Antigen-driven Convergent Evolution of Polysaccharide-specific "DH-less" B Cells in Glycoconjugate Immunized Mice.

Glycoconjugate vaccines elicit robust anti-polysaccharide Ab response by recruiting T-cell help. Multiple doses of glycoconjugate vaccine are required to induce long-lasting immunity. The characteristics of anti-polysaccharide Ab response have been reported previously. However, the effect of glycoconjugate booster immunization on anti-polysaccharide and anti-carrier protein Ab repertoire remains poorly understood. In this study, we used clinically relevant pneumococcal capsular polysaccharide type 14 (PCP14) conjugated with cross-reactive material 197 (CRM197) as a model glycoconjugate Ag (PCP14-CRM197). We performed a comprehensive sequence analysis of mouse mAbs generated against PCP14 and CRM197 following immunization with one or three doses of PCP14-CRM197. Analysis of the paired Ig H and L chain transcripts revealed that anti-PCP14 Ab repertoire is extremely restricted. The reoccurrence of five replacement mutations at identical positions in anti-polysaccharide mAbs generated from different mice provided evidence for Ag-driven selection in PCP14-specific B cells. Convergent evolution was observed wherein distinct V(D)J rearrangements resulted in identical or nearly identical CDR3 in anti-PCP14 mAbs. Abs that lacked DH encoded amino acids dominated the anti-PCP14 Ab response. In contrast, anti-CRM197 Ab response was quite diverse, with fewer mutations compared with the anti-PCP14 mAbs, suggesting that conjugation of the polysaccharide to a carrier protein interferes with the development of carrier protein-specific Ab responses. Our findings provide molecular insights into the maturation of Ab responses driven by booster doses of glycoconjugate. This has fundamental implications for the design of glycoconjugate vaccines, especially where the development of Ab response against the carrier protein is also crucial.

Analyses of the association between Helicobacter pylori antibody titre and pathogenicity before and after eradication: results of the Kyushu and Okinawa population study, a retrospective observational cohort study.

OBJECTIVES: To assess the utility of Helicobacter pylori antibody testing, we evaluated the correlation between the H. pylori antibody titre and H. pylori-associated pathogenicity and the changes in antibody titre after H. pylori eradication therapy. DESIGN: A retrospective observational cohort study. SETTING AND PARTICIPANTS: From 2004 to 2016, medical check-ups were performed in different regions of Japan. In total, 324 subjects infected with H. pylori who received H. pylori eradication therapy were enrolled; H. pylori was eradicated in 266 of these subjects. We examined the associations between H. pylori antibody titre with pepsinogen and the presence or absence of H. pylori-associated pathogenic proteins, such as cytotoxin-associated gene A and vacuolating cytotoxin gene A, at baseline and after H. pylori eradication therapy. RESULTS: The H.pylori antibody titre showed a positive correlation with pepsinogen II and a negative correlation with the pepsinogen I/II ratio. Moreover, the H.pylori antibody titre significantly correlated with the positive rates of H. pylori-associated pathogenic protein before eradication therapy. Antibody titres decreased after eradication, the pepsinogen I/II ratio increased and the H. pylori-associated pathogenic protein-positive rate decreased in patients with successful eradication. The determination of eradication using the decline in antibody titre 6 months after eradication therapy was useful (area under the receiver operating characteristic curve: 0.98). CONCLUSIONS: Our data indicate that the H. pylori antibody titre may represent the degree of pathogenicity. The H. pylori antibody titre was associated with attenuation of pathogenicity in patients with H. pylori eradication, indicating the clinical utility of H. pylori antibody testing.

Immunoinformatics design and synthesis of a multi-epitope vaccine against Helicobacter pylori based on lipid nanoparticles.

