YF vaccine in CKD patients undergoing dialysis. This was a multicenter, retrospective cohort study involving 223 individuals with CKD who were on dialysis after primary vaccination against YF. Clinical and epidemiologic characteristics were collected and the vaccine adverse event (VAE) were assessed. Around 35 months after vaccination, the serological response was evaluated in 71 (32%) patients using neutralization tests. No serious VAE occurred in any patient. Local reactions were reported in 13 individuals (5.8%), while 6 (2.7%) reported generalized systemic reactions and 205 (91.9%) did not display any VAE. No clinical or epidemiologic characteristic predicted the occurrence of VAE. Adequate serological response was found in 38% of participants and none of the clinical or epidemiological characteristics were associated with immunogenicity. The outcomes of our study suggest that the yellow YF vaccine is well-tolerated in CKD patients undergoing dialysis, but it does not induce adequate immune response. Future research should focus on evaluating both cellular and humoral immune responses following administration of various doses of the YF vaccine.

Elevated Soluble ACE2 Activity in Children and Adults After SARS-CoV-2 Exposure Irrespective of Laboratory-Confirmed Infection.

The pivotal role of the cell entry receptor ACE2 for SARS-CoV-2 infection is well-established. When ACE2 is shed from cell surface into plasma as soluble ACE2 (sACE2), it can effectively neutralize SARS-CoV-2. This longitudinal prospective cohort study analyzed sACE2 activity in 1192 participants, aged 4 months to 81 years, 3 and 12 months after SARS-CoV-2 household exposure. Following SARS-CoV-2 exposure, participants exhibited significantly elevated sACE2 activity, irrespective of confirmed infection, with the highest levels observed in exposed children. Longitudinal analysis revealed a decline in sACE2 levels over time, reaching levels comparable to age- and sex-matched pre-pandemic controls. An increase in sACE2 activity was also confirmed in vitro in Calu-3 (human lung) cells within hours of SARS-CoV-2 exposure, providing a direct link between SARS-CoV-2 exposure and elevated sACE2. This study, therefore, challenges the dichotomy of categorizing SARS-CoV-2 exposed participants as infected or not infected solely on currently established diagnostic assays. It demonstrates lasting host responses independent of B- and T-cell memory and may help to keep SARS-CoV-2 infections in balance and contribute to successful virus clearance in children and adults lacking humoral and cellular immune responses following SARS-CoV-2 exposure. Trial Registration: German Registry for Clinical Studies; Identifier: D 00021521.

Papillomaviruses and Papillomaviral Disease in Dogs and Cats: A Comprehensive Review

Papillomaviruses (PVs) frequently infect humans as well as non-human species. While most PV infections are asymptomatic, PVs can also cause hyperplastic papillomas (warts) as well as pre-neoplastic and neoplastic lesions. In this review, the life cycle of PVs is discussed, along with the mechanisms by which PVs cause hyperplastic and neoplastic diseases. The humoral and cell-mediated immune responses to PVs are reviewed, giving context to the later discussion on the use of vaccines to reduce canine and feline PV-associated disease. Both dogs and cats are infected by numerous different PV types classified into multiple different PV genera. The taxonomic classification of PVs is reviewed, along with the significance of this classification. The PV-associated diseases of dogs and cats are then described. These descriptions include the clinical presentation of the disease, the causative PV types, the histological features that allow diagnosis, and, where appropriate, possible treatment options. The review is comprehensive and contains the latest information about PVs and the diseases they cause in dogs and cats.

