Immune response induced by a Streptococcus suis multi-serotype autogenous vaccine used in sows to protect post-weaned piglets.

Streptococcus suis is a bacterial pathogen that causes important economic losses to the swine industry worldwide. Since there are no current commercial vaccines, the use of autogenous vaccines applied to gilts/sows to enhance transfer of passive immunity is an attractive alternative to protect weaned piglets. However, there is no universal standardization in the production of autogenous vaccines and the vaccine formulation may be highly different among licenced manufacturing laboratories. In the present study, an autogenous vaccine that included S. suis serotypes 2, 1/2, 5, 7 and 14 was prepared by a licensed laboratory and administrated to gilts using a three-dose program prior to farrowing. The antibody response in gilts as well as the passive transfer of antibodies to piglets was then evaluated. In divergence with previously published data with an autogenous vaccine produced by a different company, the increased response seen in gilts was sufficient to improve maternal antibody transfer to piglets up to 5 weeks of age. However, piglets would still remain susceptible to S. suis disease which often appears during the second part of the nursery period. Vaccination did not affect the shedding of S. suis (as well as that of the specific S. suis serotypes included in the vaccine) by either gilts or piglets. Although all antibiotic treatments were absent during the trial, the clinical protective effect of the vaccination program with the autogenous vaccine could not be evaluated, since limited S. suis cases were present during the trial, confirming the need for a complete evaluation of the clinical protection that must include laboratory confirmation of the aetiological agent involved in the presence of S. suis-associated clinical signs. Further studies to evaluate the usefulness of gilt/sow vaccination with autogenous vaccines to protect nursery piglets should be done.

Development of novel Streptococcus equi vaccines with an assessment of their immunizing potentials and protective efficacies.

Strangles is a highly contagious disease of the equine upper respiratory tract caused by Streptococcus equi subspecies. Streptococcus equi subsp. equi (S. equi) and Streptococcus equi subsp. zooepidemicus (S. zooepidemicus) was isolated, as local, hot, and field strains, from horses clinically suffering from respiratory distress. The isolated Streptococci were identified using bacteriological and molecular techniques. Four formulations of inactivated S. equi vaccines were developed and evaluated. The first formulation was prepared using the S. equi isolates, adjuvanted with MONTANIDE GEL adjuvant, while the second formulation was adjuvanted with MONTANIDE ISA-70 adjuvant. The other 2 formulations were inactivated combined vaccines prepared from both S. equi and S. zooepidemicus isolates. The 3rd formulation was the combined isolates adjuvanted with MONTANIDE GEL while the 4th formulation was the combined isolates adjuvanted with MONTANIDE ISA-70. The developed vaccines' physical properties, purity, sterility, safety, and potency were ensured. The immunizing efficacy was determined in isogenic BALB/c mice and white New Zealand rabbits using the passive hemagglutination test. Also, the antibodies' titer of the combined S. equi and S. zooepidemicus vaccine adjuvanted with MONTANIDE ISA-70 in foals was tracked using an indirect enzyme-linked immunosorbent assay. The protective efficacy of the developed vaccines was determined using a challenge test in both laboratory and field animal models, where a 75% protection rate was achieved. The combined vaccine proved to be more efficacious than the monovalent vaccine. Also, the MONTANIDE ISA-70 adjuvant provided significant protective efficacy than the MONTANIDE GEL. The current work is introducing a very promising mitigative and strategic controlling solution for strangles.

An optimized caries model of Streptococcus mutans in rats and its application for evaluating prophylactic vaccines.

