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As a zoonotic pathogen, S. suis serotype 2 (SS2) can cause severe diseases in both pigs and humans, and develop resistance to antibiotics. Plant natural compounds are regarded as promising alternatives to conventional antibiotics. Phillyrin is the major bioactive components of Chinese herbal medicine Forsythia suspensa. In this study, we explored the activity and action mechanism of phillyrin against SS2. The results showed that phillyrin could disrupt membrane integrity, destroy intracellular structures, and increase the exosmosis of DNA. Results of PCR revealed that phillyrin affected bacterial-virulence-related genes' expression levels. Meanwhile, phillyrin significantly decreased the adhesion activity, inhibited lactate dehydrogenase (LDH) secretion, and reduced biofilm formation of SS2 in Newborn pig trachea epithelial (NPTr) cells. Furthermore, phillyrin protected tight junction protein of NPTr cells from SS2. We reported that phillyrin (0.1 mg/kg) treatment after bacterial challenge significantly improved the survival rate, ameliorated pulmonary inflammation, and inhibited the accumulation of multiple cytokines (IL-1, IL-6, IL-8, and TNF-α). Molecular docking showed that phillyrin had a good binding activity with the Ala88 and Asp111 of suilysin (SLY), one of the most important virulence factors of SS2. Collectively, phillyrin possesses antibacterial and anti-inflammatory activities, and is a promising candidate for preventing SS2 infection. [ABSTRACT FROM AUTHOR]

Streptococcus (S.) suis presents a serious threat to the pig industry as well as food safety and public health. Although several LAMP assays have been developed for the identification of S. suis, no universal assay is so far available for the field-suitable examination of clinical pig specimens. Based on the thrA housekeeping gene, a new loop-mediated isothermal amplification (LAMP) assay was developed and validated for the detection of S. suis in the brain and joints of pigs. For this LAMP assay, two different methods for the extraction of DNA from brain and joint swabs were compared. Using the LPTV boiling method, the detection limit of LAMP was 1.08 CFU/reaction, while the detection limit was 53.8 CFU/reaction using a commercial DNA extraction kit. The detection limits of thrA-LAMP in combination with the LPTV boiling method were 104–105 CFU/swab in the presence of brain tissue and 103–104 CFU/swab in the presence of joint tissue. The diagnostic quality criteria of LAMP were determined by the examination of 49 brain swabs and 34 joint swabs obtained during routine diagnostic necropsies. Applying the LPTV boiling method to brain swabs, the sensitivity, specificity, and positive and negative predictive values of thrA-LAMP were 88.0, 95.8, 95.7, and 88.5% using cultural investigation as a reference method, and 76.7, 100, 100, and 73.1% using real-time PCR as a reference method. Based on these results, the thrA-LAMP assay combined with the LPTV boiling method is suitable for rapid detection of S. suis from brain swabs. [ABSTRACT FROM AUTHOR]