Helicobacter pylori (H. pylori) is responsible for various chronic or acute diseases, such as stomach ulcers, dyspepsia, peptic ulcers, gastroesophageal reflux, gastritis, lymphoma, and stomach cancers. Although specific drugs are available to treat the bacterium's harmful effects, there is an urgent need to develop a preventive or therapeutic vaccine. Therefore, the current study aims to create a multi-epitope vaccine against H. pylori using lipid nanoparticles. Five epitopes from five target proteins of H. pylori, namely, Urease, CagA, HopE, SabA, and BabA, were used. Immunogenicity, MHC (Major Histocompatibility Complex) bonding, allergenicity, toxicity, physicochemical analysis, and global population coverage of the entire epitopes and final construct were carefully examined. The study involved using various bioinformatic web tools to accomplish the following tasks: modeling the three-dimensional structure of a set of epitopes and the final construct and docking them with Toll-Like Receptor 4 (TLR4). In the experimental phase, the final multi-epitope construct was synthesized using the solid phase method, and it was then enclosed in lipid nanoparticles. After synthesizing the construct, its loading, average size distribution, and nanoliposome shape were checked using Nanodrop at 280 nm, dynamic light scattering (DLS), and atomic force microscope (AFM). The designed vaccine has been confirmed to be non-toxic and anti-allergic. It can bind with different MHC alleles at a rate of 99.05%. The construct loading was determined to be about 91%, with an average size of 54 nm. Spherical shapes were also observed in the AFM images. Further laboratory tests are necessary to confirm the safety and immunogenicity of the multi-epitope vaccine.

From pathogenesis to antigens: the key to shaping the future of TB vaccines.

Tuberculosis (TB) remains one of the gravest global health challenges. Mycobacterium tuberculosis (M. tuberculosis), the causative agent, employs sophisticated immune evasion and pathogenesis strategies. Its capability to thrive within immune cells and incite robust inflammatory responses prolongs infection and dissemination. Mycobacterial advanced adaptations facilitate navigation through the human immune system and present a variable antigenic profile throughout different infection stages. Investigating these strategies unfolds targeted approaches to effective vaccine development against TB. This review delves into the most advanced and exhaustive insights into the immune evasion tactics and pathogenic processes of M. tuberculosis across various infection stages. The knowledge distilled from this analysis holds the promise of guiding the creation of innovative TB vaccines and translating theoretical groundwork into practical immunological defenses.

Laboratory diagnosis and treatment of Mycoplasma pneumoniae infection in children: a review.

Mycoplasma pneumoniae (MP) is the cause of Mycoplasma pneumoniae pneumonia (MPP) in children and adolescents, with the clinical manifestations highlighted by intermittent irritating cough, accompanied by headache, fever and muscle pain. This paper aimed to study the research status and focal points in MP infection, especially the common laboratory diagnostic methods and clinical treatment of Mycoplasma pneumoniae. Laboratory diagnostic methods include molecular assay, serological antibody detection, rapid antigen detection and isolation and culture. Polymerase chain reaction (PCR) is the gold standard with high sensitivity and specificity. The serological antibody can detect various immune antibodies qualitatively or quantitatively in serum. Rapid antigen can be detected faster, with no equipment environment requirements, which can be used for the early diagnosis of MP infection. While the culture growth cycle is long and insensitive, not recommended for routine diagnosis. Macrolides were the preferred drug for children with MPP, while the drug resistance rate was rising in China. Tetracycline can be substituted but was not recommended for children under 8 years of age, quinolone drugs are not necessary, severe MPP can be combined with glucocorticoids, involving the nervous or immune system can choose gamma globulin. Other treatments for MPP including symptomatic treatment which can alleviate symptoms, improve lung function and improve prognosis. A safe and effective vaccine needed to be developed which can provide protective immunity to children and will reduce the incidence of MPP.

Assay for interferon gamma release as a novel marker in pediatric patients with systemic lupus erythematosus.

