Streptococcus suis 5'-nucleotidases contribute to adenosine-mediated immune evasion and virulence in a mouse model.

Streptococcus suis (S. suis) is an important swine bacterial pathogen and causes human infections, leading to a wide range of diseases. However, the role of 5'-nucleotidases in its virulence remains to be fully elucidated. Herein, we identified four cell wall-anchored 5'-nucleotidases (Snts) within S. suis, named SntA, SntB, SntC, and SntD, each displaying similar domains yet exhibiting low sequence homology. The malachite green reagent and HPLC assays demonstrated that these recombinant enzymes are capable of hydrolysing ATP, ADP, and AMP into adenosine (Ado), with the hierarchy of catalytic efficiency being SntC>SntB>SntA>SntD. Moreover, comprehensive enzymatic activity assays illustrated slight variances in substrate specificity, pH tolerance, and metal ion requirements, yet highlighted a conserved substrate-binding pocket, His-Asp catalytic dyad, metal, and phosphate-binding sites across Snts, with the exception of SntA. Through bactericidal assays and murine infection assays involving in site-mutagenesis strains, it was demonstrated that SntB and SntC collaboratively enhance bacterial survivability within whole blood and polymorphonuclear leukocytes (PMNs) via the Ado-A2aR pathway in vitro, and within murine blood and organs in vivo. This suggests a direct correlation between enzymatic activity and enhancement of bacterial survival and virulence. Collectively, S. suis 5'-nucleotidases additively contribute to the generation of adenosine, influencing susceptibility within blood and PMNs, and enhancing survival within blood and organs in vivo. This elucidation of their integral functions in the pathogenic process of S. suis not only enhances our comprehension of bacterial virulence mechanisms, but also illuminates new avenues for therapeutic intervention aimed at curbing S. suis infections.

A bifunctional amylopullulanase of Streptococcus suis ApuA contributes to immune evasion by interaction with host complement C3b.

The complement system is the first defense line of the immune system. However, pathogens have evolved numerous strategies to evade complement attacks. Streptococcus suis is an important zoonotic bacterium, harmful to both the pig industry and human health. ApuA has been reported as a bifunctional amylopullulanase and also contributed to virulence of S. suis. Herein, we found that ApuA could activate both classical and alternative pathways of the complement system. Furthermore, by using bacterial two-hybrid, far-western blot and ELISA assays, it was confirmed that ApuA could interact with complement C3b. The interaction domain of ApuA with C3b was found to be its α-Amylase domain (ApuA_N). After construction of an apuA mutant (ΔapuA) and its complementary strain, it was found that compared to the wild-type strain (WT), ΔapuA had significantly increased C3b deposition and membrane attack complex formation. Additionally, ΔapuA showed significantly lower survival rates in human serum and blood and was more susceptible to engulfment by neutrophils and macrophages. Mice infected with ΔapuA had significantly higher survival rates and lower bacterial loads in their blood, lung and brains, compared to those infected with WT. In summary, this study identified ApuA as a novel factor involved in the complement evasion of S. suis and suggested its multifunctional role in the pathogenesis of S. suis.

The granulocyte colony-stimulating factor produced during Streptococcus suis infection controls neutrophil recruitment in the blood without affecting bacterial clearance.

Streptococcus suis causes diseases in pigs and has emerged as a zoonotic agent. When infected, the host develops an exacerbated inflammation that can lead to septic shock and meningitis. Although neutrophils greatly infiltrate the lesions, their dynamics during S. suis infection remain poorly described. Moreover, very few studies reported on the production and role of a key factor in the regulation of neutrophils: the colony-stimulating granulocyte factor (G-CSF). In this study, we characterized the G-CSF-neutrophil axis in the pathogenesis of S. suis induced disease. Using a mouse model of S. suis infection, we first evaluated the recruitment of neutrophils and their activation profile by flow cytometry. We found that infection provokes a massive neutrophil recruitment from the bone marrow to the blood and spleen. In both compartments, neutrophils displayed multiple activation markers. In parallel, we observed high systemic levels of G-CSF, with a peak of production coinciding with that of neutrophil recruitment. We then neutralized the effects of G-CSF and highlighted its role in the release of neutrophils from the bone marrow to the blood. However, it did not affect bacteremia nor the cytokine storm induced by S. suis. In conclusion, systemic G-CSF induces the release of neutrophils from the bone marrow to the blood, but its role in inflammation or bacterial clearance seems to be compensated by unknown factors. A better understanding of the role of neutrophils and inflammatory mediators could lead to better strategies for controlling the infection caused by S. suis.