ABSTRACT Streptococcus suis is a zoonotic pathogen that causes disease in humans after exposure to infected pigs or pig-derived food products. In this study, we examined the serotype distribution, antimicrobial resistance phenotypes and genotypes, integrative and conjugative elements (ICEs), and associated genomic environments of S. suis isolates from humans and pigs in China from 2008 to 2019. We identified isolates of 13 serotypes, predominated by serotype 2 (40/96; 41.7%), serotype 3 (10/96; 10.4%), and serotype 1 (6/96; 6.3%). Whole-genome sequencing analysis revealed that these isolates possessed 36 different sequence types (STs), and ST242 and ST117 were the most prevalent. Phylogenetic analysis revealed possible animal and human clonal transmission, while antimicrobial susceptibility testing indicated high-level resistance to macrolides, tetracyclines, and aminoglycosides. These isolates carried 24 antibiotic resistance genes (ARGs) that conferred resistance to 7 antibiotic classes. The antibiotic resistance genotypes were directly correlated with the observed phenotypes. We also identified ICEs in 10 isolates, which were present in 4 different genetic environments and possessed differing ARG combinations. We also predicted and confirmed by PCR analysis the existence of a translocatable unit (TU) in which the oxazolidinone resistance gene optrA was flanked by IS1216E elements. One-half (5/10) of the ICE-carrying strains could be mobilized by conjugation. A comparison of the parental recipient with an ICE-carrying transconjugant in a mouse in vivo thigh infection model indicated that the ICE strain could not be eliminated with tetracycline treatment. S. suis therefore poses a significant challenge to global public health and requires continuous monitoring, especially for the presence of ICEs and associated ARGs that can be transferred via conjugation. IMPORTANCE S. suis is a serious zoonotic pathogen. In this study, we investigated the epidemiological and molecular characteristics of 96 S. suis isolates from 10 different provinces of China from 2008 to 2019. A subset of these isolates (10) carried ICEs that were able to be horizontally transferred among isolates of different S. suis serotypes. A mouse thigh infection model revealed that ICE-facilitated ARG transfer promoted resistance development. S. suis requires continuous monitoring, especially for the presence of ICEs and associated ARGs that can be transferred via conjugation.

The Netherlands has been very successful in the last decade in reducing antimicrobial use in animals. On about a quarter of farms, antimicrobial use in weaned pigs remains relatively high. As Streptococcus suis (S. suis) infections are responsible for a high consumption of antimicrobials, a specific veterinary guideline to control S. suis was developed, but seemed to be poorly adopted by veterinarians. Guided by the theoretical domains framework, the aim of this study was to identify determinants influencing veterinarians' adherence to this guideline. We interviewed 13 pig veterinarians. Interviewees described multiple approaches to managing S. suis problems and adherence to the guideline. Mentioned determinants could be categorized into 12 theoretical domains. The following six domains were mentioned in all interviews: knowledge, skills, beliefs about capabilities, beliefs about consequences, social influences, and environmental context and resources. The insights derived from this study are relevant for understanding factors influencing veterinarians' adoption of scientific evidence and guidelines and can be used to develop evidence-based implementation strategies for veterinary guidelines. [ABSTRACT FROM AUTHOR]

Streptococcus (S.) suis presents a serious threat to the pig industry as well as food safety and public health. Although several LAMP assays have been developed for the identification of S. suis, no universal assay is so far available for the field-suitable examination of clinical pig specimens. Based on the thrA housekeeping gene, a new loop-mediated isothermal amplification (LAMP) assay was developed and validated for the detection of S. suis in the brain and joints of pigs. For this LAMP assay, two different methods for the extraction of DNA from brain and joint swabs were compared. Using the LPTV boiling method, the detection limit of LAMP was 1.08 CFU/reaction, while the detection limit was 53.8 CFU/reaction using a commercial DNA extraction kit. The detection limits of thrA-LAMP in combination with the LPTV boiling method were 104–105 CFU/swab in the presence of brain tissue and 103–104 CFU/swab in the presence of joint tissue. The diagnostic quality criteria of LAMP were determined by the examination of 49 brain swabs and 34 joint swabs obtained during routine diagnostic necropsies. Applying the LPTV boiling method to brain swabs, the sensitivity, specificity, and positive and negative predictive values of thrA-LAMP were 88.0, 95.8, 95.7, and 88.5% using cultural investigation as a reference method, and 76.7, 100, 100, and 73.1% using real-time PCR as a reference method. Based on these results, the thrA-LAMP assay combined with the LPTV boiling method is suitable for rapid detection of S. suis from brain swabs.