Using New Technologies to Analyze Gut Microbiota and Predict Cancer Risk

The gut microbiota significantly impacts human health, influencing metabolism, immunological responses, and disease prevention. Dysbiosis, or microbial imbalance, is linked to various diseases, including cancer. It is crucial to preserve a healthy microbiome since pathogenic bacteria, such as Escherichia coli and Fusobacterium nucleatum, can cause inflammation and cancer. These pathways can lead to the formation of tumors. Recent advancements in high-throughput sequencing, metagenomics, and machine learning have revolutionized our understanding of the role of gut microbiota in cancer risk prediction. Early detection is made easier by machine learning algorithms that improve the categorization of cancer kinds based on microbiological data. Additionally, the investigation of the microbiome has been transformed by next-generation sequencing (NGS), which has made it possible to fully profile both cultivable and non-cultivable bacteria and to understand their roles in connection with cancer. Among the uses of NGS are the detection of microbial fingerprints connected to treatment results and the investigation of metabolic pathways implicated in the development of cancer. The combination of NGS with machine learning opens up new possibilities for creating customized medicine by enabling the development of diagnostic tools and treatments that are specific to each patient’s microbiome profile, even in the face of obstacles like data complexity. Multi-omics studies reveal microbial interactions, biomarkers for cancer detection, and gut microbiota’s impact on cancer progression, underscoring the need for further research on microbiome-based cancer prevention and therapy.

Effects of immune response on different nano-adjuvants combined with H11 antigen of Haemonchus contortus

The intestinal native protein H11 is one of the most immunodominant antigen of Haemonchus contortus. However, H11 combined with QuilA adjuvant shows only short-lived protection for animals. Nano-adjuvants have the huge potential to extend the immune response in human and animals. Here, we compared the immune responses induced by H11 combined with three nano-adjuvants including lipid nanoparticles (LNP), immunostimulating complex matrix (IMX) and AddaS03™. We measured the cytokine levels of goat PBMCs stimulated by H11 mixed with three nano-adjuvants and QuilA. Results showed that the transcriptions of IL-6, IL-8 and IFN-γ genes were significantly inhibited in all adjuvant group compared with PBS control. H11-IMX combination significantly up-regulated IL-2, IL-17 and TNF-α gene transcriptions. The H11-LNP group showed a significant increase in IL-17 and TNF-α transcriptions, while the H11-AddaS03™ group significantly increased IL-2 transcription. Additionally, mice immunized with these formulations were assessed for splenic lymphocyte proliferation. All H11-adjuvant combinations, except H11-LNP, significantly stimulated lymphocyte proliferation compared to the H11 alone and PBS control groups, with the H11-AddaS03™ combination showing the strongest effect. Analysis of serum antibody levels revealed that all immune groups induced high IgM levels, with H11-IMX inducing the highest IgG levels. Our results demonstrated that IMX or LNP combined with H11 mainly induced a Th17 cell immune response in goat PBMCs, while IMX or AddaS03™ combined with H11 could induce a strong humoral immune response in mice. This study elucidates the immune response profiles of different nano-adjuvant combined with H11 antigen, providing important insights for vaccine development against parasitic nematodes.

Effects of Baseline Blood Zinc Levels on the Humoral Immune Response After COVID-19 mRNA Vaccination: A Prospective Study in a Japanese Population

Background/Objectives: Although the protective effects of zinc against COVID-19 are documented, its impact on COVID-19 vaccine immunogenicity remains unknown. Methods: We conducted a prospective study involving a cohort of 79 Japanese individuals (aged 21–56 years; comprising three subcohorts) and measured their serum zinc levels pre-vaccination and anti-SARS-CoV-2 IgM/IgG levels pre- and post-vaccination over 4 months. Results: Serum zinc concentrations ranged between 74–140 and 64–113 μg/dL in male and female individuals, respectively, with one male and 11 female participants exhibiting subclinical zinc deficiency (60–80 μg/dL). Mixed models for antibody titers, accounting for the subcohorts, repeat measurements, and covariates (e.g., vaccine type, sex, age, height, steroid use, medical history, smoking and drinking habits, perceived stress, and sleep disturbances) showed positive effects of zinc on IgM (p = 0.012) and IgG (p = 0.013) in 45 female individuals with 255 observations. However, a similar association was not found in the 34 male participants with 162 observations. This discrepancy may be attributed to one participant being included in the subcohort with frequent repeat measurements (10 repeats in 4 months). COVID-19 mRNA vaccine immunogenicity was enhanced in the participants with high baseline blood zinc levels within the reference range. Conclusions: Our findings underscore the relevance of maintaining adequate zinc levels before vaccination, which can be achieved through a balanced diet and healthy lifestyle choices.