The hypervirulent Group B Streptococcus HvgA adhesin promotes central nervous system invasion through transcellular crossing of the choroid plexus.

BACKGROUND: Group B Streptococcus (GBS) is the leading cause of neonatal meningitis responsible for a substantial cause of death and disability worldwide. The vast majority of GBS neonatal meningitis cases are due to the CC17 hypervirulent clone. However, the cellular and molecular pathways involved in brain invasion by GBS CC17 isolates remain largely elusive. Here, we studied the specific interaction of the CC17 clone with the choroid plexus, the main component of the blood-cerebrospinal fluid (CSF) barrier. METHODS: The interaction of GBS CC17 or non-CC17 strains with choroid plexus cells was studied using an in vivo mouse model of meningitis and in vitro models of primary and transformed rodent choroid plexus epithelial cells (CPEC and Z310). In vivo interaction of GBS with the choroid plexus was assessed by microscopy. Bacterial invasion and cell barrier penetration were examined in vitro, as well as chemokines and cytokines in response to infection. RESULTS: GBS CC17 was found associated with the choroid plexus of the lateral, 3rd and 4th ventricles. Infection of choroid plexus epithelial cells revealed an efficient internalization of the bacteria into the cells with GBS CC17 displaying a greater ability to invade these cells than a non-CC17 strain. Internalization of the GBS CC17 strain involved the CC17-specific HvgA adhesin and occurred via a clathrin-dependent mechanism leading to transcellular transcytosis across the choroid plexus epithelial monolayer. CPEC infection resulted in the secretion of several chemokines, including CCL2, CCL3, CCL20, CX3CL1, and the matrix metalloproteinase MMP3, as well as immune cell infiltration. CONCLUSION: Our findings reveal a GBS strain-specific ability to infect the blood-CSF barrier, which appears to be an important site of bacterial entry and an active site of immune cell trafficking in response to infection.

Immunogenicity of a 30-valent M protein mRNA group A Streptococcus vaccine.

BACKGROUND: Group A Streptococcus (Strep A) causes both uncomplicated and severe invasive infections, as well as the post-infection complications acute rheumatic fever and rheumatic heart disease. Despite the high global burden of disease resulting from Strep A infections, there is not a licensed vaccine. A 30-valent M protein-based vaccine has previously been shown to be immunogenic in animal models and in a Phase I clinical trial (NCT02564237). Here, we assessed the immunogenicity of a 30-valent messenger (m)RNA vaccine designed to express the same M peptide targets as the 30-valent protein vaccine and compared it with the protein vaccine. METHODS: Female New Zealand white rabbits were immunized with one of four vaccine formulations (3 doses of each formulation at days 1, 28, and 56): soluble mRNA (100 µg/animal), C-terminal transmembrane mRNA (100 µg/animal), protein vaccine (400 µg/animal), or a non-translatable RNA control (100 µg/animal). Serum was collected one day prior to the first dose and on days 42 and 70. Rabbit serum samples were assayed for antibody levels against synthetic M peptides by ELISA. HL-60 opsonophagocytic killing (OPK) assays were performed to assess functional antibody levels. RESULTS: Serum IgG levels were similar for the mRNA and protein vaccines. The CtTM version of the mRNA vaccine elicited slightly higher antibody levels than the mRNA designed to express soluble proteins. OPK activity was similar for the mRNA and protein vaccines, regardless of M type. CONCLUSIONS: The total antibody responses and functional antibody levels elicited by the 30-valent mRNA Strep A vaccines were similar to those observed following immunization with the analogous protein vaccine. The mRNA vaccine platform provides potential advantages to protein-based vaccines including inherent adjuvant activity, increased production efficiency, lower cost, and the potential to rapidly change epitopes/peptides, all of which are important considerations related to multivalent Strep A vaccine development.

New vaccination approach using formalin-killed Streptococcus pyogenes vaccine on the liver of Oreochromis niloticus fingerlings.