BACKGROUND: The interferon-gamma (IFN-γ) release assay (IGRA) is an important laboratory diagnosis for latent Mycobacterium tuberculosis (TB) infection. The TB-IGRA measures the release of IFN-γ from peripheral blood cells, who are exposed to TB antigen (Ag), mitogen (MT), or negative/nil control (NL) in vitro. While, an exceptional higher TB Ag-NL level will reflect an elevation of peripheral lymphocytes released IFN-γ in a same condition. Therefore, we found that the elevated levels of TB Ag-NL could become a new biomarker for the diagnosis and treatment of pediatric systemic lupus erythematosus (SLE) patients. METHODS: We have analyzed the clinical data of 776 children who are underwent TB-IGRA testing in the Department of Allergy and Rheumatology of Guangzhou Women and Children's Medical Center from 2018 to 2020. To investigate the association between TB Ag-NL and SLE, we have analyzed the clinical data of 47 SLE patients and TB Ag-NL testing results, and then evaluated the association between TB Ag-NL and SLE disease activity. RESULTS: The TB Ag-NL levels were significantly higher in patients with active SLE than those in inactive SLE (p = 0.0002). The TB Ag-NL levels were positively correlated with the SLE disease activity index (SLEDAI) and laboratory diagnosis parameters. The mean value of TB Ag-NL in SLE patients (0.04191 ± 0.07955, IU/mL) were significantly higher than those in patients with juvenile dermatomyositis (JDM) (0.0158 ± 0.0337, IU/mL, p = 0.036), juvenile idiopathic arthritis (JIA) (0.0162 ± 0.0388, IU/mL, p = 0.001), and healthy controls (HC) (0.0001 ± 0.0027, IU/mL, p = 0.0003). Therefore, the elevated TB Ag-NL levels could serve as a potential diagnostic biomarker of SLE, especially for the active SLE. CONCLUSION: The detection of IFN-γ release levels by the TB-IGRA may be useful to assess SLE disease activity in pediatric patients with active SLE.

Profiling IgG and IgA antibody responses during vaccination and infection in a high-risk gonorrhoea population.

Development of a vaccine against gonorrhoea is a global priority, driven by the rise in antibiotic resistance. Although Neisseria gonorrhoeae (Ng) infection does not induce substantial protective immunity, highly exposed individuals may develop immunity against re-infection with the same strain. Retrospective epidemiological studies have shown that vaccines containing Neisseria meningitidis (Nm) outer membrane vesicles (OMVs) provide a degree of cross-protection against Ng infection. We conducted a clinical trial (NCT04297436) of 4CMenB (Bexsero, GSK), a licensed Nm vaccine containing OMVs and recombinant antigens, comprising a single arm, open label study of two doses with 50 adults in coastal Kenya who have high exposure to Ng. Data from a Ng antigen microarray established that serum IgG and IgA reactivities against the gonococcal homologs of the recombinant antigens in the vaccine peaked at 10 but had declined by 24 weeks. For most reactive OMV-derived antigens, the reverse was the case. A cohort of similar individuals with laboratory-confirmed gonococcal infection were compared before, during, and after infection: their reactivities were weaker and differed from the vaccinated cohort. We conclude that the cross-protection of the 4CMenB vaccine against gonorrhoea could be explained by cross-reaction against a diverse selection of antigens derived from the OMV component.

A study of antigen selection by extracellular vesicles as vaccine candidates against Mycobacterium tuberculosis infection.

