Development Of Mucosal Vaccines Research Articles

Harnessing Bacillus subtilis Spore Surface Display (BSSD) Technology for Mucosal Vaccines and Drug Delivery: Innovations in Respiratory Virus Immunization

Respiratory viruses present significant global health challenges due to their rapid evolution, efficient transmission, and zoonotic potential. These viruses primarily spread through aerosols and droplets, infecting respiratory epithelial cells and causing diseases of varying severity. While traditional intramuscular vaccines are effective in reducing severe illness and mortality, they often fail to induce sufficient mucosal immunity, thereby limiting their capacity to prevent viral transmission. Mucosal vaccines, which specifically target the respiratory tract’s mucosal surfaces, enhance the production of secretory IgA (sIgA) antibodies, neutralize pathogens, and promote the activation of tissue-resident memory B cells (BrMs) and local T cell responses, leading to more effective pathogen clearance and reduced disease severity. Bacillus subtilis spore surface display (BSSD) technology is emerging as a promising platform for the development of mucosal vaccines. By harnessing the stability and robustness of Bacillus subtilis spores to present antigens on their surface, BSSD technology offers several advantages, including enhanced stability, cost-effectiveness, and the ability to induce strong local immune responses. Furthermore, the application of BSSD technology in drug delivery systems opens new avenues for improving patient compliance and therapeutic efficacy in treating respiratory infections by directly targeting mucosal sites. This review examines the potential of BSSD technology in advancing mucosal vaccine development and explores its applications as a versatile drug delivery platform for combating respiratory viral infections.

Single cell transcriptional analysis of human adenoids identifies molecular features of airway microfold cells.

The nasal, oropharyngeal, and bronchial mucosa are primary contact points for airborne pathogens like Mycobacterium tuberculosis (Mtb), SARS-CoV-2, and influenza virus. While mucosal surfaces can function as both entry points and barriers to infection, mucosa-associated lymphoid tissues (MALT) facilitate early immune responses to mucosal antigens. MALT contains a variety of specialized epithelial cells, including a rare cell type called a microfold cell (M cell) that functions to transport apical antigens to basolateral antigen-presenting cells, a crucial step in the initiation of mucosal immunity. M cells have been extensively characterized in the gastrointestinal (GI) tract in murine and human models. However, the precise development and functions of human airway M cells is unknown. Here, using single-nucleus RNA sequencing (snRNA-seq), we generated an atlas of cells from the human adenoid and identified 16 unique cell types representing basal, club, hillock, and hematopoietic lineages, defined their developmental trajectories, and determined cell-cell relationships. Using trajectory analysis, we found that human airway M cells develop from progenitor club cells and express a gene signature distinct from intestinal M cells. Surprisingly, we also identified a heretofore unknown epithelial cell type demonstrating a robust interferon-stimulated gene signature. Our analysis of human adenoid cells enhances our understanding of mucosal immune responses and the role of M cells in airway immunity. This work also provides a resource for understanding early interactions of pathogens with airway mucosa and a platform for development of mucosal vaccines.

Early, robust mucosal secretory IgA but not IgG response to SARS-CoV-2 spike in oral fluid is associated with faster viral clearance and COVID-19 symptom resolution.

High priority efforts are underway to support the development of novel mucosal COVID-19 vaccines, such as the US Government's Project NextGen and the Center for Epidemic Preparedness Innovations' goal to respond to the next pandemic with a new vaccine in 100 days. However, there is limited consensus about the complementary role of mucosal immunity in disease progression and how to evaluate immunogenicity of mucosal vaccines. This study investigated the role of oral mucosal antibody responses in viral clearance and COVID-19 symptom duration. Participants with PCR-confirmed SARS-CoV-2 infection provided oral fluid for testing with SARS-CoV-2 antibody multiplex assays, nasal swabs for RT-PCR and symptom information at up to eight follow-ups from April 2020 to February 2022. High and moderate oral fluid anti-spike (S) secretory IgA (SIgA) post infection was associated with significantly faster viral clearance and symptom resolution across age groups with effect sizes equivalent to having COVID-19 vaccine immunity at the time of infection. Those with high and moderate anti-S SIgA cleared the virus 14 days (95% CI: 10-18) and recovered 9-10 days (95% CI: 6-14) earlier. Delayed and higher anti-S IgG was associated with significantly longer time to clearance and recovery. Experiencing symptoms longer than four weeks was associated with lower anti-RBD SIgA 15-30 days after infection onset (p<0.001). Robust mucosal SIgA early post infection appears to support faster clearance of SARS-CoV-2 and recovery from COVID-19 symptoms. This research underscores the importance of harmonizing mucosal immune response assays to evaluate new mucosal vaccines.