Newly synthesized vaccines prepared from formalin-killed bacteria Streptococcus pyogenes were investigated in the current study to evaluate the effectiveness of the newly synthesized vaccine as well as their safety by injected intraperitoneal. The study involved several steps 1st step is the preparation of the vaccine followed by the 2nd step: Evaluate the effectiveness and vaccine safety against pathogenic S. pyogenes through 4 different groups including control (Group I). Group II (Bacterial, infected group), Group III (Vaccine), and the Last group was the challenged group after the vaccination (Vacc + Bac). Different Immunological and biochemical parameters were measured in addition to hematological and histopathological examinations. For example, oxidative/antioxidants, inflammatory biomarkers, fragmentation and cell damage, and finally the histopathological study. The current study showed an increase in all oxidative, inflammatory, and cell damage (DNA fragmentation assays), additionally markedly elevation in histopathological cell damage in the infected group (Group II) compared with the control group. The vaccine and challenged after vaccination group (vaccine + Bacteria), showed great improvement in oxidative biomarkers (LPO) and an increase in antioxidants biomarkers (GSH, SOD, GST, DPPH, ABTS, GR and GPx), Also the inflammation and histopathological examination. The newly synthesized vaccine improved the resistance of Oreochromis niloticus and can be used as a preventive therapy agent for pathogenic bacteria S. pyogenes.

Inhibition of Macrophage Recruitment to Heart Valves Mediated by the C-C Chemokine Receptor Type 2 Attenuates Valvular Inflammation Induced by Group A Streptococcus in Lewis Rats.

BACKGROUND: Rheumatic heart disease (RHD) is an autoimmune disease caused by recurrent infections of Group A streptococcus (GAS), ultimately leading to inflammation and the fibrosis of heart valves. Recent studies have highlighted the crucial role of C-C chemokine receptor type 2-positive (CCR2+) macrophages in autoimmune diseases and tissue fibrosis. However, the specific involvement of CCR2+ macrophages in RHD remains unclear. METHODS: This study established an RHD rat model using inactivated GAS and complete Freund's adjuvant, demonstrating a correlation between CCR2+ macrophages and fibrosis in the mitral valves of these rats. RESULTS: Intraperitoneal injection of the CCR2 antagonist Rs-504393 significantly reduced macrophage infiltration, inflammation, and fibrosis in valve tissues of RHD rats compared to the solvent-treated group . Existing evidence suggests that C-C motif chemokine ligand 2 (CCL2) acts as the primary recruiting factor for CCR2+ cells. To validate this, human monocytic leukemia cells (THP-1) were cultured in vitro to assess the impact of recombinant CCL2 protein on macrophages. CCL2 exhibited pro-inflammatory effects similar to lipopolysaccharide (LPS), promoting M1 polarization in macrophages. Moreover, the combined effect of LPS and CCL2 was more potent than either alone. Knocking down CCR2 expression in THP-1 cells using small interfering RNA suppressed the pro-inflammatory response and M1 polarization induced by CCL2. CONCLUSIONS: The findings from this study indicate that CCR2+ macrophages are pivotal in the valvular remodeling process of RHD. Targeting the CCL2/CCR2 signaling pathway may therefore represent a promising therapeutic strategy to alleviate valve fibrosis in RHD.

Molecular structure and functional responses of IgM, IgT and IgD to Flavobacterium covae and Streptococcus iniae infection in Asian seabass (Lates calcarifer Bloch, 1790).

The Asian seabass (Lates calcarifer) faces significant disease threats, which are exacerbated by intensive farming practices and environmental changes. Therefore, understanding its immune system is crucial. The current study presents a comprehensive analysis of immune-related genes in Asian seabass peripheral blood leukocytes (PBLs) using Iso-seq technology, identifying 16 key pathways associated with 7857 immune-related genes, comprising 634 unique immune-related genes. The research marks the first comprehensive report on the entire immunoglobulin repertoire in Asian seabass, revealing specific characteristics of immunoglobulin heavy chain constant region transcripts, including IgM (Cµ, ighm), IgT (Cτ, ight), and IgD (Cδ, ighd). The study confirms the presence of membrane-bound form, ighmmb, ightmb, ighdmb of IgM, IgT and IgD and secreted form, ighmsc and ightsc of IgM and IgT, respectively, with similar structural patterns and conserved features in amino acids across immunoglobulin molecules, including cysteine residues crucial for structural integrity observed in other teleost species. In response to bacterial infections by Flavobacterium covae (formerly F. columnare genomovar II) and Streptococcus iniae, both secreted and membrane-bound forms of IgM (ighmmb and ighmsc) and IgT (ightmb and ightsc) show significant expression, indicating their roles in systemic and mucosal immunity. The expression of membrane-bound form IgD gene, ighdmb, predominantly exhibits targeted upregulation in PBLs, suggesting a regulatory role in B cell-mediated immunity. The findings underscore the dynamic and tissue-specific expression of immunoglobulin repertoires, ighmmb, ighmsc, ightmb, ightsc and ighdmb in Asian seabass, indicating a sophisticated immune response to bacterial pathogens. These findings have practical implications for fish aquaculture, and disease control strategies, serving as a valuable resource for advancing research in Asian seabass immunology.