Dental caries is a prevalent oral disease that mainly results from Streptococcus mutans. Susceptibility to S. mutans decreased rapidly after weaning in a well-known rat model. However, owing to the lack of time to establish protective immunity ahead of challenge, the weaning rat model is suboptimal for assessing prophylactic vaccines against S. mutans infection. In this study, we found that, in adult rats, S. mutans cultured under air-restricted conditions showed dramatically increased colonization efficacy and accelerated development of dental caries compared with those cultured under air-unrestricted conditions. We propose that S. mutans cultured under air-restricted conditions can be used to develop an optimal caries model, especially for the evaluation of prophylactic efficacy against S. mutans. Therefore, we used the anti-caries vaccine, KFD2-rPAc, to reevaluate the protection against the challenge of S. mutans. In immunized rats, rPAc-specific protective antibodies were robustly elicited by KFD2-rPAc before the challenge. In addition to inhibiting the initial and long-term colonization of S. mutans in vivo, KFD2-rPAc immunization showed an 83% inhibitory efficacy against the development of caries, similar to that previously evaluated in a weaning rat model. These results demonstrate that culturing under air-restricted conditions can promote S. mutans infection in adult rats, thereby helping establish a rat infection model to evaluate the prophylactic efficacy of vaccines and anti-caries drugs.

Recombinant lactococcal-based oral vaccine for protection against Streptococcus agalactiae infections in tilapia (Oreochromis niloticus).

Streptococcosis outbreaks caused by Streptococcus agalactiae infection in tilapia aquaculture have been consistently reported and associated with high mortality and morbidity leading to significant economic losses. Existing vaccine candidates against Streptococcus spp. are designed for intraperitoneal injections that are not practical and labor-intensive which have prompted farmers to protect aquatic animals with antibiotics, thus encouraging the emergence of multidrug resistant bacteria. In this study, a live recombinant L. lactis vaccine expressing a 1403 bp surface immunogenic protein (SIP) and a 1100 bp truncated SIP (tSIP) gene was developed and evaluated against S. agalactiae infection in tilapia. Both SIP and tSIP sequences were cloned and transformed into L. lactis. The recombinant L.lactis vaccine was orally administered to juvenile tilapia for a month. Detection of SIP-specific serum IgM in vaccinated groups compared to control groups indicated that recombinant proteins expressed from L. lactis could elicit immunogenic reactions in tilapia. Fish immunized with the tSIP vaccine also showed the highest level of protection compared to other test groups, and the mortality rate was significantly reduced compared to both control groups. The relative percentage of survival (RPS) against S. agalactiae for both SIP and tSIP-vaccinated groups was 50 % and 89 %, respectively, at 14 days post-challenge. Significant up-regulation of IgM, IL-1ß, IL-10, TNF-α and IFN-γ were observed at day 34 between the vaccinated and control groups. These results indicated that the recombinant lactococcal tSIP vaccine can elicit both cell-mediated and humoral responses and is recommended as a potential oral vaccine against S. agalactiae infection. Future work will include further in vivo challenge assessments of this vaccine candidate fused with adjuvants to boost immunogenicity levels in tilapia.

Evaluation of the efficacy of MONTANIDE™ GR01, a new adjuvant for feed-based vaccines, on the immune response and protection against Streptococcus agalactiae in oral vaccinated Nile tilapia (Oreochromis niloticus) under laboratory and on-farm conditions.

Streptococcosis, an emerging infectious disease caused by Streptococcus agalactiae, has had adverse effects on farmed tilapia. Several vaccines have been developed to prevent this disease and induce a specific immune response against S. agalactiae infection. In this study the use of MONTANIDE™ GR01, a new adjuvant for oral vaccination, was optimized for use in tilapia under laboratory and field studies. In the laboratory trial the immune response and protective efficacy of two doses of MONTANIDE™ GR01, 20 % (w/w) and 2 % (w/w), included into the feed-based adjuvanted vaccines were assessed comparatively. Following immunization, the innate immune parameters studied in serum, including lysozyme, myeloperoxidase, catalase and glutathione peroxidase activity, were all increased significantly. Furthermore, specific IgM antibodies against S. agalactiae were induced significantly in serum post-vaccination, with higher levels observed in both groups that received the feed-based adjuvanted vaccine. Under both injection and immersion challenge conditions, the relative percent survival for the feed-based adjuvanted vaccine groups ranged from 78 % to 84 %. Following use of the low dose concentration of MONTANIDE™ GR01 for oral vaccination of tilapia in cage culture systems, several innate immune parameters were effectively enhanced in the immunized fish. Similarly, the levels of specific IgM antibodies in the serum of feed-based vaccinated fish were significantly enhanced, reaching their highest levels 2-5 months post-vaccination. Cytokines associated with innate and adaptive immunity were also examined, and the expression levels of several genes showed significant up-regulation. This indicates that both cellular and humoral immune responses were induced by the feed-based adjuvanted vaccine. The economic impact of a feed-based adjuvanted vaccine was examined following vaccination, considering the growth performance and feed utilization of the fish. It was found that the Economic Performance Index and Economic Conversion Ratio were unaffected by vaccination, further demonstrating that there are no negative impacts associated with administering a feed-based vaccine to fish. In conclusion, the data from this study indicate that MONTANIDE™ GR01 is a highly valuable adjuvant for oral vaccination, as demonstrated by its ability to induce a strong immune response and effectively prevent streptococcal disease in Nile tilapia.