Intranasal Immunization with a Recombinant Adenovirus Encoding Multi-Stage Antigens of Mycobacterium tuberculosis Preferentially Elicited CD8+ T Cell Immunity and Conferred a Superior Protection in the Lungs of Mice than Bacillus Calmette-Guerin.

The development of a tuberculosis (TB) vaccine is imperative. Employing multi-stage Mycobacterium tuberculosis (Mtb) antigens as targeted antigens represents a critical strategy in establishing an effective novel TB vaccine. In this investigation, we evaluated the immunogenicity and protective efficacy of a recombinant adenovirus vaccine expressing two fusion proteins, Ag85B-ESAT6 (AE) and Rv2031c-Rv2626c (R2), derived from multi-stage antigens of Mtb via intranasal administration in mice. Intranasal delivery of Ad-AE-R2 induced both long-lasting mucosal and systemic immunities, with a preferential elicitation of CD8+ T cell immunity demonstrated by the accumulation and retention of CD8+ T cells in BALF, lung, and spleen, as well as the generation of CD8+ TRM cells in BALF and lung tissues. Compared to subcutaneous immunization with Bacillus Calmette-Guerin (BCG), Ad-AE-R2 provided superior protection against high-dose intratracheal BCG challenge, specifically within the lungs of mice. Our findings support the notion that empowering T cells within the respiratory mucosa is crucial for TB vaccine development while highlighting targeting CD8+ T cell immunity as an effective strategy for optimizing TB vaccines and emphasizing that eliciting systemic memory immunity is also vital for the successful development of a TB mucosal vaccine. Furthermore, our results demonstrate that the BCG challenge serves as a convenient and efficient method to evaluate candidate vaccine efficacy.

Harnessing the potential of the NALT and BALT as targets for immunomodulation using engineering strategies to enhance mucosal uptake.

Mucosal barrier tissues and their mucosal associated lymphoid tissues (MALT) are attractive targets for vaccines and immunotherapies due to their roles in both priming and regulating adaptive immune responses. The upper and lower respiratory mucosae, in particular, possess unique properties: a vast surface area responsible for frontline protection against inhaled pathogens but also simultaneous tight regulation of homeostasis against a continuous backdrop of non-pathogenic antigen exposure. Within the upper and lower respiratory tract, the nasal and bronchial associated lymphoid tissues (NALT and BALT, respectively) are key sites where antigen-specific immune responses are orchestrated against inhaled antigens, serving as critical training grounds for adaptive immunity. Many infectious diseases are transmitted via respiratory mucosal sites, highlighting the need for vaccines that can activate resident frontline immune protection in these tissues to block infection. While traditional parenteral vaccines that are injected tend to elicit weak immunity in mucosal tissues, mucosal vaccines (i.e., that are administered intranasally) are capable of eliciting both systemic and mucosal immunity in tandem by initiating immune responses in the MALT. In contrast, administering antigen to mucosal tissues in the absence of adjuvant or costimulatory signals can instead induce antigen-specific tolerance by exploiting regulatory mechanisms inherent to MALT, holding potential for mucosal immunotherapies to treat autoimmunity. Yet despite being well motivated by mucosal biology, development of both mucosal subunit vaccines and immunotherapies has historically been plagued by poor drug delivery across mucosal barriers, resulting in weak efficacy, short-lived responses, and to-date a lack of clinical translation. Development of engineering strategies that can overcome barriers to mucosal delivery are thus critical for translation of mucosal subunit vaccines and immunotherapies. This review covers engineering strategies to enhance mucosal uptake via active targeting and passive transport mechanisms, with a parallel focus on mechanisms of immune activation and regulation in the respiratory mucosa. By combining engineering strategies for enhanced mucosal delivery with a better understanding of immune mechanisms in the NALT and BALT, we hope to illustrate the potential of these mucosal sites as targets for immunomodulation.

Host Functional Response to a Prototypic Orally Delivered Self-Replicating Vaccine Platform.