The role of HMGB2 in the immune response of Nile tilapia (Oreochromis niloticus) to streptococcal infection.

High mobility group protein B2 (HMGB2) is an abundant chromatin-associated protein with pivotal roles in transcription, cell proliferation, differentiation, inflammation, and tumorigenesis. However, its immune function in Nile tilapia (Oreochromis niloticus) remains unclear. In this study, we identified a homologue of HMGB2 from Nile tilapia (On-HMGB2) and investigated its functions in the immune response against streptococcus infection. The open reading frame (ORF) of On-HMGB2 spans 642 bp, encoding 213 amino acids, and contains two conserved HMG domains. On-HMGB2 shares over 80 % homology with other fish species and 74%-76 % homology with mammals. On-HMGB2 was widely distributed in various tissues, with its highest transcript levels in the liver and the lowest in the intestine. Knockdown of On-HMGB2 promoted the inflammatory response in Nile tilapia, increased the bacterial load in the tissues, and led to elevated mortality in Nile tilapia following Streptococcus agalactiae infection. Taken together, On-HMGB2 significantly influences the immune system of Nile tilapia in response to streptococcus infection.

Vaccines for Streptococcus agalactiae: current status and future perspectives.

A maternal vaccine to protect newborns against invasive Streptococcus agalactiae infection is a developing medical need. The vaccine should be offered during the third trimester of pregnancy and induce strong immune responses and placental transfer of protective antibodies. Polysaccharide vaccines against S. agalactiae conjugated to protein carriers are in advanced stages of development. Additionally, protein-based vaccines are also in development, showing great promise as they can provide protection regardless of serotype. Furthermore, safety concerns regarding a new vaccine are the main barriers identified. Here, we present vaccines in development and identified safety, cost, and efficacy concerns, especially in high-need, low-income countries.

Neonatal intestinal colonization of Streptococcus agalactiae and the multiple modes of protection limiting translocation.

Streptococcus agalactiae, also known as Group B Streptococcus (GBS), is a predominant pathogen of neonatal sepsis, commonly associated with early-onset neonatal sepsis. GBS has also been associated with cases of late-onset sepsis potentially originating from the intestine. Previous findings have shown GBS can colonize the infant intestinal tract as part of the neonatal microbiota. To better understand GBS colonization dynamics in the neonatal intestine, we collected stool and milk samples from prematurely born neonates for identification of potential pathogens in the neonatal intestinal microbiota. GBS was present in approximately 10% of the cohort, and this colonization was not associated with maternal GBS status, delivery route, or gestational weight. Interestingly, we observed the relative abundance of GBS in the infant stool negatively correlated with maternal IgA concentration in matched maternal milk samples. Using a preclinical murine model of GBS infection, we report that both vertical transmission and direct oral introduction resulted in intestinal colonization of GBS; however, translocation beyond the intestine was limited. Finally, vaccination of dams prior to breeding induced strong immunoglobulin responses, including IgA responses, which were associated with reduced mortality and GBS intestinal colonization. Taken together, we show that maternal IgA may contribute to infant immunity by limiting the colonization of GBS in the intestine.

Streptococcus suis Serotype 2 Type IV Secretion Effector SspA-1 Induces Proinflammatory Cytokine Production via TLR2 Endosomal and Type I Interferon Signaling.

The subtilisin-like protease-1 (SspA-1) plays an important role in the pathogenesis of a highly virulent strain of Streptococcus suis 2. However, the mechanism of SspA-1-triggered excessive inflammatory response is still unknown. In this study, we demonstrated that activation of type I IFN signaling is required for SspA-1-induced excessive proinflammatory cytokine production. Further experiments showed that the TLR2 endosomal pathway mediates SspA-1-induced type I IFN signaling and the inflammatory response. Finally, we mapped the major signaling components of the related pathway and found that the TIR adaptor proteins Mal, TRAM, and MyD88 and the downstream activation of IRF1 and IRF7 were involved in this pathway. These results explain the molecular mechanism by which SspA-1 triggers an excessive inflammatory response and reveal a novel effect of type I IFN in S. suis 2 infection, possibly providing further insights into the pathogenesis of this highly virulent S. suis 2 strain.