Application of Transthoracic Echocardiography for Cardiac Safety Evaluation in the Clinical Development Process of Vaccines Against Streptococcus pyogenes.

Superficial infections with Streptococcus pyogenes (Strep A), pharyngitis and impetigo can induce acute rheumatic fever, an autoimmune sequela manifesting mostly with arthritis and rheumatic carditis. Valvular heart damage can persist or advance following repeated episodes of acute rheumatic fever, causing rheumatic heart disease. Acute rheumatic fever and rheumatic heart disease disproportionately affect children and young adults in developing countries and disadvantaged communities in developed countries. People living with rheumatic heart disease are at risk of experiencing potentially fatal complications such as heart failure, bacterial endocarditis or stroke. Transthoracic echocardiography plays a central role in diagnosing both rheumatic carditis and rheumatic heart disease. Despite the obvious medical need, no licensed Strep A vaccines are currently available, as their clinical development process faces several challenges, including concerns for cardiac safety. However, the development of Strep A vaccines has been recently relaunched by many vaccine developers. In this context, a reliable and consistent safety evaluation of Strep A vaccine candidates, including the use of transthoracic echocardiography for detecting cardiac adverse events, could greatly contribute to developing a safe and efficacious product in the near future. Here, we propose a framework for the consistent use of transthoracic echocardiography to proactively detect cardiac safety events in clinical trials of Strep A vaccine candidates.

Throat and skin infections caused by certain types of bacteria, named Streptococcus pyogenes, are frequent worldwide; however, in many children from less developed countries and disadvantaged communities, infections with S. pyogenes lead to a condition called acute rheumatic fever, which usually affects the joints and the heart. Damage to the heart valves may evolve to rheumatic heart disease, a permanent condition with often life-threatening complications. Rheumatic heart disease is an important health problem in places and communities where S. pyogenes infections occur frequently. A vaccine against these bacteria would help lower the number of people with valvular heart disease; however, no such vaccine exists yet. Research on vaccines against S. pyogenes was on hold for almost 30 years because of initial concerns that vaccinated children might develop acute rheumatic fever more frequently. Recently, researchers started working again on vaccines against S. pyogenes, but concerns about the safety of such vaccines persist. Doctors can reliably use echocardiography to diagnose cases of rheumatic carditis (as a sign of acute rheumatic fever) and rheumatic heart disease. Here, we propose a simple approach for the consistent use of echocardiography in clinical research of vaccines against S. pyogenes that will allow the detection of any potential heart-related side effects of the vaccine.

Potential for Maternally Administered Vaccine for Infant Group B Streptococcus.

