Synthetic BSA-conjugated disaccharide related to the Streptococcus pneumoniae serotype 3 capsular polysaccharide increases IL-17A Levels, γδ T cells, and B1 cells in mice.

The disaccharide (ß-D-glucopyranosyluronic acid)-(1→4)-ß-D-glucopyranoside represents a repeating unit of the capsular polysaccharide of Streptococcus pneumoniae serotype 3. A conjugate of the disaccharide with BSA (di-BSA conjugate) adjuvanted with aluminum hydroxide induced - in contrast to the non-adjuvanted conjugate - IgG1 antibody production and protected mice against S. pneumoniae serotype 3 infection after intraperitoneal prime-boost immunization. Adjuvanted and non-adjuvanted conjugates induced production of Th1 (IFNγ, TNFα); Th2 (IL-5, IL-13); Th17 (IL-17A), Th1/Th17 (IL-22), and Th2/Th17 cytokines (IL-21) after immunization. The concentration of cytokines in mice sera was higher in response to the adjuvanted conjugate, with the highest level of IL-17A production after the prime and boost immunizations. In contrast, the non-adjuvanted conjugate elicited only weak production of IL-17A, which gradually decreased after the second immunization. After boost immunization of mice with the adjuvanted di-BSA conjugate, there was a significant increase in the number of CD45+/CD19+ B cells, TCR+ γδ T cell, CD5+ В1 cells, and activated cells with MHC II+ expression in the spleens of the mice. IL-17A, TCR+ γδ T cells, and CD5+ В1 cells play a crucial role in preventing pneumococcal infection, but can also contribute to autoimmune diseases. Immunization with the adjuvanted and non-adjuvanted di-BSA conjugate did not elicit autoantibodies against double-stranded DNA targeting cell nuclei in mice. Thus, the molecular and cellular markers associated with antibody production and protective activity in response to immunization with the di-BSA conjugate adjuvanted with aluminum hydroxide are IL-17A, TCR+ γδ T cells, and CD5+ В1 cells against the background of increasing MHC II+ expression.

A combination influenza mRNA vaccine candidate provided broad protection against diverse influenza virus challenge.

Influenza viruses present a significant threat to global health. The production of a universal vaccine is considered essential due to the ineffectiveness of current seasonal influenza vaccines against mutant strains. mRNA technology offers new prospects in vaccinology, with various candidates for different infectious diseases currently in development and testing phases. In this study, we encapsulated a universal influenza mRNA vaccine. The vaccine encoded influenza hemagglutinin (HA), nucleoprotein (NP), and three tandem repeats of matrix protein 2 (3M2e). Twice-vaccinated mice exhibited strong humoral and cell-mediated immune responses in vivo. Notably, these immune responses led to a significant reduction in viral load of the lungs in challenged mice, and also conferred protection against future wild-type H1N1, H3N2, or H5N1 influenza virus challenges. Our findings suggest that this mRNA-universal vaccine strategy for influenza virus may be instrumental in mitigating the impact of future influenza pandemics.

Vaccine and Non-Vaccine HPV Types Presence in Adolescents with Vertically Acquired HIV Five Years Post Gardasil Quadrivalent Vaccination: The ZIMGARD Cohort.

BACKGROUND: Human papillomavirus (HPV) vaccination programs are a key intervention in protecting individuals against HPV-related disease. HIV1-infected individuals are at increased risk of HPV-associated cancers. This study was conducted to evaluate the potential role of prophylactic HPV vaccines in preventing new HPV infections among participants with perinatally acquired HIV who received the quadrivalent HPV vaccine at least five years before this study. METHODS: This cross-sectional study was conducted at Newlands Clinic, Harare, Zimbabwe. The clinic provided the Gardasil quadrivalent HPV vaccine (4vHPV) to 624 adolescents living with HIV starting in December 2015. Vaginal and penile swabs were collected and tested for HPV types from the study participants who had received the 4vHPV vaccine 5-6 years before enrolment. RESULTS: We present the results of 98 participants (44.6% female) vaccinated at a median age of 15 years (IQR 12-16). The mean amount of time since vaccination was 6 years (SD: ±0.4). The HPV-positive rate amongst the analyzed swabs was 69% (68/98). Among 30/98 (31%) HPV-positive participants, 13/98 (13%) had low-risk HPV types, and 17/98 (17%) had high-risk HPV types. Twelve participants tested positive for HPV18, only one participant tested positive for HPV16, and an additional four (4.3%) tested positive for either type 6 or 11, with respect to vaccine-preventable low-risk HPV types. CONCLUSION: The Gardasil quadrivalent HPV vaccine (4vHPV) was expected to protect against infection with HPV types 16, 18, 6, and 11. We demonstrated a possible waning of immunity to HPV18 in 17% of the participants, and an associated loss in cross-protection against HPV45. We observed a relatively high prevalence of 'opportunistic non-vaccine HPV types' or 'ecological niche occupiers' in this cohort, and suggest further research on the involvement of these types in cervical and other genital cancers. Our study is one of the few, if not the first, to report on HPV vaccine immunoprotection among people living with HIV (PLWH), thereby setting a baseline for further studies on HPV vaccine effectiveness among PLWH.