Streptococcus suis is an emerging bacterial pathogen of huge economic impact to the swine industry worldwide. The information regarding the carrier status of S. suis in the slaughtered pigs along with its genetic characterization is not available in Indian pig population, which needs to be addressed for the therapeutic and preventive measures. In the present study, 563 palatine tonsils of apparently healthy slaughtered pigs were probed for the prevalence, and genetic characterization of S. suis and prevalence were found to be 15.45% and 32.68% by bacteriological and molecular methods, respectively. In 87 isolates recovered, 6 cps-types were detected showing the predominance of serotype 7 (24.13%) and 5 (18.39%), whereas 11 cps-types were detected in tonsillar DNA involving cps-types 9 (28.26%) and 7 (14.13%) as the major serotypes with arcA+/sly+/epf+/mrp− being the prevalent genotype. The histopathological changes with the immunodetection of S. suis antigen confirmed its persistence in asymptomatic carriers. Of 87 bacterial isolates, 7 isolates (serotypes 7 & 2) were pathogenic to Swiss albino mice showing the classical lesions of meningitis and septicemia. The presence of virulent serotypes of S. suis in healthy slaughtered pigs suggests a great health risk to the people engaged in piggery operations and in-contact pigs. [ABSTRACT FROM AUTHOR]

Abstract Background Laboratory facilities for etiological diagnosis of central nervous system (CNS) infection are limited in developing countries; therefore, patients are treated empirically, and the epidemiology of the pathogens is not well-known. Tubercular meningitis is one of the common causes of meningitis, which has high morbidity and mortality, but lacks sensitive diagnostic assays. The objectives of this study were to determine the causes of meningitis in adult patients by using molecular assays, to assess the risk factors associated with them, and to explore whether biomarkers can differentiate tubercular meningitis from bacterial meningitis. Methods We conducted a cross-sectional study in the Department of Infectious Diseases, Bach Mai Hospital, Hanoi, Vietnam, from June 2012 to May 2014. All patients who were ≥ 16 years old and who had meningoencephalitis suggested by abnormal cerebrospinal fluid (CSF) findings (CSF total cell >5/mm3 or CSF protein ≥40 mg/dL) were included in the study. In addition to culture, CSF samples were tested for common bacterial and viral pathogens by polymerase chain reaction (PCR) and for biomarkers: C-reactive protein and adenosine deaminase (ADA). Results Total number of patients admitted to the department was 7506; among them, 679 were suspected to have CNS infection, and they underwent lumbar puncture. Five hundred eighty-three patients had abnormal CSF findings (meningoencephalitis); median age was 45 (IQR 31–58), 62.6% were male, and 60.9% were tested for HIV infection. Among 408 CSF samples tested by PCR, out of them, 358 were also tested by culture; an etiology was identified in 27.5% (n=112). S. suis (8.8%), N. meningitis (3.2%), and S. pneumoniae (2.7%) were common bacterial and HSV (2.2%), Echovirus 6 (0.7%), and Echovirus 30 (0.7%) were common viral pathogens detected. M. tuberculosis was found in 3.2%. Mixed pathogens were detected in 1.8% of the CSF samples. Rural residence (aOR 4.1, 95% CI 1.2–14.4) and raised CSF ADA (≥10 IU/L) (aOR 25.5, 95% CI 3.1–212) were associated with bacterial meningitis when compared with viral meningitis; similarly, raised CSF ADA (≥10 IU/L) (aOR 42.2, 95% CI 2.0–882) was associated with tubercular meningitis. Conclusions Addition of molecular method to the conventional culture had enhanced the identification of etiologies of CNS infection. Raised CSF ADA (≥10 IU/L) was strongly associated with bacterial and tubercular meningitis. This biomarker might be helpful to diagnose tubercular meningitis once bacterial meningitis is ruled out by other methods.