The development of mucosal vaccines has been limited and could be aided by a systems vaccinology approach to identify platforms and adjuvant strategies that induce protective immune responses. The induction of local immune responses by mucosal-delivered vaccines has been difficult to evaluate from peripheral samples, as systemic responses often do not correlate with the mucosal response. Here, we utilized transcriptomics in combination with Gene Set Enrichment Analysis (GSEA) to assess innate immune activation by an oral probiotic Lactobacillus acidophilus-based vaccine platform in mice. The goal was to explore the earliest immune responses elicited after oral immunization at the Peyer's patch. Twenty-four hours after oral delivery of the L. acidophilus vaccine platform, we found an abundance of L. acidophilus at Peyer's patches and detected expression of the vaccine viral proteins and adjuvants, confirming in vivo vaccine delivery. Compared to mice orally dosed with buffer or wild-type L. acidophilus, we identified enhanced responses in immune pathways related to cytokine and gene signaling, T and B cell activation, phagocytosis, and humoral responses. While more work is needed to correlate these pathways with protection from infection and/or disease, they indicate this method's potential to evaluate and aid in the iterative development of next-generation mucosal vaccines.

Development of an anti-tauopathy mucosal vaccine specifically targeting pathologic conformers

Alzheimer’s disease (AD) and related tauopathies are associated with pathological tau protein aggregation, which plays an important role in neurofibrillary degeneration and dementia. Targeted immunotherapy to eliminate pathological tau aggregates is known to improve cognitive deficits in AD animal models. The tau repeat domain (TauRD) plays a pivotal role in tau-microtubule interactions and is critically involved in the aggregation of hyperphosphorylated tau proteins. Because TauRD forms the structural core of tau aggregates, the development of immunotherapies that selectively target TauRD-induced pathological aggregates holds great promise for the modulation of tauopathies. In this study, we generated recombinant TauRD polypeptide that form neurofibrillary tangle-like structures and evaluated TauRD-specific immune responses following intranasal immunization in combination with the mucosal adjuvant FlaB. In BALB/C mice, repeated immunizations at one-week intervals induced robust TauRD-specific antibody responses in a TLR5-dependent manner. Notably, the resulting antiserum recognized only the aggregated form of TauRD, while ignoring monomeric TauRD. The antiserum effectively inhibited TauRD filament formation and promoted the phagocytic degradation of TauRD aggregate fragments by microglia. The antiserum also specifically recognized pathological tau conformers in the human AD brain. Based on these results, we engineered a built-in flagellin-adjuvanted TauRD (FlaB-TauRD) vaccine and tested its efficacy in a P301S transgenic mouse model. Mucosal immunization with FlaB-TauRD improved quality of life, as indicated by the amelioration of memory deficits, and alleviated tauopathy progression. Notably, the survival of the vaccinated mice was dramatically extended. In conclusion, we developed a mucosal vaccine that exclusively targets pathological tau conformers and prevents disease progression.

Inflammatory Profiles Induced by Intranasal Immunization with Ricin Toxin-immune Complexes.

The underlying contribution of immune complexes in modulating adaptive immunity in mucosal tissues remains poorly understood. In this report, we examined, in mice, the proinflammatory response elicited by intranasal delivery of the biothreat agent ricin toxin (RT) in association with two toxin-neutralizing mAbs, SylH3 and PB10. We previously demonstrated that ricin-immune complexes (RICs) induce the rapid onset of high-titer toxin-neutralizing Abs that persist for months. We now demonstrate that such responses are dependent on CD4+ T cell help, because treatment of mice with an anti-CD4 mAb abrogated the onset of RT-specific Abs following intranasal RICs exposure. To define the inflammatory environment associated with RIC exposure, we collected bronchoalveolar lavage fluid (BALF) and sera from mice 6, 12, and 18 h after they had received RT or RICs by the intranasal route. A 32-plex cytometric bead array revealed an inflammatory profile elicited by RT that was dominated by IL-6 (>1500-fold increase in BALF) and secondarily by KC (CXCL1), G-CSF, GM-CSF, and MCP-1. RICs induced inflammatory profiles in both BALF and serum response that were similar to RT, albeit at markedly reduced levels. These results demonstrate that RICs retain the capacity to induce local and systemic inflammatory cytokines/chemokines that, in turn, may influence Ag sampling and presentation in the lung mucosa and draining lymph nodes. A better understanding of the fate of immune complexes following intranasal delivery has implications for the development of mucosal vaccines for biothreats and emerging infectious diseases.