BACKGROUND: Natural history studies have correlated serotype-specific anti-capsular polysaccharide (CPS) IgG in newborns with a reduced risk of group B streptococcal disease. A hexavalent CPS-cross-reactive material 197 glycoconjugate vaccine (GBS6) is being developed as a maternal vaccine to prevent invasive group B streptococcus in young infants. METHODS: In an ongoing phase 2, placebo-controlled trial involving pregnant women, we assessed the safety and immunogenicity of a single dose of various GBS6 formulations and analyzed maternally transferred anti-CPS antibodies. In a parallel seroepidemiologic study that was conducted in the same population, we assessed serotype-specific anti-CPS IgG concentrations that were associated with a reduced risk of invasive disease among newborns through 89 days of age to define putative protective thresholds. RESULTS: Naturally acquired anti-CPS IgG concentrations were associated with a reduced risk of disease among infants in the seroepidemiologic study. IgG thresholds that were determined to be associated with 75 to 95% reductions in the risk of disease were 0.184 to 0.827 µg per milliliter. No GBS6-associated safety signals were observed among the mothers or infants. The incidence of adverse events and of serious adverse events were similar across the trial groups for both mothers and infants; more local reactions were observed in the groups that received GBS6 containing aluminum phosphate. Among the infants, the most common serious adverse events were minor congenital anomalies (umbilical hernia and congenital dermal melanocytosis). GBS6 induced maternal antibody responses to all serotypes, with maternal-to-infant antibody ratios of approximately 0.4 to 1.3, depending on the dose. The percentage of infants with anti-CPS IgG concentrations above 0.184 µg per milliliter varied according to serotype and formulation, with 57 to 97% of the infants having a seroresponse to the most immunogenic formulation. CONCLUSIONS: GBS6 elicited anti-CPS antibodies against group B streptococcus in pregnant women that were transferred to infants at levels associated with a reduced risk of invasive group B streptococcal disease. (Funded by Pfizer and the Bill and Melinda Gates Foundation; C1091002 ClinicalTrials.gov number, NCT03765073.).

ß-glucan mimics tissue damage signaling and generates a trade-off between head kidney and spleen to activate acquired immunity in vaccinated tilapia (Oreochromis niloticus).

The association of vaccines with immunostimulants such as ß-glucan, promote the production of cytokines, competent immune cells and antibodies. However, differences between ß-glucan types and trials make it difficult to understand ß-glucan's mechanism of action. In this study, three trials were carried out with control and fish fed ß-glucan, the first trial occurred at 15 days; the second trial occurred at 30 days when we associated ß-glucan and vaccine; and the third trial occurred at 15 days post-challenge with Streptococcus agalactiae in tilapia (O. niloticus) in order to investigate immune-related gene expression in the head kidney and spleen using real-time qPCR. We found increases in HSP70, IL-6, IL-1ß, TNF-α, IL-10, Lys and C3 predominantly in the head kidney, except for IgM expression, which prevailed in the spleen, under vaccinated + ß-glucan action. This demonstrates the trade-off presented by the head kidney and spleen after immunostimulation in order to produce acquired immunity, as well as an increase in HSP70 expression in vaccinated + ß-glucan fish. The results suggest that ß-glucan stimulates the immune response through damage-associated molecular patterns (DAMPs) recognition. Therefore, these dynamics of the immune response promote a more robust defense against disease.

Immunogenicity and protective efficacy of a Streptococcus suis vaccine composed of six conserved immunogens.

A vaccine protecting against different Streptococcus suis serotypes is highly needed in porcine practice to improve animal welfare and reduce the use of antibiotics. We hypothesized that immunogens prominently recognized by convalescence sera but significantly less so by sera of susceptible piglets are putative protective antigens. Accordingly, we investigated immunogenicity and protective efficacy of a multicomponent vaccine including six main conserved immunogens, namely SSU0934, SSU1869, SSU0757, SSU1950, SSU1664 and SSU0187. Flow cytometry confirmed surface expression of all six immunogens in S. suis serotypes 2, 9 and 14. Although prime-booster vaccination after weaning resulted in significantly higher specific IgG levels against all six immunogens compared to the placebo-treated group, no significant differences between bacterial survival in blood from either vaccinated or control animals were recorded for serotype 2, 9 and 14 strains. Furthermore, vaccinated piglets were not protected against morbidity elicited through intranasal challenge with S. suis serotype 14. As ~50% of animals in both groups did not develop disease, we investigated putative other correlates of protection. Induction of reactive oxygen species (ROS) in blood granulocytes was not associated with vaccination but correlated with protection as all piglets with >5% ROS survived the challenge. Based on these findings we discuss that the main immunogens of S. suis might actually not be a priori good candidates for protective antigens. On the contrary, expression of immunogens that evoke antibodies that do not mediate killing of this pathogen might constitute an evolutionary advantage conserved in many different S. suis strains.