Introduction. Tuberculosis (TB), an infectious disease caused by Mycobacterium tuberculosis (M. tb), remains a significant global public health concern. It is crucial to develop more effective vaccines for TB in order to achieve global control of the disease. Extracellular vesicles (EVs) are spherical membrane-bound structures released by pathogens and host cells. During the course of an infection, both pathogen- and host-derived EVs are produced and play important roles in determining the course of the infection. EVs offer intriguing tools as potential vaccines due to their ability to deliver multiple pathogen or host antigens.Hypothesis /Gap Statement. We hypothesized that EVs derived from M. tb and EVs from M. tb-infected macrophages may serve as potential vaccine candidates against M. tb infection.Aim. This study aims to compare the immunogenicity and immune protection between M. tb EVs and M. tb-infected macrophage-derived EVs.Methodology. In this study, EVs were extracted from culture supernatants of M. tb and M. tb-infected macrophages, respectively. Mass spectrometry was employed to explore the antigen composition of H37Rv-Mφ-EVs and H37Rv-EVs. Cytokine profiling and antibody response assays were used to analyse the immunogenicity offered by EVs. Additionally, we used histological examination to evaluate and protective efficacy of the EVs.Results. Our results demonstrated that mice immunized by EVs released from M. tb-infected macrophages induced stronger inflammatory cytokine response than M. tb. Moreover, EVs from M. tb-infected macrophages reinforced T-cell activation and antibody response compared to M. tb EVs. Proteomic analysis revealed that EVs from M. tb-infected macrophages containing immunodominant cargos have stronger binding ability with major histocompatibility complex molecules, which may contribute to the protection from M. tb infection. Indeed, immunization of EVs released from M. tb-infected macrophages significantly reduced the bacterial load and better protection against M. tb infection than EVs from M. tb. Importantly, the selected antigens (Ag85B, ESAT-6 and the Rv0580c) from EVs of M. tb-infected macrophages exhibited effective immunogenicity.Conclusion. Our results suggested that EVs derived from M. tb-infected macrophages might serve as a proper antigenic library for vaccine candidates against M. tb challenge.

A novel mRNA multi-epitope vaccine of Acinetobacter baumannii based on multi-target protein design in immunoinformatic approach.

Acinetobacter baumannii is a gram-negative bacillus prevalent in nature, capable of thriving under various environmental conditions. As an opportunistic pathogen, it frequently causes nosocomial infections such as urinary tract infections, bacteremia, and pneumonia, contributing to increased morbidity and mortality in clinical settings. Consequently, developing novel vaccines against Acinetobacter baumannii is of utmost importance. In our study, we identified 10 highly conserved antigenic proteins from the NCBI and UniProt databases for epitope mapping. We subsequently screened and selected 8 CTL, HTL, and LBL epitopes, integrating them into three distinct vaccines constructed with adjuvants. Following comprehensive evaluations of immunological and physicochemical parameters, we conducted molecular docking and molecular dynamics simulations to assess the efficacy and stability of these vaccines. Our findings indicate that all three multi-epitope mRNA vaccines designed against Acinetobacter baumannii are promising; however, further animal studies are required to confirm their reliability and effectiveness.

Comparative analysis of natural resistance-associated macrophage protein 1 gene expression and anti-PGL-1 antibodies in multibacillary leprosy and household contacts.

Leprosy continues to pose a significant challenge to public health, particularly in certain global regions. Accurate diagnosis and understanding of the disease's etiology are crucial for effective management and prevention. This study aimed to explore the contribution of Natural resistance-associated macrophage protein 1 (NRAMP1) and its genetic variations, as well as the levels of anti-PGL-1 antibodies, to the pathology of multibacillary leprosy in affected individuals and their household contacts. The study included 23 multibacillary leprosy patients and 28 household contacts. NRAMP1 protein expression and anti-PGL-1 IgG and IgM levels were measured using PCR and ELISA techniques, respectively. Genotypic variants of the NRAMP1 gene were also examined. Statistical analyses, including Mann-Whitney tests and univariate logistic regression, were employed to evaluate the data. Significant differences were observed in NRAMP1 protein expression and IgG and IgM levels between the patient and household contact groups. The study also highlighted the role of the NRAMP1 gene and its D543N and 3'UTR polymorphisms in leprosy susceptibility. No significant differences were observed in the genotype variants of INT4 between the two groups. These findings emphasize the potential of integrating PCR technology with serological tests to enhance diagnostic precision in leprosy. They also suggest the need for further research to clarify the role of NRAMP1 and its polymorphisms in leprosy susceptibility and resistance.

On-Demand Vaccine Production via Dock-and-Display of Biotinylated Antigens on Bacterial Extracellular Vesicles.