In vivo functional immunoprotection correlates for vaccines against invasive bacteria.

Vaccination has significantly reduced the incidence of invasive infections caused by several bacterial pathogens, including Streptococcus pneumoniae, Haemophilus influenzae, and Neisseria meningitidis. However, no vaccines are available for many other invasive pathogens. A major hurdle in vaccine development is the lack of functional markers to quantify vaccine immunity in eliminating pathogens during the process of infection. Based on our recent discovery of the liver as the major organ of vaccine-induced clearance of blood-borne virulent bacteria, we here describe a new vaccine evaluation system that quantitatively characterizes the key features of effective vaccines in shuffling virulent bacteria from the blood circulation to the liver resident macrophage Kupffer cells (KCs) and sinusoidal endothelial cells (LSECs) in mouse septic infection model. This system consists of three related correlates or assays: pathogen clearance from the bloodstream, pathogen trapping in the liver, and pathogen capture by KCs/LSECs. These readouts were consistently associated with the serotype-specific immunoprotection levels of the 13-valent pneumococcal polysaccharide conjugate vaccine (PCV13) against lethal infection of S. pneumoniae, a major invasive Gram-positive pathogen of community-acquired infections in humans. Furthermore, the reliability and sensitivity of these correlates in reflecting vaccine efficacy were verified with whole cell vaccines of Klebsiella pneumoniae and Escherichia coli, two major Gram-negative pathogens in hospital-acquired invasive infections. This system may be used as effective readouts to evaluate the immunoprotective potential of vaccine candidates in the preclinical phase by filling the current technical gap in vaccine evaluation between the conventional in vitro approaches (e.g. antibody production and pathogen neutralization/opsonophagocytosis) and survival of immunized animals.

Evaluation of the immunoprotective effects of porcine deltacoronavirus subunit vaccines.

Porcine deltacoronavirus (PDCoV), a new porcine enteric coronavirus, has seriously endangered the pig breeding industry and caused great economic losses. However, a PDCoV vaccine is not commercially available. Therefore, new and efficient PDCoV vaccines must be developed without delay. In this study, we used the ExpiCHO eukaryotic expression system to express and purify the following 3 structural proteins of PDCoV: S, N and M. Subsequently, the level of humoral and cellular immunity induced by the S protein (immunization with the S protein alone) and a protein mixture (immunization with a mixture of S, N and M proteins) were evaluated in mice and piglets, respectively, and the performances of the 2 immunizations in a challenge protection test were assessed in piglets. The results showed that both the S protein and the protein mixture induced the production of high levels of specific IgG antibodies and neutralizing antibodies and effectively neutralized PDCoV-infected LLC-PK cells in vitro. Furthermore, compared with the S protein, the N and M proteins in the protein mixture promoted the expression of CD8+ T cells and IFN-γ, induced a stronger cellular immune response, and effectively protected 4/5 of the piglets from PDCoV infection. In conclusion, the results of this study showed that the N and M proteins play important roles in inducing an immunoprotective response. Using N and M antigens as effective antigenic components in the development of PDCoV vaccines in the future will effectively increase the immune efficacy of the vaccines.

A novel chimeric recombinant FliC-Pgp3 vaccine promotes immunoprotection against Chlamydia muridarum infection in mice.