Double synergic chitosan-coated poly (lactic-co-glycolic) acid nanospheres loaded with nucleic acids as an intranasally administered vaccine delivery system to control the infection of foot-and-mouth disease virus

The spread of foot-and-mouth disease virus (FMDV) through aerosol droplets among cloven-hoofed ungulates in close contact is a major obstacle for successful animal husbandry. Therefore, the development of suitable mucosal vaccines, especially nasal vaccines, to block the virus at the initial site of infection is crucial. Here, we constructed eukaryotic expression plasmids containing the T and B-cell epitopes (pTB) of FMDV in tandem with the molecular mucosal adjuvant Fms-like tyrosine kinase receptor 3 ligand (Flt3 ligand, FL) (pTB-FL). Then, the constructed plasmid was electrostatically attached to mannose-modified chitosan-coated poly(lactic-co-glycolic) acid (PLGA) nanospheres (MCS-PLGA-NPs) to obtain an active nasal vaccine targeting the mannose-receptor on the surface of antigen-presenting cells (APCs). The MCS-PLGA-NPs loaded with pTB-FL not only induced a local mucosal immune response, but also induced a systemic immune response in mice. More importantly, the nasal vaccine afforded an 80% protection rate against a highly virulent FMDV strain (AF72) when it was subcutaneously injected into the soles of the feet of guinea pigs. The nasal vaccine prepared in this study can effectively induce a cross-protective immune response against the challenge with FMDV of same serotype in animals and is promising as a potential FMDV vaccine.

Polymeric Caffeic Acid Acts as an Antigen Delivery Carrier for Mucosal Vaccine Formulation by Forming a Complex with an Antigenic Protein.

The development of mucosal vaccines, which can generate antigen-specific immune responses in both the systemic and mucosal compartments, has been recognized as an effective strategy for combating infectious diseases caused by pathogenic microbes. Our recent research has focused on creating a nasal vaccine system in mice using enzymatically polymerized caffeic acid (pCA). However, we do not yet understand the molecular mechanisms by which pCA stimulates antigen-specific mucosal immune responses. In this study, we hypothesized that pCA might activate mucosal immunity at the site of administration based on our previous findings that pCA possesses immune-activating properties. However, contrary to our initial hypothesis, the intranasal administration of pCA did not enhance the expression of various genes involved in mucosal immune responses, including the enhancement of IgA responses. Therefore, we investigated whether pCA forms a complex with antigenic proteins and enhances antigen delivery to mucosal dendritic cells located in the lamina propria beneath the mucosal epithelial layer. Data from gel filtration chromatography indicated that pCA forms a complex with the antigenic protein ovalbumin (OVA). Furthermore, we examined the promotion of OVA delivery to nasal mucosal dendritic cells (mDCs) after the intranasal administration of pCA in combination with OVA and found that OVA uptake by mDCs was increased. Therefore, the data from gel filtration chromatography and flow cytometry imply that pCA enhances antigen-specific antibody production in both mucosal and systemic compartments by serving as an antigen-delivery vehicle.

Vaccine Strategies to Elicit Mucosal Immunity.

Vaccines are essential tools to prevent infection and control transmission of infectious diseases that threaten public health. Most infectious agents enter their hosts across mucosal surfaces, which make up key first lines of host defense against pathogens. Mucosal immune responses play critical roles in host immune defense to provide durable and better recall responses. Substantial attention has been focused on developing effective mucosal vaccines to elicit robust localized and systemic immune responses by administration via mucosal routes. Mucosal vaccines that elicit effective immune responses yield protection superior to parenterally delivered vaccines. Beyond their valuable immunogenicity, mucosal vaccines can be less expensive and easier to administer without a need for injection materials and more highly trained personnel. However, developing effective mucosal vaccines faces many challenges, and much effort has been directed at their development. In this article, we review the history of mucosal vaccine development and present an overview of mucosal compartment biology and the roles that mucosal immunity plays in defending against infection, knowledge that has helped inform mucosal vaccine development. We explore new progress in mucosal vaccine design and optimization and novel approaches created to improve the efficacy and safety of mucosal vaccines.

Role of cellular effectors in the induction and maintenance of IgA responses leading to protective immunity against enteric bacterial pathogens.