Engineered outer membrane vesicles (OMVs) derived from Gram-negative bacteria are a promising vaccine technology for developing immunity against diverse pathogens. However, antigen display on OMVs can be challenging to control and highly variable due to bottlenecks in protein expression and localization to the bacterial host cell's outer membrane, especially for bulky and complex antigens. Here, we describe methods related to a universal vaccine technology called AvidVax (avidin-based vaccine antigen crosslinking) for rapid and simplified assembly of antigens on the exterior of OMVs during vaccine development. The AvidVax platform involves remodeling the OMV surface with multiple copies of a synthetic antigen-binding protein (SNAP), which is an engineered fusion protein comprised of an outer membrane scaffold protein linked to a biotin-binding protein. The resulting SNAPs enable efficient decoration of OMVs with a molecularly diverse array of biotinylated subunit antigens, including globular and membrane proteins, glycans and glycoconjugates, haptens, lipids, nucleic acids, and short peptides. We detail the key steps in the AvidVax vaccine production pipeline including preparation and isolation of SNAP-OMVs, biotinylation and enrichment of vaccine antigens, and formulation and characterization of antigen-loaded SNAP-OMVs.

Harnessing microbial antigens as cancer antigens: a promising avenue for cancer immunotherapy.

Immunotherapy has revolutionized cancer treatment by leveraging the immune system's innate capabilities to combat malignancies. Despite the promise of tumor antigens in stimulating anti-tumor immune responses, their clinical utility is hampered by limitations in eliciting robust and durable immune reactions, exacerbated by tumor heterogeneity and immune evasion mechanisms. Recent insights into the immunogenic properties of host homologous microbial antigens have sparked interest in their potential for augmenting anti-tumor immunity while minimizing off-target effects. This review explores the therapeutic potential of microbial antigen peptides in tumor immunotherapy, beginning with an overview of tumor antigens and their challenges in clinical translation. We further explore the intricate relationship between microorganisms and tumor development, elucidating the concept of molecular mimicry and its implications for immune recognition of tumor-associated antigens. Finally, we discuss methodologies for identifying and characterizing microbial antigen peptides, highlighting their immunogenicity and prospects for therapeutic application.

Investigation of cross-opsonic effect leads to the discovery of PPIase-domain containing protein vaccine candidate to prevent infections by Gram-positive ESKAPE pathogens.

BACKGROUND: Enterococcus faecium and Staphylococcus aureus are the Gram-positive pathogens of the ESKAPE group, known to represent a great threat to human health due to their high virulence and multiple resistances to antibiotics. Combined, enterococci and S. aureus account for 26% of healthcare-associated infections and are the most common organisms responsible for blood stream infections. We previously showed that the peptidyl-prolyl cis/trans isomerase (PPIase) PpiC of E. faecium elicits the production of specific, opsonic, and protective antibodies that are effective against several strains of E. faecium and E. faecalis. Due to the ubiquitous characteristics of PPIases and their essential function within Gram-positive cells, we hypothesized a potential cross-reactive effect of anti-PpiC antibodies. RESULTS: Opsonophagocytic assays combined with bioinformatics led to the identification of the foldase protein PrsA as a new potential vaccine antigen in S. aureus. We show that PrsA is a stable dimeric protein able to elicit opsonic antibodies against the S. aureus strain MW2, as well as cross-binding and cross-opsonic in several S. aureus, E. faecium and E. faecalis strains. CONCLUSIONS: Given the multiple antibiotic resistances S. aureus and enterococci present, finding preventive strategies is essential to fight those two nosocomial pathogens. The study shows the potential of PrsA as an antigen to use in vaccine formulation against the two dangerous Gram-positive ESKAPE bacteria. Our findings support the idea that PPIases should be further investigated as vaccine targets in the frame of pan-vaccinomics strategy.

Overlooked, dismissed, and downplayed: reversion of Mycobacterium tuberculosis immunoreactivity.