The Pgp3 subunit vaccine elicits immune protection against Chlamydia trachomatis infection, but additional adjuvants are still required to enhance its immunoprotective efficacy. Flagellin can selectively stimulate immunity and act as an adjuvant. In this research, the FliC-Pgp3 recombinant was successfully expressed and purified. Tri-immunization with the FliC-Pgp3 vaccine in Balb/C mice induced rapid and persistent germinal center B-cell response and Tfh differentiation, promoting a significantly higher IgG antibody titer compared to the Pgp3 group. FliC-Pgp3 immunization primarily induced Th1-type cellular immunity, leading to higher levels of IFN-γ, TNF-α, and IL-2 secreted by CD4+ T cells than in Pgp3-vaccinated mice. Chlamydia muridarum challenge results showed that FliC-Pgp3-vaccinated mice exhibited more rapid clearance of Chlamydia muridarum colonization in the lower genital tract, ensuring a lower hydrosalpinx rate and cumulative score. Histological analysis showed reduced dilation and inflammatory infiltration in the oviduct and uterine horn of FliC-Pgp3-vaccinated mice compared to the PBS and Pgp3 control. Importantly, tri-immunization with FliC-Pgp3 effectively activated CD4+ T cells and dendritic cells, as confirmed by the adoptive transfer, resulting in better immune protection in recipient mice. In summary, the novel FliC-Pgp3 chimeric is hoped to be a novel vaccine with improved immunoprotection against Chlamydia muridarum.

Expression of a SAG protein with a CAP domain from Eimeria necatrix and its role in invasion and immunoprotection.

Eimeria necatrix is a high pathogenic pathogen, which seriously endangers the poultry industry. The surface antigens (SAGs) of Apicomplexa are a kind of membrane protein anchored on the surface of the parasites through its carboxyl terminal glycosylphosphatidylinositol (GPI) structure. However, little is known about GPI-linked surface proteins in E. necatrix. In the present work, the E. necatrix sag gene (Ensag-CAP) was amplified and cloned for expression of the recombinant protein (rEnSAG-CAP). The full length Ensag-CAP gene was 813 bp, coding 270 amino acids with a predicated molecular weight of 28.86 kDa and contained a CAP domain with four sequence motifs CAP1, CAP2, CAP3 and CAP4. The rEnSAG-CAP was about 32 kDa and mainly expressed in a soluble form. Western blot analysis indicated that the rEnSAG-CAP could be recognized by anti-rEnSAG-CAP monoclonal antibody (anti-rEnSAG-CAP McAb) and the convalescent serum of chicken infected with E. necatrix. Native protein of EnSAG-CAP was detected in second-generation merozoites (MZ-2) using anti-rEnSAG-CAP polyclonal antibody (anti-rEnSAG-CAP pAb). The findings from the indirect immunofluorescence assay and enzyme digestion utilizing Bacillus cereus phosphoinositide-specific phospholipase C (PI-PLC) revealed that EnSAG-CAP predominantly localized at the surfaces of SZ and MZ-2 via a GPI anchor. It was observed that EnSAG-CAP can be cleaved from MZ-2 by PI-PLC. Real-time quantitative PCR (qPCR) analysis showed that transcript levels of Ensag-CAP in MZ-2 was significantly higher than that in SZ (P < 0.05). The anti-rEnSAG-CAP McAb in vitro could significantly inhibit the sporozoite invasion into MDBK cells (P < 0.01), which suggests that the protein might participate in sporozoite invasion into MDBK cells. rEnSAG-CAP afforded an immune protection against E. necatrix. The ACI value was 164.99 in the chickens immunized with 200 µg rEnSAG-CAP. Chickens immunized with rEnSAG-CAP had a significantly higher antigen-specific serum IgY response (P < 0.0001). The data indicates that EnSAG-CAP could serve as a potential candidate antigen for the development of a recombinant coccidiosis vaccine.

Vitamin D3 can effectively and rapidly clear largemouth bass ranavirus by immunoregulation.