The mucosal immune system is a critical first line of defense to infectious diseases, as many pathogens enter the body through mucosal surfaces, disrupting the balanced interactions between mucosal cells, secretory molecules, and microbiota in this challenging microenvironment. The mucosal immune system comprises of a complex and integrated network that includes the gut-associated lymphoid tissues (GALT). One of its primary responses to microbes is the secretion of IgA, whose role in the mucosa is vital for preventing pathogen colonization, invasion and spread. The mechanisms involved in these key responses include neutralization of pathogens, immune exclusion, immune modulation, and cross-protection. The generation and maintenance of high affinity IgA responses require a delicate balance of multiple components, including B and T cell interactions, innate cells, the cytokine milieu (e.g., IL-21, IL-10, TGF-β), and other factors essential for intestinal homeostasis, including the gut microbiota. In this review, we will discuss the main cellular components (e.g., T cells, innate lymphoid cells, dendritic cells) in the gut microenvironment as mediators of important effector responses and as critical players in supporting B cells in eliciting and maintaining IgA production, particularly in the context of enteric infections and vaccination in humans. Understanding the mechanisms of humoral and cellular components in protection could guide and accelerate the development of more effective mucosal vaccines and therapeutic interventions to efficiently combat mucosal infections.

Mucosa-Associated Lymphoid Tissue: Guardians of Immunity at Mucosal Frontiers

The mucosa-associated lymphoid tissue (MALT) stands as an intricate and versatile arm of the immune system, positioned strategically at the body's mucosal interfaces. This specialized lymphoid tissue serves as a sentinel against myriad pathogenic challenges while fostering immune tolerance to commensal microorganisms. This article explores the structural and functional attributes of MALT, delving into its critical role in immune surveillance, protection, and homeostasis within mucosal environments. We delve into the intricate interplay between MALT and its cellular constituents, focusing on the lymphocytes, follicular dendritic cells, and the diverse array of immunoglobulins. This comprehensive investigation illuminates the relevance of MALT in the context of infectious diseases, autoimmunity, and inflammation, emphasizing its potential as a therapeutic target. Additionally, we discuss the implications of MALT in the development of mucosal vaccines and the ongoing research avenues poised to unveil novel insights into this remarkable immunological asset.

Nasal Immunization Using Chitosan Nanoparticles with Glycoprotein B of Murine Cytomegalovirus.

The use of nanoparticles as a delivery system for a specific antigen could solve many limitations of mucosal vaccine applications, such as low immunogenicity, or antigen protection and stabilization. In this study, we tested the ability of nasally administered chitosan nanoparticles loaded with glycoprotein B of murine cytomegalovirus to induce an immune response in an animal model. The choice of chitosan nanoparticle type was made by in vitro evaluation of sorption efficiency and antigen release. Three types of chitosan nanoparticles were prepared: crosslinked with tripolyphosphate, coated with hyaluronic acid, and in complex with polycaprolactone. The hydrodynamic size of the nanoparticles by dynamic light scattering, zeta potential, Fourier transform infrared spectroscopy, scanning electron microscopy, stability, loading efficiency, and release kinetics with ovalbumin were evaluated. Balb/c mice were immunized intranasally using the three-dose protocol with nanoparticles, gB, and adjuvants Poly(I:C) and CpG ODN. Subsequently, the humoral and cell-mediated antigen-specific immune response was determined. On the basis of the properties of the tested nanoparticles, the cross-linked nanoparticles were considered optimal for further investigation. The results show that nanoparticles with Poly(I:C) and with gB alone raised IgG antibody levels above the negative control. In the case of mucosal IgA, only gB alone weakly induced the production of IgA antibodies compared to saline-immunized mice. The number of activated cells increased slightly in mice immunized with nanoparticles and gB compared to those immunized with gB alone or to negative control. The results demonstrated that chitosan nanoparticles could have potential in the development of mucosal vaccines.