Tuberculosis (TB) is caused by Mycobacterium tuberculosis (Mtb). Following infection, immune responses to Mtb antigens can be measured using the tuberculin skin test or an interferon-γ release assay. The gain of Mtb immunoreactivity, a change from a negative to a positive tuberculin skin test or interferon-γ release assay result, is called conversion and has long been used as a measure of Mtb exposure. However, the loss of immunoreactivity (reversion; a positive followed by a negative result) has often been overlooked. Instead, in clinical and epidemiological circles, Mtb immunoreactivity is commonly considered to persist lifelong and confer a lifetime of disease risk. We present a critical review, describing the evidence for reversion from cohort studies, ecological studies and studies of TB progression risk. We outline the inconsistent reasons why reversion has been dismissed from common understanding and present evidence demonstrating that, just as conversion predominantly indicates prior exposure to Mtb antigens, so its opposite, reversion, suggests the reduction or absence of exposure (endogenous or exogenous). Mtb immunoreactivity is dynamic in both individuals and populations and this is why it is useful for stratifying short-term TB progression risk. The neglect of reversion has shaped TB research and policy at all levels, influencing clinical management and skewing Mtb infection risk estimation and transmission modelling, leading to an underestimation of the contribution of re-exposure to the burden of TB, a serious oversight for an infectious disease. More than a century after it was first demonstrated, it is time to incorporate reversion into our understanding of the natural history of TB.

Immunoinformatics and structural aided approach to develop multi-epitope based subunit vaccine against Mycobacterium tuberculosis.

Tuberculosis is a highly contagious disease caused by Mycobacterium tuberculosis (Mtb), which is one of the prominent reasons for the death of millions worldwide. The bacterium has a substantially higher mortality rate than other bacterial diseases, and the rapid rise of drug-resistant strains only makes the situation more concerning. Currently, the only licensed vaccine BCG (Bacillus Calmette-Guérin) is ineffective in preventing adult pulmonary tuberculosis prophylaxis and latent tuberculosis re-activation. Therefore, there is a pressing need to find novel and safe vaccines that provide robust immune defense and have various applications. Vaccines that combine epitopes from multiple candidate proteins have been shown to boost immunity against Mtb infection. This study applies an immunoinformatic strategy to generate an adequate multi-epitope immunization against Mtb employing five antigenic proteins. Potential B-cell, cytotoxic T lymphocyte, and helper T lymphocyte epitopes were speculated from the intended proteins and coupled with 50 s ribosomal L7/L12 adjuvant, and the vaccine was constructed. The vaccine's physicochemical profile demonstrates antigenic, soluble, and non-allergic. In the meantime, docking, molecular dynamics simulations, and essential dynamics analysis revealed that the multi-epitope vaccine structure interacted strongly with Toll-like receptors (TLR2 and TLR3). MM-PBSA analysis was performed to ascertain the system's intermolecular binding free energies accurately. The immune simulation was applied to the vaccine to forecast its immunogenic profile. Finally, in silico cloning was used to validate the vaccine's efficacy. The immunoinformatics analysis suggests the multi-epitope vaccine could induce specific immune responses, making it a potential candidate against Mtb. However, validation through the in-vivo study of the developed vaccine is essential to assess its efficacy and immunogenicity profile, which will assure active protection against Mtb.

Putative new combination vaccine candidates identified by reverse vaccinology and genomic approaches to control enteric pathogens.

OBJECTIVES: The pathogenic microorganisms that cause intestinal diseases can significantly jeopardize people's health. Currently, there are no authorized treatments or vaccinations available to combat the germs responsible for intestinal disease. METHODS: Using immunoinformatics, we developed a potent multi-epitope Combination (combo) vaccine versus Salmonella and enterohemorrhagic E. coli. The B and T cell epitopes were identified by performing a conservancy assessment, population coverage analysis, physicochemical attributes assessment, and secondary and tertiary structure assessment of the chosen antigenic polypeptide. The selection process for vaccine development included using several bioinformatics tools and approaches to finally choose two linear B-cell epitopes, five CTL epitopes, and two HTL epitopes. RESULTS: The vaccine had strong immunogenicity, cytokine production, immunological properties, non-toxicity, non-allergenicity, stability, and potential efficacy against infections. Disulfide bonding, codon modification, and computational cloning were also used to enhance the stability and efficacy of expression in the host E. coli. The vaccine's structure has a strong affinity for the TLR4 ligand and is very durable, as shown by molecular docking and molecular modeling. The results of the immunological simulation demonstrated that both B and T cells had a heightened response to the vaccination component. CONCLUSIONS: The comprehensive in silico analysis reveals that the proposed vaccine will likely elicit a robust immune response against pathogenic bacteria that cause intestinal diseases. Therefore, it is a promising option for further experimental testing.