Streptococcus equi-derived extracellular vesicles as a vaccine candidate against Streptococcus equi infection.

Streptococcus equi subspecies equi is a pathogenic bacterium that causes strangles, a highly contagious respiratory infection in horses and other equines. The limitations of current vaccines against S. equi infection warrants the development of an affordable, safe, and effective vaccine. Because gram-positive extracellular vesicles (EVs) transport various immunogenic antigens, they are attractive vaccine candidates. Here, we purified the EVs of S. equi ATCC 39506 and evaluated them as a vaccine candidate against S. equi infection in mice. As an initial step, comparative proteomic analysis was performed to characterize the functional features of the EVs. Reverse vaccinology and knowledge-based annotations were then used to screen potential vaccine candidates (PVCs) for S. equi ATCC 39506. Finally, 32 PVCs were found to be enriched in the EV fraction, suggesting the usefulness of this fraction as a vaccine. Importantly, a significantly higher survival rate after S. equi infection was detected in mice immunized with S. equi-derived EVs via the intraperitoneal route than in mice immunized with heat-killed bacteria. Of note, immunoprecipitation-mass spectrometry results validated various immunogenic antigens within the EV proteome. In conclusion, our results suggest that S. equi-derived EVs can serve as a vaccine candidate against S. equi infection.

Immune response and protective efficacy of two new adjuvants, Montanide™ ISA 763B VG and Montanide™ GEL02, administered with a Streptococcus agalactiae ghost vaccine in Nile tilapia (Oreochromis niloticus).

Streptococcus agalactiae is one of the most important pathogens infecting tilapia worldwide and causes meningoencephalitis, septicemia and high mortalities with considerable losses. Various types of vaccines have been developed against S. agalactiae infection, such as inactivated vaccines, live attenuated vaccines and subunit vaccines. Bacterial ghosts (BGs) are nonliving, empty cell envelopes and have been reported as novel vaccine candidates. Therefore, the main aims of this study were to develop an S. agalactiae ghost vaccine (SAGV) and to evaluate the immune response and protective effect of SAGV against S. agalactiae with two novel adjuvants, Montanide™ ISA 763B VG and Montanide™ GEL02. Nile tilapia, mean weight 50 g, were divided into four groups as follows; 1) fish injected with PBS as control, 2) fish injected with the SAGV alone; 3) fish injected with the SAGV+Montanide™ ISA 763B VG; and 4) fish injected with SAGV+Montanide™ GEL02. Following vaccination, innate immunity parameters including serum lysozyme, myeloperoxidase, catalase, and bactericidal activity were all significantly enhanced. Moreover, specific serum IgM antibodies were induced and reached their highest level 2-8 weeks post vaccination. Importantly, the relative percent survival of tilapia vaccinated against the SAGV formulated with both adjuvants was 80-93%. Furthermore, the transcription of immune-related genes (IgM, TCRß, IL-1ß, IL-8 and TNFα) were up-regulated in tilapia after vaccination, indicating that both cellular and humoral immune responses were induced by these adjuvanted vaccines. In summary, Montanide™ ISA 763B VG and Montanide™ GEL02 can enhance immunoprotection induced by the SAGV vaccine against streptococcosis, demonstrating that both have value as potential adjuvants of fish vaccines.

Immunogenicity in Oreochromis niloticus vaccinated with sonicated antigens against streptococcosis.