Largemouth bass ranavirus (LMBV) is a highly destructive pathogen that causes significant mortality rates among largemouth bass populations. Unfortunately, there is a dearth of drug development efforts specifically aimed at treating LMBV. To address this, our study sought to investigate the potential effectiveness of incorporating varying doses of VD3 into the diet as a treatment for LMBV. Through qRT-PCR and semi-qPCR, we observed significant suppression and clearance of LMBV pathogens in largemouth bass fed with 15000 IU/Kg and 20000 IU/Kg of VD3 within 14 days. In addition, VD3 treatment significantly increased the expression levels of key immune-related genes such as IL-1ß, IFN-γ, Mx, and IgM. Encouragingly, we observed that VD3 significantly increased antioxidant and immune activities such as TSOD, TAOC and C3 in serum and maintained total protein levels. Additionally, tissue pathology sections highlighted a dose-dependent relationship between VD3 supplementation and tissue damage, with the 15000 IU and 20000 IU groups exhibiting minimal damage. In conclusion, a reasonable concentration of VD3 effectively reduced LMBV replication and tissue damages, while improved immune-related genes expression and serum biochemical indices. These findings declare the considerable therapeutic potential of VD3 supplementation for combating LMBV disease and provide an alternative treatment option for fish farming.

Probiotic Potential of Bacillus sp. 62A Isolated from a Marine Extreme Environment.

Antimicrobial resistance is an important health concern globally, and probiotics are considered an alternative to minimize it. The present study examined the in vitro probiotic characteristics and in vivo immunomodulatory potential of Bacillus sp. 62A - an extremophile bacterium. Bacillus sp. 62A was evaluated in vitro for its cytotoxicity, hemolytic activity, antibiotic susceptibility, and resistance to gastrointestinal conditions (bile salts, low pH, and intestinal adherence). Additionally, the immunomodulatory effect of Bacillus sp. 62A was studied in mice. The animals were supplemented daily with phosphate-buffered saline (control) and Bacillus sp. 62A at 1 × 108 colony forming units (CFU). Samples were taken on days 5 and 10. Isolated splenocytes were challenged with Escherichia coli for immunological analyses and immune-related gene expression. Serum and feces were collected for IgA and IgG determination. Bacillus sp. 62A did not show cytotoxicity, hemolytic activity, or resistance to antibiotics. Furthermore, the bacterium has autoaggregation and intestinal adhesion capacities and grows in the presence of bile salts and low pH. Bacillus supplementation in mice improved respiratory burst activity, nitric oxide production, and IL-1ß and IL-6 gene expressions, mainly at 10 days. After E. coli challenge, Bacillus supplementation in mice induced an anti-inflammatory response through a decrease in immunological parameters and an increase in IL-10 gene expression. Moreover, serum IgA and IgG and fecal IgG augmented in supplemented mice. In conclusion, Bacillus sp. 62A has biosafe and immunomodulatory probiotic potential.

Enhanced immunoprotection against Acinetobacter baumannii infection: Synergistic effects of Bap and BauA in a murine model.

BACKGROUND: The rise of multi-drug resistant Acinetobacter baumannii poses a grave threat to hospital settings, resulting in increased mortality rates and garnering global attention. The formation of biofilms facilitated by biofilm-associated protein (Bap) and the iron absorption capabilities mediated by Baumannii acinetobactin utilization A (BauA) contribute to the persistence and survival of multidrug-resistant strains. In this study, we aimed to investigate the potential of disrupting the function of BauA and Bap simultaneously as a strategy for controlling A. baumannii. METHODS: Recombinant Bap and BauA were expressed, purified, and subcutaneously administered individually and in combination to BALB/c mice. Subsequently, mice were intraperitoneally challenged with A. baumannii, and the bacterial load and tissue damage in the spleen, lung, and liver were assessed. Serum samples were evaluated to determine antibody titers in surviving mice. RESULTS: Specific IgG antibodies were significantly increased. A combination of the antigens resulted in enhanced titer of specific IgGs in comparison to either BauA or Bap alone. The antibodies remained stable over a seven-month period. The combination of Bap and BauA exhibited superior immunoprotection against A. baumannii infection compared to individual administration, resulting in a further reduction in bacterial load in the liver, spleen, and lungs. The histopathological analysis demonstrated successful protection of the tissues against A. baumannii-induced damage upon administration of the two immunogens. CONCLUSIONS: The combination of Bap and BauA has the potential to target a broader range of A. baumannii strains, including those expressing either Bap or BauA, thereby increasing its efficacy against a diverse array of strains.

MAIT cells in bacterial infectious diseases: heroes, villains, or both?