Targeting plasmid-encoded proteins that contain immunoglobulin-like domains to combat antimicrobial resistance.

Antimicrobial resistance (AMR) poses a significant threat to human health. Although vaccines have been developed to combat AMR, it has proven challenging to associate specific vaccine antigens with AMR. Bacterial plasmids play a crucial role in the transmission of AMR. Our recent research has identified a group of bacterial plasmids (specifically, IncHI plasmids) that encode large molecular mass proteins containing bacterial immunoglobulin-like domains. These proteins are found on the external surface of the bacterial cells, such as in the flagella or conjugative pili. In this study, we show that these proteins are antigenic and can protect mice from infection caused by an AMR Salmonella strain harboring one of these plasmids. Furthermore, we successfully generated nanobodies targeting these proteins, that were shown to interfere with the conjugative transfer of IncHI plasmids. Considering that these proteins are also encoded in other groups of plasmids, such as IncA/C and IncP2, targeting them could be a valuable strategy in combating AMR infections caused by bacteria harboring different groups of AMR plasmids. Since the selected antigens are directly linked to AMR itself, the protective effect extends beyond specific microorganisms to include all those carrying the corresponding resistance plasmids.

HLA-A02 restricted T-cell cross-reactivity to a microbial antigen.

Molecular mimicry has been proposed to be a possible mechanism of induction of autoimmunity. In some cases, it is believed that such events could lead to a disease such as Type 1 diabetes (T1D). One of the primary MHC-I epitopes in the non-obese diabetic (NOD) mouse model of T1D has been identified as a peptide from the islet-specific glucose-6-phosphatase catalytic subunit-related protein (IGRP) protein. In humans, the most common MHC-I model allele is HLA-A02; based on this, the study here identified a potential HLA-A0201-restricted human IGRP epitope as YLKTNLFLFL and also found a homologous A0201-restricted peptide in an Enterococcal protein. Using cells obtained from healthy human donors, it was seen that after a 2-week incubation with the synthetic bacterial protein, healthy A0201+ donor CD8+ cells displayed increased staining for human IGRP-peptide-dextramer. On the other hand, in control cultures, no significant levels of dextramer-staining CD8+ T-cells were detectable. From these outcomes, it is possible to conclude that certain bacterial proteins may initiate CD8+ T-cell-mediated immune reaction toward homologous human antigens.

The innate memory response of macrophages to Mycobacterium tuberculosis is shaped by the nature of the antigenic stimuli.