Abstract Streptococcus suis (S. suis) is an important zoonotic pathogen that causes septicaemia, meningitis and streptococcal toxic shock-like syndrome in its host, and recent studies have shown that S. suis could be competent for natural genetic transformation. Transformation is an important mechanism for the horizontal transfer of DNA, but some elements that affect the transformation process need to be further explored. Upon entering the competent state, Streptococcus species stimulate the transcription of competence-related genes that are responsible for exogenous DNA binding, uptake and processing. In this study, we performed conserved promoter motif and qRT-PCR analyses and identified CrfP as a novel murein hydrolase that is widespread in S. suis and stimulated with a peptide pheromone in the competent state through a process controlled by ComX. A bioinformatics analysis revealed that CrfP consists of a CHAP hydrolase domain and two bacterial Src homology 3-binding (SH3b) domains. Further characterization showed that CrfP could be exported to extracellular bacterial cells and lytic S. suis strains of different serotypes, and this finding was verified by TEM and a turbidity assay. To investigate the potential effect of CrfP in vivo, a gene-deletion mutant (ΔcrfP) was constructed. Instead of stopping the natural transformation process, the inactivation of CrfP clearly reduced the effective transformation rate. Overall, these findings provide evidence showing that CrfP is important for S. suis serovar 2 competence.

Gamithromycin is a long-acting azalide antibiotic that has been developed recently for the treatment of swine respiratory diseases. In this study, the pharmacokinetic/pharmacodynamic (PK/PD) targets, PK/PD cutoff, and optimum dosing regimen of gamithromycin were evaluated in piglets against Streptococcus suis in China, including a subset with capsular serotype 2. Short post-antibiotic effects (PAEs) (0.5–2.6 h) and PA-SMEs (2.4–7.7 h) were observed for gamithromycin against S. suis. The serum matrix dramatically facilitated the intracellular uptake of gamithromycin by S. suis strains, thus contributing to the potentiation effect of serum on their susceptibilities, with a Mueller-Hinton broth (MHB)/serum minimum inhibitory concentration (MIC) ratio of 28.86 for S. suis. Dose-response relationship demonstrated the area under the concentration (AUC)/MIC ratio to be the predictive PK/PD index closely linked to activity (R2 > 0.93). For S. suis infections, the net stasis, 1–log10, and 2–log10 kill effects were achieved at serum AUC24h/MIC targets of 17.9, 49.1, and 166 h, respectively. At the current clinical dose of 6.0 mg/kg, gamithromycin PK/PD cutoff value was determined to be 8 mg/L. A PK/PD-based dose assessment demonstrated that the optimum dose regimen of gamithromycin to achieve effective treatments for the observed wild-type MIC distribution of S. suis in China with a probability of target attainment (PTA) ≥ 90% was 2.53 mg/kg in this study. These results will aid in the development of clinical dose-optimization studies and the establishment of clinical breakpoints for gamithromycin in the treatment of swine respiratory infections due to S. suis.

The Netherlands has been very successful in the last decade in reducing antimicrobial use in animals. On about a quarter of farms, antimicrobial use in weaned pigs remains relatively high. As Streptococcus suis (S. suis) infections are responsible for a high consumption of antimicrobials, a specific veterinary guideline to control S. suis was developed, but seemed to be poorly adopted by veterinarians. Guided by the theoretical domains framework, the aim of this study was to identify determinants influencing veterinarians’ adherence to this guideline. We interviewed 13 pig veterinarians. Interviewees described multiple approaches to managing S. suis problems and adherence to the guideline. Mentioned determinants could be categorized into 12 theoretical domains. The following six domains were mentioned in all interviews: knowledge, skills, beliefs about capabilities, beliefs about consequences, social influences, and environmental context and resources. The insights derived from this study are relevant for understanding factors influencing veterinarians’ adoption of scientific evidence and guidelines and can be used to develop evidence-based implementation strategies for veterinary guidelines.