Due to the aggravation of bacterial drug resistance and the lag in the development of new antibiotics, it is crucial to develop novel therapeutic regimens for bacterial infectious diseases. Currently, immunotherapy is a promising regimen for the treatment of infectious diseases. Mucosal-associated invariant T (MAIT) cells, a subpopulation of innate-like T cells, are abundant in humans and can mount a rapid immune response to pathogens, thus becoming a potential target of immunotherapy for infectious diseases. At the site of infection, activated MAIT cells perform complex biological functions by secreting a variety of cytokines and cytotoxic substances. Many studies have shown that MAIT cells have immunoprotective effects because they can bridge innate and adaptive immune responses, leading to bacterial clearance, tissue repair, and homeostasis maintenance. MAIT cells also participate in cytokine storm generation, tissue fibrosis, and cancer progression, indicating that they play a role in immunopathology. In this article, we review recent studies of MAIT cells, discuss their dual roles in bacterial infectious diseases and provide some promising MAIT cell-targeting strategies for the treatment of bacterial infectious diseases.

Long-term immunoprotection after live attenuated measles-mumps-rubella booster vaccination in children with juvenile idiopathic arthritis.

INTRODUCTION: Vaccines, especially live attenuated vaccines, in children with JIA pose a great challenge due to both potential lower immunogenicity and safety as a result of immunosuppressive treatment. For many years, in the Netherlands, JIA patients receive a measles-mumps-rubella (MMR) booster vaccine at the age of nine years as part of the national immunization program. OBJECTIVES: To study long-term humoral immunoprotection in a large cohort of JIA patients who received the MMR booster vaccine while being treated with immunomodulatory therapies at the Wilhelmina Children's Hospital in Utrecht, the Netherlands. METHODS: MMR-specific IgG antibody concentrations in stored serum samples of vaccinated JIA patients were determined with chemiluminescent microparticle immunoassays (CMIA). Samples were analyzed five years after MMR booster vaccination and at last available follow-up visit using both crude and adjusted analyses. Additional clinical data were collected from electronic medical records. RESULTS: In total, 236 samples from 182 patients were analyzed, including 67 samples that were available five years post-vaccination, and an additional 169 samples available from last visits with a median duration after vaccination of 6.9 years (IQR: 2.8-8.8). Twenty-eight patients were using biologic disease-modifying antirheumatic drugs (bDMARDS) of whom 96% anti-TNF agents and 4% tocilizumab. Percentages of protective antibody levels against measles after five years were significantly lower for patients who used bDMARD therapy at vaccination compared to patients who did not: 60% versus 86% (P = 0.03). For mumps (80% versus 94%) and rubella (60% versus 83%) this difference did not reach statistical significance (P = 0.11 and P = 0.07, respectively). Antibody levels post-vaccination decreased over time, albeit not significantly different between bDMARD users and non-bDMARD users. CONCLUSION: The MMR booster vaccine demonstrated long-term immunogenicity in the majority of children with JIA from a large cohort, although lower percentages of protective measles antibody levels were observed in bDMARD users. Hence, it might be indicated to measure antibody levels at least five years after MMR booster vaccination in the latter group and advice an extra booster accordingly.

A novel live attenuated duck Tembusu virus vaccine targeting N7 methyltransferase protects ducklings against pathogenic strains.

Duck Tembusu virus (DTMUV), an emerging pathogenic flavivirus, causes markedly decreased egg production in laying duck and neurological dysfunction and death in ducklings. Vaccination is currently the most effective means for prevention and control of DTMUV. In previous study, we have found that methyltransferase (MTase) defective DTMUV is attenuated and induces a higher innate immunity. However, it is not clear whether MTase-deficient DTMUV can be used as a live attenuated vaccine (LAV). In this study, we investigated the immunogenicity and immunoprotection of N7-MTase defective recombinant DTMUV K61A, K182A and E218A in ducklings. These three mutants were highly attenuated in both virulence and proliferation in ducklings but still immunogenic. Furthermore, a single-dose immunization with K61A, K182A or E218A could induce robust T cell responses and humoral immune responses, which could protect ducks from the challenge of a lethal-dose of DTMUV-CQW1. Together, this study provides an ideal strategy to design LAVs for DTMUV by targeting N7-MTase without changing the antigen composition. This attenuated strategy targeting N7-MTase may apply to other flaviviruses.

Preparation of graphene oxide-stabilized Pickering emulsion adjuvant for Pgp3 recombinant vaccine and enhanced immunoprotection against Chlamydia Trachomatis infection.