Human Toll-like receptor activation by pathogenic Arcobacter species

The increase of Arcobacter spp. infection cases in humans, coupled with varying symptomatology, highlights the need to study the virulence mechanisms of these bacteria. Arcobacter butzleri can induce the release of several proinflammatory cytokines in human monocytic-derived macrophages, but the mechanism used to achieve this is still unclear. Therefore, we aimed to investigate the human innate immune response triggered by pathogenic Arcobacter spp., by studying the activation of the human Toll-like receptors (TLRs). Arcobacter skirrowii was the only species that showed the ability to activate all tested TLRs. Arcobacter cryaerophilus demonstrated to be able to activate TLR1/2, TLR4, and TLR2/6. A. butzleri hardly activated the TLRs, only TLR2/6 and TLR1/2 to a small extent. While all the Arcobacter species tested possess flagellum, as shown by motility assays and electron microscopy, only the flagellum of A. skirrowii was able to activate TLR5. The alignment of the flagellin amino acid data revealed that A. skirrowii shares a greater number of crucial amino acids for TLR5 recognition with the FliC of Salmonella than the other Arcobacter species, which might explain why A. skirrowii activates TLR5. Our results demonstrated that the activation of the different TLRs is Arcobacter species dependent, and there might be a correlation between the activation of the TLRs and the pathogenicity of the Arcobacter species.

Longitudinal Immunological Analysis of Portuguese Healthcare Workers Across the COVID-19 Pandemic Reveals Differences in the Humoral Immune Response to Vaccines

Background: A vaccination programme against severe acute respiratory syndrome coronavirus 2 was initiated in Portugal in December 2020. In this study, we report the findings of a prospective cohort study implemented with the objective of monitoring antibody production in response to COVID-19 vaccination. Methods: The humoral immune response to vaccination was followed up using blood samples collected from 191 healthcare workers. Participants were split into three groups: the Oxford-AstraZeneca (Vaxzevria) vaccine group (n = 68), the Pfizer-BioNTech COVID-19 (Comirnaty) vaccine group (n = 51), and the Post-COVID group (n = 72). The kinetics of anti-spike antibody production were evaluated until 56 days on average after the third dose (booster). Results: We observed that antibody titres peaked approximately one month after full vaccination and declined steadily thereafter. We also found that mRNA vaccination induces higher titres of antibodies than viral vector vaccination, and both generate greater antibody responses than mild or moderate COVID-19. Additionally, whilst the booster for the Oxford-AstraZeneca and Pfizer-BioNTech groups led to antibody levels higher than those at any previous sample collection point, the booster for the Post-COVID group (persons with a history of COVID-19 prior to vaccination) led to antibody levels lower than those attained one month after the second dose. Interpretation: Our results indicate that there are different kinetics of antibody production between individuals who received the Pfizer-BioNtech mRNA vaccine and those who received the Oxford-AstraZeneca vector vaccine, or individuals who had COVID-19 before being vaccinated. Additionally, we observed that exposure to either natural infection or vaccination modulates the response to subsequent vaccination. This is particularly evident after administration of the third dose to the Post-COVID group, where our findings point to a hindrance in vaccine boosting, probably due to unwanted feedback by high titres of pre-existing antibodies.

Construction and evaluation of glycoprotein-based nucleic acid vaccines for Marburg virus.

Marburg virus (MARV) is a zoonotic virus that can infect humans and non-human primates (NHPs) and lead to a fatal Marburg hemorrhagic fever (MHF), while there is no approved vaccine or antiviral treatment for MHF. The nucleic acid vaccine has unique advantages, including fast and simple preparation, easy to follow the virus mutation situation, and less adverse reactions. Therefore, we constructed the DNA and mRNA candidate vaccines based on codon-optimized MARV glycoprotein sequence, and evaluated the immune effect in mice through ELISA, ELISpot, and Flow cytometry. After the second booster immunization, both of the candidate vaccines induced strong humoral immune response, enhanced T cell response, and elicited neutralizing antibodies. Notably, DNA candidate vaccine induced stronger humoral immune response, while mRNA candidate vaccine elicited higher levels of IFN-γ and IL-4. In addition, transcriptome analysis revealed that the candidate vaccines activated immune response related pathways. Our study shed new light on the nucleic acid vaccines for MARV and further confirmed the potential of nucleic acid vaccine for future MHF prevention and control.

