Recent Proceedings on Prevalence and Pathogenesis of Streptococcus suis.

Streptococcus suis (S. suis) is an important zoonotic pathogen that causes huge economic losses in the pig industry, as well as severe illness and even death in humans. The outbreak of human infection of S. suis in China in 2005 led to significant human morbidity and death, prompting an increase in global studies of S. suis. In recent years, important advances have been made regarding the etiology, genomics, excavation of virulence genes, and vaccine research in S. suis. A number of countries and regions have identified their predominantly serotypes. The development of genome sequencing technology has laid an important foundation for the study of pathogenic mechanisms. For example, 89K PAI was found in representative virulence strains in China, and several studies have been carried out to confirm multiple genes which carries are closely related to virulence. Also, the functions of some regulatory genes represented by the two-component signal transduction system have been analyzed. The development of inactivated vaccines, natural avirulent vaccines, gene-deletion attenuated vaccines, subunit vaccines, and glycoconjugate vaccines have greatly contributed to the prevention and control of the disease in the future. This article aims to summarize the research progress to provide directions for future research and the prevention of S. suis.

Molecular Evolution of Clinical Pathogenic Streptococci.

The genus Streptococcus comprises a wide variety of pathogenic and commensal gram-positive bacteria, many of which the pathogenic species cause severe, invasive infections that account for a high burden of morbidity and mortality. Here, we reviewed the evolution of representative virulence factors, capsule in Streptococcus pneumoniae, M protein in Streptococcus pyogenes (GAS), biofilm in Streptococcus agalactiae (GBS) and some oral Streptococcus, as well as the effect caused by evolution, antibiotic resistance and vaccine escape. Thanks to the rapid development of whole genome sequence (WGS) data, the impact of genetic recombination to the Streptococcus evolution has been proved. As to adaptive evolution caused by antibiotics, vaccine and so on, continuous surveillance is an essential to monitor evolution of Streptococcus causing disease. This knowledge is invaluable to the development of preventative and control strategies against this important pathogen.

Recent Advances in Pathogenic Streptococcus Vaccine Development.

Streptococcus pneumoniae (Spn) and Streptococcus pyogenes (Spy) cause many invasive and noninvasive diseases responsible for high morbidity and mortality worldwide. Safe, efficacious and affordable vaccines could have a significant, positive impact on the global infectious disease burden. Since the implementation of pneumococcal vaccine in the 1980s, the incidence of Spn infection has decreased significantly. Still so, these currently used multivalent polysaccharides and conjugated pneumococcal vaccines have some limitations. For Spy, there are even no vaccines available yet. There is an urgent need of new vaccines against Spn and Spy. Encouragingly, with the hard work of many investigators worldwide, a number of new vaccines candidates are developed with promising results. Of them, many have already entered the clinical trial stage. This review will describe the current status of Spn and Spy vaccine development, with particular focus on protein-based strategy.

Maternal immunization against Group B streptococcus: World Health Organization research and development technological roadmap and preferred product characteristics.

Group B streptococcus, found in the vagina or lower gastrointestinal tract of about 10-40% of women of reproductive age, is a leading cause of early life invasive bacterial disease, potentially amenable to prevention through maternal immunization during pregnancy. Following a consultation process with global stakeholders, the World Health Organization is herein proposing priority research and development pathways and preferred product characteristics for GBS vaccines, with the aim to facilitate and accelerate vaccine licensure, policy recommendation for wide scale use and implementation.

Comparative immunosecretome analysis of prevalent Streptococcus suis serotypes.

Streptococcus suis is a major Gram-positive swine pathogen associated with a wide variety of diseases in pigs. The efforts made to develop vaccines against this pathogen have failed because of lack of common cross-reactive antigens against different serotypes. Nowadays the interest has moved to surface and secreted proteins, as they have the highest chances to raise an effective immune response because they are in direct contact with host cells and are really exposed and accessible to antibodies. In this work, we have performed a comparative immunosecretomic approach to identify a set of immunoreactive secreted proteins common to the most prevalent serotypes of S. suis. Among the 67 proteins identified, three (SSU0020, SSU0934, and SSU0215) were those predicted extracellular proteins most widely found within the studied serotypes. These immunoreactive proteins may be interesting targets for future vaccine development as they could provide possible cross-reactivity among different serotypes of this pathogen.

WHO/IVI global stakeholder consultation on group A Streptococcus vaccine development: Report from a meeting held on 12-13 December 2016.