Innate immune cells, such as macrophages, mount an immune response upon exposure to antigens and pathogens. Emerging evidence shows that macrophages exposed to an antigen can generate a "memory-like" response (a.k.a. trained immunity), which confers a non-specific and enhanced response upon subsequent stimulation with a second antigen/microbe. This trained immunity has been implicated in the enhanced response of macrophages against several invading pathogens. However, the association between the nature of the antigen and the corresponding immune correlate of elicited trained immunity is not fully understood. Similarly, the response of macrophages trained and restimulated with homologous stimulants to subsequent infection by pathogenic Mycobacterium tuberculosis (Mtb) remains unexplored. Here, we report the immune and metabolic profiles of trained immunity in human THP-1-derived macrophages after homologous training and restimulation with BCG, LPS, purified protein Derivative (PPD), heat-killed Mtb strains HN878 (hk-HN), and CDC1551 (hk-CDC). Furthermore, the impact of training on the autophagic and antimicrobial responses of macrophages with or without subsequent infection by clinical Mtb isolates HN878 and CDC1551 was evaluated. Results show that repeated stimulation of macrophages with different antigens displays distinct pro-inflammatory, metabolic, antimicrobial, and autophagy induction profiles. These macrophages also induce a differential antimicrobial response upon infection with clinical Mtb HN878 and CDC1551 isolates. A significantly reduced intracellular bacterial load was noted in the stimulated macrophages, which was augmented by the addition of rapamycin, an autophagy inducer. These observations suggest that the nature of the antigen and the mode of stimulation shape the magnitude and breadth of macrophage innate memory response, which impacts subsequent response to Mtb infection. IMPORTANCE: Trained immunity (a.k.a. innate memory response) is a novel concept that has been rapidly emerging as a mechanism underpinning the non-specific immunity of innate immune cells, such as macrophages. However, the association between the nature of the stimuli and the corresponding immune correlate of trained immunity is not fully understood. Similarly, the kinetics of immunological and metabolic characteristics of macrophages upon "training" by the same antigen as primary and secondary stimuli (homologous stimulation) are not fully characterized. Furthermore, the ability of antigens such as purified protein derivative (PPD) and heat-killed-Mtb to induce trained immunity remains unknown. Similarly, the response of macrophages primed and trained by homologous stimulants to subsequent infection by pathogenic Mtb is yet to be reported. In this study, we evaluated the hypothesis that the nature of the stimuli impacts the depth and breadth of trained immunity in macrophages, which differentially affects their response to Mtb infection.

Immunogenicity and vaccine potential of clinical isolate Mycobacterium kansasii strain against Mycobacterium tuberculosis infection.

Mycobacterium kansasii is a bacterium included in non-tuberculous mycobacteria (NTM) that can cause lung disease. It shares a significant number of antigens with Mycobacterium tuberculosis (Mtb), suggesting that it has the potential to be used as a tuberculosis (TB) vaccine. Therefore, we subcutaneously vaccinated mice with reference strain, M. kansasii-ATCC12478 [M. kansasii-American Type Culture Collection (ATCC)], and clinically isolated strain, M. kansasii-SM-1 to evaluate potential as a TB vaccine by comparing with bacille Calmette-Guerin (BCG) vaccine. Ten weeks after vaccination, we evaluated immunogenicity of M. kansasii-ATCC and M. kansasii-SM-1, and M. kansasii-SM-1 immunization induces potent Mtb antigen-specific IFN-γ-producing CD4+ T cells than M. kansasii-ATCC. Upon Mtb infection, M. kansasii-SM-1 provided better protection than M. kansasii-ATCC, which was comparable to the efficacy of BCG. These results showed that the clinical strain M. kansasii-SM-1, which exhibits an enhanced Mtb antigen-specific Th1 response, shows greater vaccine efficacy compared to M. kansasii-ATCC. In this study, we demonstrated that vaccine efficacy can vary depending on the strain of M. kansasii and that its efficacy can be comparable to BCG. This suggests that M. kansasii has the potential to be a live TB vaccine candidate.IMPORTANCEMycobacterium kansasii, a non-tuberculous mycobacteria (NTM) species causing lung disease, shares key antigens with Mycobacterium tuberculosis (Mtb), indicating its potential for TB vaccine development. Subcutaneous vaccination of mice with M. kansasii strains reference strain M. kansasii-ATCC12478 [(M. kansasii-American Type Culture Collection (ATCC)] and clinically isolated strain M. kansasii-SM-1 revealed differences in immunogenicity. M. kansasii-SM-1 induced a robust Mtb antigen-specific IFN-γ-producing CD4+ T cell response compared to M. kansasii-ATCC. Additionally, M. kansasii-SM-1 conferred better protection against Mtb infection than M. kansasii-ATCC, which is comparable to bacille Calmette-Guerin (BCG). These findings underscore the variable vaccine efficacy among M. kansasii strains, with M. kansasii-SM-1 exhibiting promising potential as a live TB vaccine candidate, suggesting its comparative effectiveness to BCG.