A Possible Protective Effect of IgA Against Severe Acute Respiratory Syndrome Corona Virus 2 (SARS-CoV-2) in Bronchoalveolar Lavage in COVID-19 Patients Admitted to Intensive Care Unit

SARS-CoV-2 infection induces a humoral immune response, producing virus-specific antibodies such as IgM, IgG, and IgA. IgA antibodies are present at mucosal sites, protecting against respiratory and other mucosal infections, including SARS-CoV-2, by neutralizing viruses or impeding attachment to epithelial cells. Since SARS-CoV-2 spreads through the nasopharynx, the specific IgAs of SARS-CoV-2 are produced quickly after infection, effectively contributing to virus neutralization. Dimeric IgA has been reported to be 10 to 15 times more potent than its equivalent IgG, suggesting that this isotype may be particularly interesting in developing new monoclonal antibodies and/or new vaccines efficiently neutralizing the virus at the mucosal sites. It is still unclear whether IgA antibodies in BAL might play a role in the disease course and if their presence may have a prognostic significance. However, a harmful effect on diseases with high IgA titers has been reported. This study evaluated mucosal-specific IgA and IgG profiles in BAL of patients with COVID-19 acute respiratory failure admitted to the ICU. We included 57 patients (41 males and 16 females), admitted to the ICU of the University of Foggia. We used a commercially available ELISA assay to evaluate the presence of SARS-CoV-2 IgG and IgA antibodies in plasma and BAL of the 57 hospitalized patients with severe COVID-19 respiratory failure. However, 40/57 BAL and plasma from infected patients were available for the ELISA test; the remaining specimens were unsuitable. IgG and IgA antibodies against SARS-CoV-2 were detectable in 37 (92.5%) and 40 (100%) plasma specimens, respectively. IgG antibodies were found in a single sample, while IgAs were detected in 19 of 40 BAL samples analyzed. Correlations between these parameters and patient outcomes reveal a signature associated with survival. Interestingly, a statistically significant inverse correlation was found between the mortality rate and the presence of IgA to SARS-CoV-2 in BAL specimens. None of the 19 patients with a positive IgA died, compared to 7 out of 12 patients with a negative IgA-BAL (p: <0.0004). Despite being limited in size, this study suggests a significant protective effect of mucosal immunity in COVID-19 patients, even in advanced disease stages, and a role of IgA in the defense against the virus, as well as the possible use of effective vaccines and therapeutic strategies based on IgA antibodies.

TbpB-based oral mucosal vaccine provides heterologous protection against Glässer’s disease caused by different serovars of Spanish field isolates of Glaesserella parasuis

BackgroundGlaesserella parasuis (G. parasuis) is the primary agent of Glässer’s disease, significantly affecting nursery and early fattening piglets. Current prophylactic measures, mainly serovar-specific bacterins administered to sows, are limited by maternal immunity, which can interfere with active immunization in piglets. Subunit vaccines containing G. parasuis-specific antigenic molecules show promise but are not yet commercially available. Transferrin-binding proteins (Tbp), which enable G. parasuis to acquire iron in low-iron environments like mucosal surfaces, have been proposed as potential vaccine antigens. The mucosal administration of a TbpB-based subunit vaccine could provide a promising solution to overcome the limitations posed by maternal immunity, offering an effective approach to control the disease in weaning piglets. This study, conducted in two phases, primarily evaluates (days 0–45) the immunogenicity of a two-dose oral mucosal TbpB-based subunit vaccine (TbpBY167A) administered to colostrum-deprived piglets, and subsequently (days 45–52), its heterologous protection by challenging these piglets with four G. parasuis clinical isolates from different TbpB clusters (I, III) and serovars (SV1, SV4, SV5, SV7) recovered from Spanish pig farms.ResultsThe oral mucosal administration of the two-dose TbpB-based vaccine induced a robust humoral immune response in immunized colostrum-deprived piglets, significantly increasing IgA and IgM concentration 15 days after the second dose (p < 0.01). Upon challenge with four G. parasuis clinical isolates, the vaccine demonstrated heterologous protection, markedly improving survival rates (OR: 8.45; CI 95%: 4.97–14.36) and significantly reducing clinical signs and lesions, regardless of the TbpB cluster and serovar. The vaccine reduced G. parasuis colonization in the respiratory tract (p < 0.0001) and G. parasuis systemic target tissues, like tarsus and carpus joints, liver, and brain (p < 0.05). Immunohistochemical analysis showed a lower macrophage count in different lung locations of immunized piglets (p < 0.0001).ConclusionsThis study demonstrates that oral mucosal administration of the TbpBY167A subunit vaccine in piglets provides effective heterologous protection against diverse virulent European G. parasuis field isolates, significantly reducing bacterial colonization and dissemination. This vaccine offers a promising alternative to traditional bacterins, overcoming limitations due to maternal immunity, and represents a strong candidate for universal vaccination against Glässer’s disease.