While progress towards a Group A Streptococcus (GAS) vaccine has been stalled by a combination of scientific, regulatory, and commercial barriers, the problem persists. The high and globally-distributed burden of disease attributable to GAS makes vaccination an imperative global public health goal. Advances across a range of scientific disciplines in understanding GAS diseases have made the goal a realistic one and focused attention on the need for coordinated global action. With a view to accelerating GAS vaccine development, the World Health Organization (WHO) and the International Vaccine Institute (IVI) convened a global stakeholder consultation on the 12th and 13th of December 2016, in Seoul, South Korea. Topics discussed included: (1) gaps in current knowledge of global GAS epidemiology, burden of disease, and molecular epidemiology; (2) contribution of pre-clinical models to candidate vaccine evaluation and new immunological assays to address GAS immunology knowledge gaps; (3) status and future of the GAS vaccine development pipeline; and (4) defining a pathway to licensure, policy recommendations and availability of a vaccine. The meeting determined to establish a GAS vaccine working group to coordinate preparation of a global vaccine values proposition, preferred product characteristics, and a technical research and development roadmap. A new global GAS vaccine consortium will drive strategic planning to anticipate requirements for licensure, prequalification, and policy recommendations.

Immunogenicity and prediction of epitopic region of antigen Ag I/II and glucosyltransferase from Streptococcus mutans.

The levels of Streptococcus (S.) mutans infections in saliva were evaluated and a comparison for specific antibody levels among children with different levels of S. mutans infection was made. The promising epitopic regions of antigen AgI/II (PAc) and glucosyltransferase (GTF) for potential vaccine targets related to S. mutans adherence were screened. A total of 94 children aged 3-4 years were randomly selected, including 53 caries-negative and 41 caries-positive children. The values of S. mutans and those of salivary total secretory immunoglobulin A (sIgA), anti-PAc and anti-Glucan binding domain (anti-GLU) were compared to determine the correlation among them. It was found the level of s-IgA against specific antigens did not increase with increasing severity of S. mutans infection, and the complete amino acid sequence of PAc and GTFB was analyzed using the DNAStar Protean system for developing specific anti-caries vaccines related to S. mutans adherence. A significantly positive correlation between the amount of S. mutans and children decayed, missing, and filled teeth index was observed. No significant difference was detected in specific sIgA against PAc or GLU between any two groups. No significant correlation was found between such specific sIgA and caries index. A total of 16 peptides from PAc as well as 13 peptides from GTFB were chosen for further investigation. S. mutans colonization contributed to early children caries as an important etiological factor. The level of sIgA against specific antigens did not increase with increasing severity of S. mutans infection in children. The epitopes of PAc and GTF have been screened to develop the peptide-based or protein-based anti-caries vaccines.

Predicted Coverage and Immuno-Safety of a Recombinant C-Repeat Region Based Streptococcus pyogenes Vaccine Candidate.

The C-terminal region of the M-protein of Streptococcus pyogenes is a major target for vaccine development. The major feature is the C-repeat region, consisting of 35-42 amino acid repeat units that display high but not perfect identity. SV1 is a S. pyogenes vaccine candidate that incorporates five 14mer amino acid sequences (called J14i variants) from differing C-repeat units in a single recombinant construct. Here we show that the J14i variants chosen for inclusion in SV1 are the most common variants in a dataset of 176 unique M-proteins. Murine antibodies raised against SV1 were shown to bind to each of the J14i variants present in SV1, as well as variants not present in the vaccine. Antibodies raised to the individual J14i variants were also shown to bind to multiple but different combinations of J14i variants, supporting the underlying rationale for the design of SV1. A Lewis Rat Model of valvulitis was then used to assess the capacity of SV1 to induce deleterious immune response associated with rheumatic heart disease. In this model, both SV1 and the M5 positive control protein were immunogenic. Neither of these antibodies were cross-reactive with cardiac myosin or collagen. Splenic T cells from SV1/CFA and SV1/alum immunized rats did not proliferate in response to cardiac myosin or collagen. Subsequent histological examination of heart tissue showed that 4 of 5 mice from the M5/CFA group had valvulitis and inflammatory cell infiltration into valvular tissue, whereas mice immunised with SV1/CFA, SV1/alum showed no sign of valvulitis. These results suggest that SV1 is a safe vaccine candidate that will elicit antibodies that recognise the vast majority of circulating GAS M-types.

Immunoproteomic assay of secreted proteins of Streptococcus suis serotype 9 with convalescent sera from pigs.