Macrolide-resistant Streptococcus suis is highly prevalent worldwide. The acquisition of the erm(B) gene mediated by mobile genetic elements (MGEs) in particular integrative and conjugative elements (ICEs) is recognized as the main reason for the rapid spread of macrolide-resistant streptococcal strains. However, knowledge about different erm(B)-carrying elements responsible for the widespread of macrolide resistance and their transferability in S. suis remains poorly understood. In the present study, two erm(B)- and tet(O)-harboring putative ICEs, designated as ICESsuYSB17_rplL and ICESsuYSJ15_rplL, and a novel erm(B)- and aadE-spw-like-carrying genomic island (GI), named GISsuJHJ17_rpsI, were identified to be excised from the chromosome and transferred among S. suis strains with different serotypes. ICESsuYSB17_rplL and ICESsuYSJ15_rplL were integrated downstream the rplL gene, a conserve locus of the ICESa2603 family. GISsuJHJ17_rpsI, with no genes belonging to the conjugation module, was integrated into the site of rpsI. All transconjugants did not exhibit obvious fitness cost by growth curve and competition assays when compared with the recipient. The results demonstrate that different erm(B)-carrying elements were presented and highlight the role of these elements in the dissemination of macrolide resistance in S. suis.

Macrolide-resistant Streptococcus suis is highly prevalent worldwide. The acquisition of the erm (B) gene mediated by mobile genetic elements (MGEs) in particular integrative and conjugative elements (ICEs) is recognized as the main reason for the rapid spread of macrolide-resistant streptococcal strains. However, knowledge about different erm (B)-carrying elements responsible for the widespread of macrolide resistance and their transferability in S. suis remains poorly understood. In the present study, two erm (B)- and tet (O)-harboring putative ICEs, designated as ICE Ssu YSB17_ rplL and ICE Ssu YSJ15_ rplL , and a novel erm (B)- and aadE-spw -like-carrying genomic island (GI), named GI Ssu JHJ17_ rpsI , were identified to be excised from the chromosome and transferred among S. suis strains with different serotypes. ICE Ssu YSB17_ rplL and ICE Ssu YSJ15_ rplL were integrated downstream the rplL gene, a conserve locus of the ICE Sa 2603 family. GI Ssu JHJ17_ rpsI , with no genes belonging to the conjugation module, was integrated into the site of rpsI. All transconjugants did not exhibit obvious fitness cost by growth curve and competition assays when compared with the recipient. The results demonstrate that different erm (B)-carrying elements were presented and highlight the role of these elements in the dissemination of macrolide resistance in S. suis. [ABSTRACT FROM AUTHOR]

Purpose: Streptococcus suis (S. suis) is an emerging zoonotic disease mainly in pigs, causing serious infections in humans with high prevalence in Southeast Asia. Despite a relatively high mortality rate, there are limited data regarding the risk factors of this life-threatening infection. Therefore, a 13-year retrospective cohort study in Chiang Mai, Thailand during 2005–2018 was conducted to explore risk factors associated with S. suis mortality and to update the outcomes of the disease. Patients and methods: S. suis positive cases were derived from those with positive S. suis isolates from microbiological culture results and Matrix-Assisted Laser Desorption Ionization Time of Flight (MALDI-TOF). Potential risk factors of mortality were identified using univariate and multivariate logistic regression. Results: Of 133 patients with culture-proven S. suis infection identified, there were 92 males and 41 females. The mean age was 56.47 years. Septicemia (55.64%) was the most common clinical manifestation followed by meningitis (37.59%) and infective endocarditis (25.56%). Alcohol drinking and raw pork consumption were documented in 66 (49.62%) and 49 (36.84%) cases respectively. The overall mortality rate was 12.03% (n=16). According to the multivariate analysis, the independent risk factors for mortality were prolonged bacteremia ≥ 6 days (OR = 43.57, 95% CI = 2.46–772.80, P =0.010), septic shock (OR = 13.34, 95% CI = 1.63–109.03, P =0.016), and direct bilirubin > 1.5 mg/dL (OR = 12.86, 95% CI = 1.91–86.59, P =0.009). Conclusion: S. suis is not infrequent in Northern Thailand, where the cultural food habit of raw pork eating is still practiced. To the best of our knowledge, this is the largest series focusing on risk factors of S. suis mortality which has been conducted in Thailand. Prolonged bacteremia ≥ 6 days, septic shock, and direct bilirubin > 1.5 mg/dL were strong predictors associated with S. suis mortality. The mortality risk factors identified may be further utilized in clinical practice and future research to improve patient outcomes. [ABSTRACT FROM AUTHOR]