Streptococcosis causes great economic losses in intensive culture of tilapia. Vaccination is the most effective and safest way to tackle infectious diseases. Thus, this study sought the more effective and safer antigenic fraction after sonication of Streptococcus agalactiae to elaborate a vaccine against streptococcosis in Nile tilapia. For this, twenty-one days after vaccination with different fractions (soluble and insoluble) of S. agalactiae, the fish were challenged with the homologous strain (LD50). Then, samples were taken at zero, 14, 28, 60 and 90 days post-vaccination (DPV, n = 7). Blood and organs (cranial kidney, spleen and liver) were collected from vaccinated and unvaccinated fish. Finally, insoluble fraction vaccine presented the best effect, resulting in a 100% relative percent of survival (RPS) and without clinical manifestations. In view of the results, it was to evaluate the role of the insoluble fraction of the antigen in the protective immunity against streptococcosis. The results indicate that the spleen might be the main organ in the vaccine response in Nile tilapia due to the great morphological and immunological differences in vaccinated fish, evidenced by the greater of melanomacrophage centers (MMC) and IgM + lymphocytes in relation to the non-vaccinated fish. At 60 DPV, it was observed the peak of the protective immunity related to the maximum concentration of proteins, circulating leukocytes, antibody titers in the serum and tissue changes with greater expression of IgM + and MMC number in the spleen and kidney of Oreochromis niloticus. Vaccination with insoluble fraction of S. agalactiae was safe and provided effective protection against streptococcosis with maximum protective response at 60 DPV.

Prime-Pull Immunization with a Bivalent M-Protein and Spy-CEP Peptide Vaccine Adjuvanted with CAF®01 Liposomes Induces Both Mucosal and Peripheral Protection from covR/S Mutant Streptococcus pyogenes.

Infections with Streptococcus pyogenes and their sequelae are responsible for an estimated 18 million cases of serious disease with >700 million new primary cases and 500,000 deaths per year. Despite the burden of disease, there is currently no vaccine available for this organism. Here, we define a combination vaccine P*17/K4S2 comprising of 20-mer B-cell peptide epitopes, p*17 (a mutant derived from the highly conserved C3-repeat region of the M-protein), and K4S2 (derived from the streptococcal anti-neutrophil factor, Spy-CEP). The peptides are chemically conjugated to either diphtheria toxoid (DT) or a nontoxic mutant form of diphtheria toxin, CRM197. We demonstrate that a prime-pull immunization regimen involving two intramuscular inoculations with P*17/K4S2 adjuvanted with a two-component liposomal adjuvant system (CAF01; developed by Statens Serum Institut [SSI], Denmark), followed by an intranasal inoculation of unadjuvanted vaccine (in Tris) induces peptide- and S. pyogenes-binding antibodies and protects from mucosal and skin infection with hypervirulent covR/S mutant organisms. Prior vaccination with DT does not diminish the response to the conjugate peptide vaccines. Detailed Good Laboratory Practice (GLP) toxicological evaluation in male and female rats did not reveal any gross or histopathological adverse effects.IMPORTANCE A vaccine to control S. pyogenes infection is desperately warranted. S. pyogenes colonizes the upper respiratory tract (URT) and skin, from where it can progress to invasive and immune-mediated diseases. Global mortality estimates for S. pyogenes-associated diseases exceeds 500,000 deaths per year. S. pyogenes utilizes antigenic variation as a defense mechanism to circumvent host immune responses and thus a successful vaccine needs to provide strain-transcending and multicompartment (mucosal and skin) immunity. By defining highly conserved and protective epitopes from two critical virulence factors (M-protein and Spy-CEP) and combining them with a potent immunostimulant, CAF®01, we are addressing an unmet clinical need for a mucosally and skin-active subunit vaccine. We demonstrate that prime-pull immunization (2× intramuscular injections followed by intranasal immunization) promotes high sustained antibody levels in the airway mucosa and serum and protects against URT and invasive disease.

Preclinical safety and immunogenicity of Streptococcus pyogenes (Strep A) peptide vaccines.