The analysis of recombinant insulin production technologies

Aim. To analyze current trends in insulin production and expression systems used to obtain recombinant proteins. Materials and methods. The specified goal was achieved using the methods of content analysis, comparative, logical, analytical and generalization of information. The research materials were publications in scientific periodicals, official websites of manufacturing companies. Results. It has been found that the following expression systems are traditionally used for the commercial production of recombinant human insulins: bacterial – Escherichia coli, and yeast – Saccharomyces cerevisiae. But to meet the global need for insulin, it is necessary to develop new effective expression systems. Therefore, the use of other eukaryotic insulin expression systems that would be suitable for large-scale production is currently being studied. Among such systems, transgenic plants, which are characterized by the absence of potential human pathogens and the presence of post-translational protein modification mechanisms similar to human ones, are promising for introduction into insulin production. Currently, studies using Arabidopsis thaliana, tobacco, lettuce and strawberry plants are successful. Stem cells – embryonic, mesenchymal and induced pluripotent are also considered promising. The methods used in the large-scale production of recombinant insulin based on the synthesis of proinsulin and the two-chain method have been characterized. The technological flowcharts of both technologies have been drawn up, the advantages and disadvantages of each are given. Conclusions. The analysis of data from the scientific literature has shown that the main methods of producing insulin preparations since the 1980s remain pro-insulin and two-chain methods, which are based on recombinant DNA technologies. The pro-insulin method is considered more efficient and cost-effective compared to the two-chain method since this technology consists of working with a single recombinant strain. The choice between these two methods depends on the specific needs of the manufacturer: for some cases a fast method with proinsulin synthesis is rational, for others – a more accurate and controlled two-chain method. The bacteria – Escherichia coli and yeast – Saccharomyces cerevisiae remain the expression systems used in the existing large-scale production technologies. But using E. coli, insulin precursors are produced in inclusion bodies, and fully functional polypeptides are obtained by solubilization and refolding steps. The yeast-based system yields a soluble insulin precursor that is secreted into the culture fluid, but requires prior humanization due to the risks of an immune response in humans. Thus, current production technologies cannot meet the growing demand for available insulin due to limited production capacity and high production costs, so research is being conducted to find new efficient expression systems, such as plant and mammalian cells, including Arabidopsis thaliana, tobacco, lettuce, strawberry, stem cells.

Expression and delivery of HA1-M2e antigen using an innovative attenuated Salmonella–mediated delivery system confers promising protection against H9N2 avian influenza challenge