Streptococcus suis is difficult to treat and responsible for various infections in humans and pigs. It can also form biofilms and induce persistent infections. Rhizoma Coptidis is a medicinal plant widely used in Traditional Chinese Medicine. Although the inhibitory effects of Rhizoma Coptidis on biofilm formation have been investigated in several studies, the ability of Rhizoma Coptidis to inhibit S. suis biofilm formation and the underlying mechanisms have not yet been reported. In this study, we showed that sub-minimal inhibitory concentrations (25 and 50 μg mL-1) of water extracts of Rhizoma Coptidis (Coptis deltoidea C.Y.Cheng & P.K.Hsiao, obtained from Sichuan Province) were sufficient to inhibit biofilm formation, as shown in the tissue culture plate (TCP) method and scanning electron microscopy. Real-time PCR and iTRAQ were used to measure gene and protein expression in S. suis. Sub-minimum inhibitory concentrations (25 and 50 μg mL-1) of Rhizoma Coptidis water extracts inhibited S. suis adhesion significantly in an anti-adherence assay. Some genes, such as gapdh, sly, and mrp, and proteins, such as antigen-like protein, CPS16V, and methyltransferase H, involved in adhesion were significantly modulated in cells treated with 50 μg mL-1 of Rhizoma Coptidis water extracts compared to untreated cells. The results from this study suggest that compounds in Rhizoma Coptidis water extracts play an important role in inhibiting adhesion of S. suis cells and, therefore, biofilm formation.

Streptococcus suis is difficult to treat and responsible for various infections in humans and pigs. It can also form biofilms and induce persistent infections. Rhizoma Coptidis is a medicinal plant widely used in Traditional Chinese Medicine. Although the inhibitory effects of Rhizoma Coptidis on biofilm formation have been investigated in several studies, the ability of Rhizoma Coptidis to inhibit S. suis biofilm formation and the underlying mechanisms have not yet been reported. In this study, we showed that subminimal inhibitory concentrations (25 and 50 mg mL-1) of water extracts of Rhizoma Coptidis (Coptis deltoidea C.Y.Cheng & P.K.Hsiao, obtained from Sichuan Province) were sufficient to inhibit biofilm formation, as shown in the tissue culture plate (TCP) method and scanning electron microscopy. Real-time PCR and iTRAQ were used to measure gene and protein expression in S. suis. Sub-minimum inhibitory concentrations (25 and 50 mg mL-1) of Rhizoma Coptidis water extracts inhibited S. suis adhesion significantly in an anti-adherence assay. Some genes, such as gapdh, sly, and mrp, and proteins, such as antigen-like protein, CPS16V, and methyltransferase H, involved in adhesion were significantly modulated in cells treated with 50 mg mL-1 of Rhizoma Coptidis water extracts compared to untreated cells. The results from this study suggest that compounds in Rhizoma Coptidis water extracts play an important role in inhibiting adhesion of S. suis cells and, therefore, biofilm formation. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Weaning piglets are susceptible to the invasive Streptococcus (S.) suis infection, which can result in septicemia. The aim of this study was to investigate the cytokine profile induced upon S. suis infection of blood, to determine the cellular sources of those cytokines, and to study the potential effects of the induced cytokines on bacterial killing. We measured TNF-α, IL-6, IFN-γ, IL-17A and IL-10 after an experimental intravenous infection with S. suis serotype 2 in vivo, and analyzed whole blood, peripheral blood mononuclear cells (PBMC) and separated leukocytes to identify the cytokine-producing cell type(s). In addition, we used a reconstituted whole blood assay to investigate the effect of TNF-α on bacterial killing in the presence of different S. suis-specific IgG levels. An increase in IL-6 and IL-10, but not in IFN-γ or IL-17A, was observed in two of three piglets with pronounced bacteremia 16 to 20 h after infection, but not in piglets with controlled bacteremia. Our results confirmed previous findings that S. suis induces TNF-α and IL-6 and could demonstrate that TNF-α is produced by monocytes in vitro. We further found that IL-10 induction resulted in reduced secretion of TNF-α and IL-6. Rapid induction of TNF-α was, however, not crucial for in vitro bacterial killing, not even in the absence of specific IgG.