Background: Traditional emulsion adjuvants are limited in clinical application because of their surfactant dependence. Graphene oxide (GO) has unique amphiphilic properties and therefore has potential to be used as a surfactant substitute to stabilize Pickering emulsions. Methods: In this study, GO-stabilized Pickering emulsion (GPE) was prepared and used as an adjuvant to facilitate an enhanced immune response to the Chlamydia trachomatis (Ct) Pgp3 recombinant vaccine. Firstly, GPE was prepared by optimizing the sonication conditions, pH, salinity, GO concentration, and water/oil ratio. GPE with small-size droplets was characterized and chosen as the candidate. Subsequently, controlled-release antigen delivery by GPE was explored. Cellular uptake behaviors, M1 polarization, and cytokine stimulation by GPE + Pgp3 was considered in terms of the production of macrophages. Finally, GPE's adjuvant effect was evaluated by vaccination with Pgp3 recombinant in BALB/c mouse models. Results: GPE with the smallest droplet sizes was prepared by sonication under 163 W for 2 min at 1 mg/mL GO in natural salinity with a pH of 2 when the water/oil ratio was 10:1 (w/w). The optimized average GPE droplet size was 1.8 µm and the zeta potential was -25.0 ± 1.3 mv. GPE delivered antigens by adsorption onto the droplet surface, demonstrating the controlled release of antigens both in vitro and in vivo. In addition, GPE promoted antigen uptake, which stimulated proinflammatory tumor necrosis factor alpha (TNF-α), enhancing the M1 polarization of macrophages in vitro. Macrophage recruitment was also significantly promoted by GPE at the injection site. In the GPE + Pgp3 treatment group, higher levels of immunoglobin (IgG), immunoglobin G1 (IgG1), immunoglobin G2a (IgG2a) sera, and immunoglobin A (IgA) were detected in vaginal fluid, and higher levels of IFN-γ and IL-2 secretion were stimulated, than in the Pgp3 group, showing a significant type 1 T helper (Th1)-type cellular immune response. Chlamydia muridarum challenging showed that GPE enhanced Pgp3's immunoprotection through its advanced clearance of bacterial burden and alleviation of chronic pathological damage in the genital tract. Conclusion: This study enabled the rational design of small-size GPE, shedding light on antigen adsorption and control release, macrophage uptake, polarization and recruitment, which enhanced augmented humoral and cellular immunity and ameliorated chlamydial-induced tissue damage in the genital tract.

Antitumor Effect of Açaí (Euterpe oleracea Mart.) Seed Extract in LNCaP Cells and in the Solid Ehrlich Carcinoma Model.

Euterpe oleracea (açaí) fruit has approximately 15% pulp, which is partly edible and commercialized, and 85% seeds. Although açaí seeds are rich in catechins-polyphenolic compounds with antioxidant, anti-inflammatory, and antitumor effects-almost 935,000 tons/year of seeds are discarded as industrial waste. This work evaluated the antitumor properties of E. oleracea in vitro and in vivo in a solid Ehrlich tumor in mice. The seed extract presented 86.26 ± 0.189 mg of catechin/g of extract. The palm and pulp extracts did not exhibit in vitro antitumor activity, while the fruit and seed extracts showed cytotoxic effects on the LNCaP prostate cancer cell line, inducing mitochondrial and nuclear alterations. Oral treatments were performed daily at 100, 200, and 400 mg/kg of E. oleracea seed extract. The tumor development and histology were evaluated, along with immunological and toxicological parameters. Treatment at 400 mg/kg reduced the tumor size, nuclear pleomorphism, and mitosis figures, increasing tumor necrosis. Treated groups showed cellularity of lymphoid organs comparable to the untreated group, suggesting less infiltration in the lymph node and spleen and preservation of the bone marrow. The highest doses reduced IL-6 and induced IFN-γ, suggesting antitumor and immunomodulatory effects. Thus, açaí seeds can be an important source of compounds with antitumor and immunoprotective properties.

Evaluation of acellular pertussis vaccine: comparisons among different strains of mice.