We have developed two candidate vaccines to protect against multiple strains of Strep A infections. The candidates are combinatorial synthetic peptide vaccines composed of a M protein epitope (J8 or p*17) and a non-M protein epitope (K4S2). To enhance immunogenicity, each peptide is conjugated to the carrier protein CRM197 (CRM) and formulated with aluminium hydroxide adjuvant Alhydrogel (Alum) to make the final vaccines, J8-CRM + K4S2-CRM/Alum and p*17-CRM + K4S2-CRM/Alum. The safety and toxicity of each vaccine was assessed. Sprague Dawley rats were administered three intramuscular doses, over a six-week study with a 4-week recovery period. A control group received CRM only formulated with Alum (CRM/Alum). There was no evidence of systemic toxicity in the rats administered either vaccine. There was an associated increase in white blood cell, lymphocyte and monocyte counts, increased adrenal gland weights, adrenocortical hypertrophy, and increased severity of granulomatous inflammation at the sites of injection and the associated inguinal lymph nodes. These changes were considered non-adverse. All rats administered vaccine developed a robust and sustained immunological response. The absence of clinical toxicity and the development of an immunological response in the rats suggests that the vaccines are safe for use in a phase 1 clinical trial in healthy humans.

Immunogenicity and protective ability of RpoE against Streptococcus suis serotype 2.

AIMS: RpoE is quite immunogenic and can be used as a candidate vaccine for Streptococcus suis infection via immunoproteomics as reported in our previous studies. In this study, we aimed to verify the immunogenicity of recombinant RpoE and its protective effect against of S. suis. METHODS AND RESULTS: The RpoE protein was successfully expressed in Escherichia coli, and the purified recombinant protein was mixed with ISA206 to prepare an S. suis subunit vaccine. Mice were immunized with the RpoE subunit vaccine and then infected with the virulent S. suis strain ZY05719. Subunit vaccine-immunized mice achieved 50% protection, less pathological damage and less bacterial distribution in each organ compared with the control mice. Furthermore, in vitro culture, showed that mouse antisera significantly (P ï¼œ 0·001) inhibited the growth of S. suis, and qRT-PCR results showed that RpoE successfully induced the up-regulation of IL-6 and TNF-α cytokines. CONCLUSIONS: RpoE mice were vaccinated to obtain immune protection, which may be candidates for S. suis subunit vaccine. SIGNIFICANCE AND IMPACT OF THE STUDY: The results of this study will provide new ideas for the development of safe and effective recombinant subunits vaccines for S. suis.

Cross-serotype protection against group A Streptococcal infections induced by immunization with SPy_2191.

Group A Streptococcus (GAS) infection causes a range of diseases, but vaccine development is hampered by the high number of serotypes. Here, using reverse vaccinology the authors identify SPy_2191 as a cross-protective vaccine candidate. From 18 initially identified surface proteins, only SPy_2191 is conserved, surface-exposed and inhibits both GAS adhesion and invasion. SPy_2191 immunization in mice generates bactericidal antibodies resulting in opsonophagocytic killing of prevalent and invasive GAS serotypes of different geographical regions, including M1 and M49 (India), M3.1 (Israel), M1 (UK) and M1 (USA). Resident splenocytes show higher interferon-γ and tumor necrosis factor-α secretion upon antigen re-stimulation, suggesting activation of cell-mediated immunity. SPy_2191 immunization significantly reduces streptococcal load in the organs and confers ~76-92% protection upon challenge with invasive GAS serotypes. Further, it significantly suppresses GAS pharyngeal colonization in mice mucosal infection model. Our findings suggest that SPy_2191 can act as a universal vaccine candidate against GAS infections.

Vaccine-Induced Th1-Type Response Protects against Invasive Group A Streptococcus Infection in the Absence of Opsonizing Antibodies.