Streptococcus suis (S. suis) is a crucial zoonotic pathogen which causes severe diseases in humans and pigs. Biofilms of S. suis can induce persistent infections which are harder to treat. Sub-minimal inhibitory concentration (sub-MIC) of tylosin can inhibit biofilm formation in bacteria. By using iTRAQ strategy, we compared the protein expression profiles of S. suis grown with sub-MIC tylosin treatment or no treatement. The result showed that 96 proteins expression were changed with 77 up-regulated and 19 down-regulated proteins. Several metabolism proteins (such as phosphoglycerate kinase), as well as cell surface proteins (such as ABC transporter proteins), were found to be involved in biofilm formation. Overall, our results indicated that S. suis metabolic regulation, cell surface proteins, and virulence proteins appear to be of importance in biofilm growth by sub-MIC tylosin treated. Thus, our data analyzed rough regulation of biofilm formation that lay the foundation for the future research of mechanism and targets.

Dry-cured hams, shoulders and loins of Iberian pigs are highly appreciated in national and international markets. Salting, additive addition and dehydration are the main strategies to produce these ready-to-eat products. Although the dry curing process is known to reduce the load of well-known food borne pathogens, studies evaluating the viability of other microorganisms in contaminated pork have not been performed. In this work, the efficacy of the dry curing process to eliminate three swine pathogens associated with pork carcass condemnation, Streptococcus suis, Streptococcus dysgalactiae and Trueperella pyogenes, was evaluated. Results of this study highlight that the dry curing process is a suitable method to obtain safe ready-to-eat products free of these microorganisms. Although salting of dry-cured shoulders had a moderate bactericidal effect, results of this study suggest that drying and ripening were the most important stages to obtain dry-cured products free of these microorganisms. [ABSTRACT FROM AUTHOR]

The efforts made to develop vaccines against Streptococcus suis have failed because of lack of common antigens cross-reactive against different serotypes of this species. The cell wall-anchored proteins can be good vaccine candidates due to their high expression and accessibility to antibodies, among these, a cell-wall protein, DNA-nuclease (SsnA), present in most of the S. suis serotypes and clinical isolates collected from infected pigs, was selected. An experimental challenge against S. suis serotype 2 in a pig model was used to validate the efficacy of recombinant SsnA combined with aluminium hydroxide plus Quil A as adjuvants, previously tested in mice by our research group with good results. In our study, clinical characteristics, bacterial load and spread, haematological and immunological parameters and the antibody response, including the opsonophagocytosis analysis of the sera were evaluated. Moreover the composition of peripheral blood leukocyte populations was studied in infected animals. The results show that the immunization of piglets with rSsnA elicits a significant humoral antibody response. However, the antibody response is not reflected in protection of pigs that are challenged with a virulent strain in our conventional vaccination model. Further studies are necessary to evaluate the use of rSsnA as a vaccine candidate for swine. [ABSTRACT FROM AUTHOR]

2-deoxyribose-5-phosphate aldolase (DERA) is a class I aldolase that catalyzes aldol condensation of two aldehydes in the active site, which is particularly germane in drug manufacture. Structural and biochemical studies have shown that the active site of DERA is typically loosely packed and displays broader substrate specificity despite sharing conserved folding architecture with other aldolases. The most distinctive structural feature of DERA compared to other aldolases is short and flexible C-terminal region. This region is also responsible for substrate recognition. Therefore, substrate tolerance may be related to the C-terminal structural features of DERA. Here, we determined the crystal structures of full length and C-terminal truncated DERA from Streptococcus suis (SsDERA). In common, both contained the typical (α/β) TIM-barrel fold of class I aldolases. Surprisingly, C-terminal truncation resulting in missing the last α9 and β8 secondary elements, allowed DERA to maintain activity comparable to the fulllength enzyme. Specifically, Arg186 and Ser205 residues at the C-terminus appeared mutually supplemental or less indispensible for substrate phosphate moiety recognition. Our results suggest that DERA might adopt a shorter C-terminal region than conventional aldolases during evolution pathway, resulting in a broader range of substrate tolerance through active site flexibility. [ABSTRACT FROM AUTHOR]