The current study was designed to comparatively analyse the reactions of different mouse strains in response to acellular pertussis (aP) vaccine, with attempt to further provide a reference for aP vaccine evaluation. NIH mice, ICR mice, and BALB/c mice adopted from different pharmacopoeias and studies were utilized to measure the immune protection and immunogenicity of the same batch of aP vaccine according to the Modified intracerebral challenge assay (MICA) from some Asian pharmacopoeias and the pertussis serological potency test (PTST) method from European Pharmacopoeia. Based on our results, the aP vaccine detected by NIH mice had the best potency. So the NIH mice were more suitable for detecting the immune protection of aP vaccine by the MICA method. Given that the levels of PT-IgG and FHA-IgG antibodies in ICR mice were the highest, and the levels of Th1 and Th2 cells were significantly increased (P < .01), it was more suitable for the detection of immunogenicity of aP vaccine by PSPT method. Spleen lymphocytes were stimulated by PT and FHA. And the levels of IL-4 in ICR mice and NIH mice were significantly increased, so were the levels of IL-17, IL-23, IL-27, and TNF-α in BALB/c mice. NIH mice have stronger adaptive immunity and the weakest inflammatory response, and ICR mice have enhanced adaptive immunity and inflammatory responses, both of which can be thereby used for evaluation by different pharmacopoeia methods. NIH was more suitable for the MICA method of Chinese Pharmacopoeia, and ICR for the PSPT method of European Pharmacopoeia.

Immunization with Tp0954, an adhesin of Treponema pallidum, provides protective efficacy in the rabbit model of experimental syphilis.

Syphilis, a chronic multisystemic disease caused by spirochete Treponema pallidum subspecies pallidum infection, continues to be a serious global health problem and congenital syphilis remains a major cause of adverse outcomes in pregnancy in developing countries. The development of an effective vaccine is the most cost-effective way to eliminate syphilis, but so far has been elusive. Here, we evaluated the immunogenicity and protective efficacy of Tp0954, a T. pallidum placental adhesin, as a potential vaccine candidate in a New Zealand White rabbit model of experimental syphilis. Animals immunized with recombinant Tp0954 (rTp0954) produced high titers of Tp0954-specific serum IgG, high levels of IFN-γ from splenocytes and specific splenocyte proliferation response when compared to control animals immunized with PBS and Freund's adjuvant (FA). Furthermore, rTp0954 immunization significantly delayed the development of cutaneous lesions, promoted inflammatory cellular infiltration at the primary lesion sites, as well as inhibited T. pallidum dissemination to distal tissues or organs when compared with that of the control animals. In addition, the naïve rabbits receiving popliteal lymph nodes from Tp0954-immunized, T. pallidum-challenged animals were not infected by T. pallidum, confirming sterile immunity. These findings suggest that Tp0954 is a potential vaccine candidate against syphilis.

Emerging of a new CD3+CD31HCD184+ tang cell phenothype in Sjögren's syndrome induced by microencapsulated human umbilical cord matrix-derived multipotent stromal cells.

Background: Sjögren's syndrome (SS) is an autoimmune disease hallmarked by infiltration and destruction of exocrine glands. Currently, there is no therapy that warrants full recovery of the affected tissues. Umbilical cord-derived multipotent stromal cells, microincapsulated in an endotoxin-free alginate gel (CpS-hUCMS), were shown to modulate the inflammatory activity of PBMCs in SS patients in vitro, through release of soluble factors (TGFß1, IDO1, IL6, PGE2, VEGF). These observations led us to set up the present study, aimed at defining the in vitro effects of CpS-hUCMS on pro- and anti-inflammatory lymphocyte subsets involved in the pathogenesis of SS. Methods and results: Peripheral blood mononuclear cells (PBMCs) upon collection from SS patients and matched healthy donors, were placed in co-culture with CpS-hUCMS for five days. Cellular proliferation and T- (Tang, Treg) and B- (Breg, CD19+) lymphocyte subsets were studied by flow cytometry, while Multiplex, Real-Time PCR, and Western Blotting techniques were employed for the analysis of transcriptome and secretome. IFNγ pre-treated hUCMS were assessed with a viability assay and Western Blotting analysis before co-culture. After five days co-culture, CpS-hUCMS induced multiple effects on PBMCs, with special regard to decrease of lymphocyte proliferation, increase of regulatory B cells and induction of an angiogenic T cell population with high expression of the surface marker CD31, that had never been described before in the literature. Conclusion: We preliminarily showed that CpS-hUCMS can influence multiple pro- and anti-inflammatory pathways that are deranged in SS. In particular, Breg raised and a new Tang phenothype CD3+CD31HCD184+ emerged. These results may considerably expand our knowledge on multipotent stromal cell properties and may open new therapeutic avenues for the management of this disease, by designing ad hoc clinical studies.