Thiolated chitosan encapsulation constituted mucoadhesive nanovaccine confers broad protection against divergent influenza A viruses

Influenza A virus (IAV) poses a significant threat to human and animal health, necessitating the development of universal influenza vaccines that can effectively activate mucosal immunity. Intranasal immunization has attracted significant attention due to its capacity to induce triple immune responses, including mucosal secretory IgA. However, inducing mucosal immunity through vaccination is challenging due to the self-cleansing nature of the mucosal surface. Thiolated chitosan (TCS) were explored for mucosal vaccine delivery, capitalizing on biocompatibility and bioadhesive properties of chitosan, with thiol modification enhancing mucoadhesive capability. The focus was on developing a universal nanovaccine by utilizing TCS-encapsulated virus-like particles displaying conserved B-cell and T-cell epitopes from M2e and NP proteins of IAV. The optimal conditions for nanoparticle formation were investigated by adjusting the thiol groups content of TCS and the amount of sodium tripolyphosphate. The nanovaccine induced robust immune responses and provided complete protection against IAVs from different species following intranasal immunization. The broad protective effect of nanovaccines can be attributed to the synergistic effect of antibodies and T cells. This study developed a universal intranasal nanovaccine and demonstrated the potential of TCS in the development of mucosal vaccines for respiratory infectious diseases.

The 2022 Vaccines Against Shigella and Enterotoxigenic Escherichia coli (VASE) Conference: Summary of breakout workshops

The global public health nonprofit organization PATH hosted the third Vaccines Against Shigella and Enterotoxigenic Escherichia coli (VASE) Conference in Washington, DC, from November 29 to December 1, 2022. This international gathering focused on cutting-edge research related to the development of vaccines against neglected diarrheal pathogens including Shigella, enterotoxigenic Escherichia coli (ETEC), Campylobacter, and non-typhoidal Salmonella. In addition to the conference’s plenary content, the agenda featured ten breakout workshops on topics of importance to the enteric vaccine field. This unique aspect of VASE Conferences allows focused groups of attendees to engage in in-depth discussions on subjects of interest to the enteric vaccine development community. In 2022, the workshops covered a range of topics. Two focused on the public health value of enteric vaccines, with one examining how to translate evidence into policy and the other on the value proposition of potential combination vaccines against bacterial enteric pathogens. Two more workshops explored new tools for the development and evaluation of vaccines, with the first on integrating antigen/antibody technologies for mucosal vaccine and immunoprophylactic development, and the second on adjuvants specifically for Shigella vaccines for children in low- and middle-income countries. Another pair of workshops covered the status of vaccines against two emerging enteric pathogens, Campylobacter and invasive non-typhoidal Salmonella. The remaining four workshops examined the assessment of vaccine impact on acute and long-term morbidity. These included discussions on the nature and severity of intestinal inflammation; cellular immunity and immunological memory in ETEC and Shigella infections; clinical and microbiologic endpoints for Shigella vaccine efficacy studies in children; and intricacies of protective immunity to enteric pathogens. This article provides a brief summary of the presentations and discussions at each workshop in order to share these sessions with the broader enteric vaccine field.

Spatiotemporal observations of host-pathogen interactions in mucosa during SARS-CoV-2 infection indicate a protective role of ILC2s

ABSTRACT Innate lymphoid cells (ILCs) play a crucial role in mucosal protection and tissue homeostasis. However, the early mucosal defense mechanisms against the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus have not been fully understood to date. This knowledge gap has motivated us to develop a coronavirus disease 2019-like mouse model to investigate the interactions between the virus, its receptor, and ILCs. In our study, we conducted intranasal challenges using pseudovirus expressing Spike (PSV-S) to examine the expression patterns of humanized ACE2 (chiACE2) and the transduction of PSV-S in lung and ileum tissues over a specified time period. Within a span of 72 hours, we observed an intense viral transduction induced by the spike protein, which was detected from the central bronchus to the bronchiole and alveolus in the lung (levels 3 to 8). Similarly, we observed the transduction of PSV-S in the villi and crypts of the ileum. Interestingly, our precise three-dimensional colocalization analysis revealed that only 73.74% ± 1.76% of the PSV-S transduction depended on chiACE2. Furthermore, type 2 innate lymphoid cells (ILC2s) exhibited the most pronounced activation in the gut mucosa. This finding indicates the significant role of ILC2s as contributors to the mucosal defense against viral infections. Our data shed light on the critical involvement of ILC2s and their intricate three-dimensional regulatory network in combating SARS-CoV-2 within the mucosa. IMPORTANCE Our study revealed the spatial interaction between humanized ACE2 and pseudovirus expressing Spike, emphasizing the role of type 2 innate lymphoid cells during the initial phase of viral infection. These findings provide a foundation for the development of mucosal vaccines and other treatment approaches for both pre- and post-infection management of coronavirus disease 2019.