Recent global advocacy efforts have highlighted the importance of development of a vaccine against group A Streptococcus (GAS). Combo5 is a non-M protein-based vaccine that provides protection against GAS skin infection in mice and reduces the severity of pharyngitis in nonhuman primates. However, Combo5 with the addition of aluminum hydroxide (alum) as an adjuvant failed to protect against invasive GAS infection of mice. Here, we show that formulation of Combo5 with adjuvants containing saponin QS21 significantly improves protective efficacy, even though all 7 adjuvants tested generated high antigen-specific IgG antibody titers, including alum. Detailed characterization of Combo5 formulated with SMQ adjuvant, a squalene-in-water emulsion containing a TLR4 agonist and QS21, showed significant differences from the results obtained with alum in IgG subclasses generated following immunization, with an absence of GAS opsonizing antibodies. SMQ, but not alum, generated strong interleukin-6 (IL-6), gamma interferon (IFN-γ), and tumor necrosis alpha (TNF-α) responses. This work highlights the importance of adjuvant selection for non-M protein-based GAS vaccines to optimize immune responses and protective efficacy.IMPORTANCE Availability of a group A Streptococcus vaccine remains an unmet public health need. Here, we tested different adjuvant formulations to improve the protective efficacy of non-M protein vaccine Combo5 in an invasive disease model. We show that novel adjuvants can dramatically shape the type of immune response developed following immunization with Combo5 and significantly improve protection. In addition, protection afforded by Combo5 is not mediated by opsonizing antibodies, believed to be the main correlate of protection against GAS infections. Overall, this report highlights the importance of adjuvant selection in raising protective immune responses against GAS invasive infection. Adjuvants that can provide a more balanced Th1/Th2-type response may be required to optimize protection of GAS vaccines, particularly those based on non-M protein antigens.

Structure-based glycoconjugate vaccine design: The example of Group B Streptococcus type III capsular polysaccharide.

Microbial surface polysaccharides are important virulence factors and targets for vaccine development. Glycoconjugate vaccines, obtained by covalently linking carbohydrates and proteins, are well established tools for prevention of bacterial infections. Elucidation of the minimal portion involved in the interactions with functional antibodies is of utmost importance for the understanding of their mechanism of induction of protective immune responses and the design of synthetic glycan based vaccines. Typically, this is achieved by combination of different techniques, which include ELISA, glycoarray, Surface Plasmon Resonance in conjunction with approaches for mapping at atomic level the position involved in binding, such as Saturation Transfer NMR and X-ray crystallography. This review provides an overview of the structural studies performed to map glycan epitopes (glycotopes), with focus on the highly complex structure of Group B Streptococcus type III (GBSIII) capsular polysaccharide. Furthermore, it describes the rational process followed to translate the obtained information into the design of a protective glycoconjugate vaccine based on a well-defined synthetic glycan epitope.

Cross-immunity in Nile tilapia vaccinated with Streptococcus agalactiae and Streptococcus iniae vaccines.

Streptococcus agalactiae and Streptococcus iniae are major bacterial pathogens of tilapia that can cause high mortality concomitant with large economic losses to aquaculture. Although development of vaccines using formalin-killed bacteria to control these diseases has been attempted, the mechanism of immunity against streptococcal infections and the cross-protective ability of these two bacteria remains unclear. To explore the immunological role of these vaccines, we compared the immune responses of tilapia after immunization with both vaccines and compared the relative percent survival (RPS) and cross-immunization protection of tilapia after separate infection with S. agalactiae and S. iniae. All results revealed that vaccinated fish had significantly higher (P < 0.05) levels of specific antibodies than control fish 14 days post secondary vaccination (PSV) and 7 days post challenge. In vaccinated fish, the mRNA expression of interleukin-8 (IL-8), interleukin-12 (IL-12), caspase-3 (C-3), tumour necrosis factor (TNF), and interferon (IFN) was significantly up regulated (P < 0.05) in the head kidney after immunized; similar results were found for IL-8, TNF and IFN in the posterior kidney, meanwhile the expression levels of C-3 and IFN were significantly increased (P < 0.05) in the spleen of vaccinated fish. Additionally, the levels of acid phosphatase (ACP), alkaline phosphatase (AKP), superoxide dismutase (SOD), and lysozyme (LZM) in vaccinated fish were improved at different degree when compared to the control fish. These results showed that vaccination with formalin-killed cells (FKCs) of either S. agalactiae or S. iniae conferred protection against infection by the corresponding pathogen in Nile tilapia, resulting in RPS values of 92.3% and 91.7%, respectively. Furthermore, cross-protection was observed, as the S. agalactiae FKC vaccine protected fish from S. iniae infection, and vice versa. These results suggested that the S. agalactiae and S. iniae FKC vaccines can induce immune responses and generate excellent protective effects in Nile tilapia.