Immunogenicity and protective efficacy of OmpA subunit vaccine against Aeromonas hydrophila infection in Megalobrama amblycephala: An effective alternative to the inactivated vaccine.

Aeromonas hydrophila is a kind of zoonotic pathogen, which can cause bacterial septicemia in fish and bring huge economic losses to global aquaculture. Outer membrane proteins (Omps) are conserved antigens of Aeromonas hydrophila, which can be developed as subunit vaccines. To evaluate the protective efficacy of inactivated vaccine and recombinant outer membrane protein A (OmpA) subunit vaccine against A. hydrophila in juvenile Megalobrama amblycephala, the present study investigated the immunogenicity and protective effects of both vaccines, as well as the non-specific and specific immune response of M. amblycephala. Compared with the non-vaccinated group, both inactivated and OmpA subunit vaccines improved the survival rate of M. amblycephala upon infection. The protective effects of OmpA vaccine groups were better than that of the inactivated vaccine groups, which should be attributed to the reduced bacterial load and enhanced host immunity in the vaccinated fish. ELISA assay showed that the titer of serum immunoglobulin M (IgM) specific to A. hydrophila up-regulated significantly in the OmpA subunit vaccine groups at 14 d post infection (dpi), which should contribute to better immune protective effects. In addition, vaccination enhanced host bactericidal abilities might also attribute to the regulation of the activities of hepatic and serum antimicrobial enzymes. Moreover, the expression of immune-related genes (SAA, iNOS, IL-1 ß, IL-6, IL-10, TNF α, C3, MHC I, MHC II, CD4, CD8, TCR α, IgM, IgD and IgZ) increased in all groups post infection, which was more significant in the vaccinated groups. Furthermore, the number of immunopositive cells exhibiting different epitopes (CD8, IgM, IgD and IgZ) that were detected by immunohistochemical assay had increased in the vaccinated groups post infection. These results show that vaccination effectively stimulated host immune response (especially OmpA vaccine groups). In conclusion, these results indicated that both the inactivated vaccine and OmpA subunit vaccine could protect juvenile M. amblycephala against A. hydrophila infection, of which OmpA subunit vaccine provided more effective immune protection and can be used as an ideal candidate for the A. hydrophila vaccine.

The Immunoprotection of OmpH Gene Deletion Mutation of Pasteurella multocida on Hemorrhagic Sepsis in Qinghai Yak.

OmpH is among the most important virulence factors of Pasteurella multocida, which mediates septicemia in yaks (Bos grunniens I) after infection with the bacteria. In the present study, yaks were infected with wild-type (WT) (P0910) and OmpH-deficient (ΔOmpH) P. multocida strains. The mutant strain was generated through the reverse genetic operation system of pathogens and proteomics technology. The live-cell bacterial count and clinical manifestations of P. multocida infection in Qinghai yak tissues (thymus, lung, spleen, lymph node, liver, kidney, and heart) were analyzed. The expression of differential proteins in the yak spleen under different treatments was analyzed using the marker-free method. We found that compared with the mutant strain, the titer of wild-type strains was significantly higher in tissues. Additionally, compared with other organs, the bacteria titer was significantly higher in the spleen. Compared with the WT p0910 strain, the mutant strain generated milder pathological changes in the tissues of yak. Proteomics analysis revealed that 57 of the 773 proteins expressed in P. multocida were significantly differentially expressed between the ΔOmpH and P0910 groups. Of the 57, 14 were over-expressed, whereas 43 were under-expressed. The differentially expressed proteins in the ΔompH group regulated the ABC transporter (ATP-powered translocation of many substrates across membranes) system, the two-component system, RNA degradation, RNA transcription, glycolysis/gluconeogenesis, biosynthesis of ubiquinone and other terpenoid-quinones, oxidative phosphorylation (citrate cycle) as well as fructose and mannose metabolism. The relationship among 54 significantly regulated proteins was analyzed using STRING. We found that WT P0910 and ΔOmpH of P. multocida infection activated the expression of ropE, HSPBP1, FERH, ATP10A, ABCA13, RRP7A, IL-10, IFN-γ, IL-17A, EGFR, and dnaJ. Overall, deletion of the OmpH gene weakened the virulence but maintained the immunogenicity of P. multocida in yak. The findings of this study provide a strong foundation for the pathogenesis of P. multocida and the management of related septicemia in yaks.