Secretory IgA impacts the microbiota density in the human nose

BackgroundRespiratory mucosal host defense relies on the production of secretory IgA (sIgA) antibodies, but we currently lack a fundamental understanding of how sIgA is induced by contact with microbes and how such immune responses may vary between humans. Defense of the nasal mucosal barrier through sIgA is critical to protect from infection and to maintain homeostasis of the microbiome, which influences respiratory disorders and hosts opportunistic pathogens.MethodsWe applied IgA-seq analysis to nasal microbiota samples from male and female healthy volunteers, to identify which bacterial genera and species are targeted by sIgA on the level of the individual host. Furthermore, we used nasal sIgA from the same individuals in sIgA deposition experiments to validate the IgA-seq outcomes.ConclusionsWe observed that the amount of sIgA secreted into the nasal mucosa by the host varied substantially and was negatively correlated with the bacterial density, suggesting that nasal sIgA limits the overall bacterial capacity to colonize. The interaction between mucosal sIgA antibodies and the nasal microbiota was highly individual with no obvious differences between potentially invasive and non-invasive bacterial species. Importantly, we could show that for the clinically relevant opportunistic pathogen and frequent nasal resident Staphylococcus aureus, sIgA reactivity was in part the result of epitope-independent interaction of sIgA with the antibody-binding protein SpA through binding of sIgA Fab regions. This study thereby offers a first comprehensive insight into the targeting of the nasal microbiota by sIgA antibodies. It thereby helps to better understand the shaping and homeostasis of the nasal microbiome by the host and may guide the development of effective mucosal vaccines against bacterial pathogens.7hP8wdG6y5p5ZRVJGsDS4ZVideo

Utility of nasal swabs for assessing mucosal immune responses towards SARS-CoV-2

SARS-CoV-2 has caused millions of infections worldwide since its emergence in 2019. Understanding how infection and vaccination induce mucosal immune responses and how they fluctuate over time is important, especially since they are key in preventing infection and reducing disease severity. We established a novel methodology for assessing SARS-CoV-2 cytokine and antibody responses at the nasal epithelium by using nasopharyngeal swabs collected longitudinally before and after either SARS-CoV-2 infection or vaccination. We then compared responses between mucosal and systemic compartments. We demonstrate that cytokine and antibody profiles differ between compartments. Nasal cytokines show a wound healing phenotype while plasma cytokines are consistent with pro-inflammatory pathways. We found that nasal IgA and IgG have different kinetics after infection, with IgA peaking first. Although vaccination results in low nasal IgA, IgG induction persists for up to 180 days post-vaccination. This research highlights the importance of studying mucosal responses in addition to systemic responses to respiratory infections. The methods described herein can be used to further mucosal vaccine development by giving us a better understanding of immunity at the nasal epithelium providing a simpler, alternative clinical practice to studying mucosal responses to infection.

Development of a spore-based mucosal vaccine against the bovine respiratory pathogen Mannheimia haemolytica

Bovine respiratory disease (BRD) is a significant health issue in the North American feedlot industry, causing substantial financial losses due to morbidity and mortality. A lack of effective vaccines against BRD pathogens has resulted in antibiotics primarily being used for BRD prevention. The aim of this study was to develop a mucosal vaccine against the BRD pathogen, Mannheimia haemolytica, using Bacillus subtilis spores as an adjuvant. A chimeric protein (MhCP) containing a tandem repeat of neutralizing epitopes from M. haemolytica leukotoxin A (NLKT) and outer membrane protein PlpE was expressed to produce antigen for adsorption to B. subtilis spores. Adsorption was optimized by comparing varying amounts of antigen and spores, as well as different buffer pH and reaction temperatures. Using the optimal adsorption parameters, spore-bound antigen (Spore-MhCP) was prepared and administered to mice via two mucosal routes (intranasal and intragastric), while intramuscular administration of free MhCP and unvaccinated mice were used as positive and negative control treatments, respectively. Intramuscular administration of MhCP elicited the strongest serum IgG response. However, intranasal immunization of Spore-MhCP generated the best secretory IgA-specific response against both PlpE and NLKT in all samples evaluated (bronchoalveolar lavage, saliva, and feces). Since proliferation of M. haemolytica in the respiratory tract is a prerequisite to lung infection, this spore-based vaccine may offer protection in cattle by limiting colonization and subsequent infection, and Spore-MhCP warrants further evaluation in cattle as a mucosal vaccine against M. haemolytica.