Elevated antibody binding to striatal cholinergic interneurons in patients with pediatric acute-onset neuropsychiatric syndrome.

Pediatric Acute-onset Neuropsychiatric Syndrome (PANS) is characterized by the abrupt onset of significant obsessive-compulsive symptoms (OCS) and/or severe food restriction, together with other neuropsychiatric manifestations. An autoimmune pathogenesis triggered by infection has been proposed for at least a subset of PANS. The older diagnosis of Pediatric Autoimmune Neuropsychiatric Disorder Associated with Streptococcus (PANDAS) describes rapid onset of OCD and/or tics associated with infection with Group A Streptococcus. The pathophysiology of PANS and PANDAS remains incompletely understood. We recently found serum antibodies from children with rigorously defined PANDAS to selectively bind to cholinergic interneurons (CINs) in the striatum. Here we examine this binding in children with relapsing and remitting PANS, a more heterogeneous condition, collected in a distinct clinical context from those examined in our previous work, from children with a clinical history of Streptococcus infection. IgG from PANS cases showed elevated binding to striatal CINs in both mouse and human brain. Patient plasma collected during symptom flare decreased a molecular marker of CIN activity, phospho-riboprotein S6, in ex vivo brain slices; control plasma did not. Neither elevated antibody binding to CINs nor diminished CIN activity was seen with plasma collected from the same children during remission. These findings replicate what we have seen previously in PANDAS and support the hypothesis that at least a subset of PANS cases have a neuroimmune pathogenesis. Given the critical role of CINs in modulating basal ganglia function, these findings confirm striatal CINs as a locus of interest in the pathophysiology of both PANS and PANDAS.

Medulla oblongata and NCCs are central defenders against Streptococcus agalactiae infection of the tilapia brain.

Various types of professional immune cells first emerge in fish and likely represent the primordial form and functions. Recent advancements revealed the direct connection between the central nervous system and the immune system in the mammalian brain. However, the specifics of brain-immune networks in the fish and the underlying mechanisms of teleost's brain against pathogen infection have not been fully elucidated. In this study, we investigated the distribution of markers representing cerebral cells associated with protection and professional lymphocytes in the seven major components of the Nile tilapia brain through RNA-Seq assay and observed the most dominant abundance in the medulla oblongata. The subsequent challenge test revealed the non-specific cytotoxic cells (NCCs) exhibited the strongest response against streptococcal infection of the brain. The presence of NCCs in the brain was then confirmed using immunofluorescence and the cytotoxic effects usually induced by NCCs under infection were determined as well. Collectively, these findings contribute significantly to comprehending the mechanism of fish neuroimmune interaction and enhancing our understanding of its evolutionary development.

Streptococcus anginosus: A new pathogen of superficial gastritis, atrophic gastritis and gastric cancer.

A wealth of research indicates that superficial gastritis (SG) and atrophic gastritis (AG) are precursors to gastric cancer (GC). While Helicobacter pylori (H. pylori) has long been recognized as a key player in GC development, recent findings by Fu et al. have identified Streptococcus anginosus (S. anginosus) as an emerging pathogen that can trigger SG, AG and GC. S. anginosus, a gram-positive coccus, leverages its surface protein T. pallidum membrane protein C (TMPC) to engage with the annexin A2 (ANXA2) receptor of gastric epithelial cells, facilitating its colonization and invasion in the gastric mucosa. This leads to an upregulation of proinflammatory chemokines Ccl20 and Ccl8, causing prolonged effects on gastric barrier function and microbiota homeostasis, leading to SG. Moreover, these bacteria activate the mitogen-activated protein kinase (MAPK) signaling pathway, which is associated with the development of AG and GC. Importantly, inhibiting TMPC or knocking down ANXA2 can reduce S. anginosus colonization and invasion, lowering the chances of SG, AG, and GC. This paper highlights the molecular mechanisms of S. anginosus in SG, AG and GC, emphasizing the importance of a multi-pathogen strategy in gastric disease management and the need for further investigation into the role of S. anginosus in GC progression.

Deficiency of hasB accelerated the clearance of Streptococcus equi subsp. Zooepidemicus through gasdermin d-dependent neutrophil extracellular traps.