Streptococcus suis is an important pathogen of pigs. In China, in addition to S. suis serotype 2, S. suis serotype 9 (SS9) is also a prevalent serotype. There is no vaccine available for SS9. An immunoproteome-based approach was developed to identify SS9 immunogenic proteins for vaccine development. Secreted proteins extracted from SS9 strain GZ0565 were screened by two-dimensional Western blotting using convalescent sera from pigs. Protein spots were excised from preparative gels and were identified by matrix-assisted laser desorption ionization time-of-flight mass spectrometry, which led to the identification of ten immunogenic proteins (sortases, ABC transporter substrate-binding protein-maltose/maltodextrin, ABC transporter periplasmic protein, CHAP domain containing protein, peptidoglycan-binding LysM, elongation factor Tu, elongation factor G, thymidine kinase, molecular chaperone DnaK, hypothetical protein SSU98_2184). These novel immunogenic proteins, which are encoded by genes that are reasonably conserved among SS9 strains, may be developed as antigens for further study of SS9 vaccine.

[Protective efficacy of a new fusion anti-caries DNA vaccine encoding antigens of both Streptococcus mutans and Streptococcus sobrinus].

OBJECTIVE: To construct a new fusion anti-caries DNA vaccine pGJGAC/VAX encoding antigens of both S. mutans and S. sobrinus so as to enhance the protective effect of DNA vaccine against S. sobrinus infection. METHODS: The CAT fragment of S. sobrinus OMZ176 gtf-I was amplified by semi-nest PCR and then inserted into the plasmid pGJA-P/VAX to construct the recombinant plasmid pGJGAC/VAX. The CHO cell was transfected and the expression of fusion protein detected using cellular immunohistochemistry and Western blot. Mice were immunized with pGJGAC/VAX and control plasmids via the intramuscular (i.m) or intranasal (i.n) routes. During the experiment, blood and saliva samples were collected at a 2-week interval for antibody assay by ELISA. Rats were orally challenged with S. mutans Ingbritt or S. sobrinus 6715 and then immunized i.n with pGJGAC/VAX, pGJA-P/VAX or pVAX1. The Keyes method was used to determine the caries activity. RESULTS: (1) CAT sequence was identical to the related sequence of gtf-I (OMZ176) in GenBank. The recombinant plasmid pGJGAC/VAX encoded the genes of antigens of both S. mutans and S. sobrinus. The expressed protein could respond to specific anti-PAc, anti-GLU and anti-CAT antibodies respectively. (2) As for antibody reactions, mice in the experiment group had significantly higher levels of anti-PAc, anti-GLU and anti-CAT IgG antibodies than those in the pVAX1 group (P < 0.01). The peak responses of specific anti-CAT antibodies were observed at 8 weeks (GAC/i.m) and 10 weeks (GAC/i.n) and were approximately 62.13 microg/ml and 11.43 microg/ml respectively. The peak responses of specific anti-CAT IgA antibodies were seen at 8 weeks (GAC/i.m) and 10 weeks (GAC/i.n) and were approximately 0.67% and 0.80% respectively. (3) In the group infected with S. mutans or S. sobrinus, the pGJGAC/VAX-immunized rats showed significantly fewer E, Ds and Dm lesions than pVAX1-immunized rats (P < 0.05) and decreased Ds and Dm levels than pGJA-P/VAX-immunized rats (P < 0.05) while there was no obvious difference in E lesions between the two groups (P > 0.05). CONCLUSION: A new fusion anti-caries DNA vaccine pGJGAC/VAX encoding antigens of both S. mutans and S. sobrinus is constructed successfully and expressed correctly in eukaryotic cells. It induces effective mucosal and systematic humoral responses so as to provide a better protection against S. sobrinus.

Identification of new candidate vaccine antigens made by Streptococcus pyogenes: purification and characterization of 16 putative extracellular lipoproteins.

Putative extracellular lipoproteins made by group A Streptococcus (GAS) are the focus of this study, which was designed to identify new candidate vaccine antigens. Bioinformatic analysis of a serotype M1 GAS strain identified 30 open-reading frames encoding putative lipoproteins. The genes encoding the mature form of 29 of these proteins were cloned, and 16 recombinant proteins were overexpressed in Escherichia coli and purified to apparent homogeneity. The genes encoding these 16 proteins were highly conserved in GAS strains for which genome sequence data are available (serotypes M1, M3, M5, M12, M18, and M28). Mice inoculated subcutaneously with GAS and humans with GAS pharyngitis and invasive infections seroconverted to most of the 16 recombinant proteins, which indicates that these lipoproteins were produced during infection. The blood of mice actively immunized with 5 of the 16 recombinant proteins had significantly (P<.05) increased growth-inhibitory activity, compared with the blood of unimmunized mice, which identified these proteins as potential new vaccine candidates.