This study explores a dual expression vector system for delivering prokaryotic and eukaryotic antigens to improve conventional vaccination strategies. To enhance immune protection against H9N2 avian influenza virus (AIV), which threatens poultry and humans, we used the previously constructed pJHL270 and pJHL305 plasmids with the Ptrc and CMV promoters to stimulate MHC class II and I responses through exogenous and endogenous antigenic presentation. Salmonella Gallinarum (SG), a delivery vector, was engineered to have defective lipopolysaccharide structures through lon, pagL, and rfaL deletion. It demonstrated a safety profile with lower induction of inflammatory cytokines than the wild-type strain. Bioinformatics tools predicted that the HA1 and M2e sequences, which were designed as consensus sequences of South Korean strains (2000–2021), would have high antigenicity and favorable structures. In vitro expression of the vaccine constructs was validated by western blotting. Birds immunized with attenuated SG harboring pJHL270 (JOL3025) or pJHL305 (JOL3027) containing HA-M2e showed significant increases in serum IgY and mucosal IgA antibodies, indicating strong humoral and mucosal immune responses, comparable with inactivated commercial vaccine. Post-immunization, we found a substantial rise in the hemagglutination inhibition titer, suggesting effective prevention of viral attachment and robust cell-mediated immunity, with a 1.96-fold and 2.80-fold increase in CD4+ and CD8+ T cells, respectively, for JOL3025 and a 1.75-fold and 2.49-fold increase for JOL3027. Furthermore, MHC class I and II expression increased 1.35-fold and 1.63-fold, for JOL3025, and 1.61-fold and 1.68-fold, respectively, for JOL3027. The IL-4 and IFN-γ levels were elevated, indicating a balanced Th-1 and Th-2 response. Post-challenge, birds immunized with vaccine candidates or the commercial vaccine exhibited minimal to no clinical signs, reduced lesions, lower lung viral titers, and negligible impacts on egg production compared to controls. In conclusion, both plasmids successfully delivered HA1-M2e immunogens through the engineered SG strains, eliciting strong humoral, mucosal, and cell-mediated immune responses and co-stimulating MHC class I and II antigen presentation pathways to provide effective protection against H9N2 AIV with minimal adverse effects.

Distinct Requirements for CD4+ T Cell Help for Immune Responses Induced by mRNA and Adenovirus-Vector SARS-CoV-2 Vaccines.

CD4+ T cells have been established as central orchestrators of cellular and humoral immune responses to infection or vaccination. However, the need for CD4+ T cell help to generate primary CD8+ T cell responses is variable depending on the infectious agent or vaccine and yet consistently required for the recall of CD8+ T cell memory responses or antibody responses. Given the deployment of new vaccine platforms such as nucleoside-modified mRNA vaccines, we sought to elucidate the requirement for CD4+ T cell help in the induction of cellular and antibody responses to mRNA and adenovirus (Ad)-vectored vaccines against SARS-CoV-2. Using antibody-mediated depletion of CD4+ T cells in a mouse immunization model, we observed that CD4+ T cell help was dispensable for both primary and secondary CD8+ T cell responses to the BNT162b2 and mRNA-1273 mRNA vaccines but required for the AZD1222 Ad-vectored vaccine. Nonetheless, CD4+ T cell help was needed by both mRNA and Ad-vectored vaccine platforms for the generation of antibodies, demonstrating the centrality of CD4+ T cells in vaccine-induced protective immunity against SARS-CoV-2. Ultimately, this highlights the shared and distinct regulation of humoral and cellular responses induced by these vaccine platforms.

Evaluation of alternative vaccination routes against paratuberculosis in goats

Paratuberculosis is a chronic granulomatous enteritis, caused by Mycobacterium avium subspecies paratuberculosis (Map), that affects ruminants worldwide. Vaccination has been considered the most cost-effective method for the control of this disease in infected dairy herds. However, currently available vaccines do not provide complete protection and interfere with the diagnosis of both paratuberculosis and bovine tuberculosis, limiting its use. Because of that, efforts are being made for the development of new vaccines. The primary objective of this study was to evaluate the efficacy of two whole-cell inactivated experimental vaccines against paratuberculosis in goats, administered through the oral (OV) and intradermal (IDV) routes, and compare them with that of the commercial subcutaneous vaccine Gudair® (SCV). Over an 11-month period, the effect of vaccination and a subsequent Map challenge on the specific peripheral immune responses and Map-DNA fecal shedding were recorded. At the end of the experiment, tissue bacterial load and lesion severity were assessed. The experimental vaccines did not induce specific humoral immune responses and only elicited mild and delayed cellular immune responses. Although the OV reduced lesion severity, neither this vaccine nor the IDV prototype was able to reduce fecal shedding or tissue bacterial load. Moreover, although the SCV did not confer sterile immunity, it outperformed both experimental vaccines in all these parameters.