Streptococcus equi subsp. zooepidemicus (S. zooepidemicus, SEZ) is an essential zoonotic bacterial pathogen that can cause various inflammation, such as meningitis, endocarditis, and pneumonia. UDP-glucose dehydrogenase (hasB) is indispensable in synthesizing SEZ virulence factor hyaluronan capsules. Our study investigated the infection of hasB on mice response to SEZ by employing a constructed capsule-deficient mutant strain designated as the ΔhasB strain. This deficiency was associated with a reduced SEZ bacterial load in the mice's blood and peritoneal lavage fluid (PLF) post-infection. Besides, the ΔhasB SEZ strain exhibited a higher propensity for neutrophil infiltration and release of cell-free DNA (cfDNA) in vivo compared to the wild-type (WT) SEZ strain. In vitro experiments further revealed that ΔhasB SEZ more effectively induced the formation of neutrophil extracellular traps (NETs) containing histone 3 (H3), neutrophil elastase (NE), and DNA, than its WT counterpart. Moreover, the release of NETs was determined to be gasdermin D (GSDMD)-dependent during the infection process. Taken together, these findings underscore that the deficiency of the hasB gene in SEZ leads to enhanced GSDMD-dependent NET release from neutrophils, thereby reducing SEZ's capacity to resist NETs-mediated eradication during infection. Our finding paves the way for the development of innovative therapeutic strategies against SEZ.

The suppressive role of NLRP6 in host defense against Streptococcus suis infection.

Streptococcus suis (S. suis) disease is a prevalent zoonotic infectious threat that elicits a systemic inflammatory response in both swine and humans, frequently culminating in high mortality rates. The excessive inflammation triggered by S. suis infection can precipitate tissue damage and sudden death; however, a comprehensive strategy to mitigate this inflammatory response remains elusive. Our study examines the role of NLRP6 in S. suis infection, with a particular focus on its involvement in pathogen regulation. A marked upregulation of NLRP6 was observed in peritoneal macrophages post-infection with S. suis SC19 strain, consequently activating the NLRP6 inflammasome. Furthermore, SC19 infection was found to augment the secretion of pro-inflammatory cytokines IL-1ß via NLRP6 activation, while NLRP6 deficiency mitigates the invasion and adhesion of SC19 to macrophages. In vivo models revealed that NLRP6 deletion enhanced survival rates of SC19-infected mice, alongside a reduction in tissue bacterial load and inflammatory cytokine levels. NLRP6-/- mice were shown to exhibit attenuated inflammatory responses in pulmonary, hepatic, and splenic tissues post-SC19 infection, as evidenced by lower inflammation scores. Flow cytometry analyses further substantiated that NLRP6 is involved in modulating macrophage and neutrophil recruitment during infection. Our findings suggest that NLRP6 negatively regulates host resistance against S. suis infection; its absence results in reduced mortality, bacterial colonization, and a milder inflammatory response. Elucidating the mechanism of NLRP6 in S. suis-induced inflammation provides novel insights and theoretical underpinnings for the prophylaxis and therapeutics of S. suis diseases.

Experimental Intraperitoneal Infection of Piglets.

Streptococcus suis is a swine bacterial pathogen that predominantly causes disease in weaned piglets characterized by swelling of joints, arthritis, septicemia, meningitis, and sudden death. Intravenous, intramuscular, intraperitoneal, and intranasal infection models were developed to study the bacterial pathogenicity and efficacy of vaccines and various therapeutics. The selection of the appropriate infection model is a critical step in any study, as it may impact the outcomes of the study. Here we describe a method for infecting weaned piglets with S. suis using intraperitoneal route as a reliable, consistent, and reproducible animal model to evaluate vaccine protection against systemic bacterial infection.

Preparation and Study of Bacterins.

Streptococcus suis is a bacterial pathogen that can cause significant economic losses in the swine industry due to high morbidity and mortality rates in infected animals. Vaccination with bacterins, which consist of inactivated bacteria and adjuvants to enhance the pig's immune response, is an effective approach to control S. suis infections in piglets. Here we provide a description of S. suis bacterins and the methods for vaccine preparation. Moreover, this chapter also describes the addition of recombinant Sao (rSao-L) protein to the S. suis bacterin, aiming to enhance the efficacy of the bacterins against S. suis in piglets. Furthermore, the methods for evaluating the immune response elicited by the bacterins are also covered in this chapter.