Membrane attack complex/perforin/cholesterol-dependent cytolysin (MACPF/CDC) proteins constitute a major superfamily of pore-forming proteins that act as bacterial virulence factors and effectors in immune defence. Upon binding to the membrane, they convert from the soluble monomeric form to oligomeric, membrane-inserted pores. Using real-time atomic force microscopy (AFM), electron microscopy (EM), and atomic structure fitting, we have mapped the structure and assembly pathways of a bacterial CDC in unprecedented detail and accuracy, focussing on suilysin from Streptococcus suis. We show that suilysin assembly is a noncooperative process that is terminated before the protein inserts into the membrane. The resulting ring-shaped pores and kinetically trapped arc-shaped assemblies are all seen to perforate the membrane, as also visible by the ejection of its lipids. Membrane insertion requires a concerted conformational change of the monomeric subunits, with a marked expansion in pore diameter due to large changes in subunit structure and packing.

Streptococcus suis serotype 2 (S. suis 2) is a zoonotic pathogen that exhibits high-level resistance and multi-drug resistance to classic antibiotics and causes serious human casualties and heavy economic losses in the swine industry worldwide. Therefore, alternative therapies or novel antibacterial agents need to be developed to combat this pathogen. A novel endolysin derived from the S. suis temperate phage phi7917, termed Ly7917, was identified, which had broad lytic activity against S. suis type 1, 2, 7 and 9. Ly7917 consisted of an N-terminal cysteine, histidine-dependent amidohydrolases/ peptidase catalytic domain and C-terminal SH3b cell wall binding domain. The endolysin maintained activity at high pH and its catalytic activity could be improved by addition of 10 μM 1.5 mM Ca2+. In animal studies, 90% of BALB/c mice challenged with typical virulent strain HA9801 of S. suis 2 were protected by Ly7917 treatment. The bacterial load in the blood of HA9801-challenged mice was efficiently reduced almost 50% by Ly7917 while that of penicillin-G-treated mice kept almost unchanged. Our data suggest that Ly7917 may be an alternative therapeutic agent for infections caused by virulent S. suis strains. [ABSTRACT FROM AUTHOR]

Streptococcus suis is one of the most important swine pathogens worldwide. In this study, a total of 22 virulence-related genes in 101 strains of S. suis serotype 2 (SS2) were detected by PCR, namely, mrp , epf , sly , fbps , rgg , ofs , srtA , pgdA , gapdh , iga , endoD , ciaRH , salKR , manN , purD , rgg , DppIV , neuB , dltA , SMU_61-like, SpyM3_0908 (Permease) and the SspA gene. The distribution of virulence-related genes among isolates was visualized using BioNumerics software to study similarities among the isolates. Two clusters of SS2 were apparent on the phylogenetic tree, namely, Clusters A and B. Both mouse and zebrafish infection models revealed that strains in Cluster B were more virulent than those in Cluster A. Statistical comparison between the two clusters was performed, and structure analysis demonstrated that epf , sly , rgg , endoD , SMU_61-like and SpyM3_0908 were possible predictive markers for SS2 virulence. The transcription levels of highly prevalent genes in both clusters were detected by qRT-PCR in representative strains. For Cluster A isolates, the transcription levels of neuB , dltA , fbps and pgdA were significantly lower, but the transcription level of iga was significantly higher than that in Cluster B isolates. Although encoded in the genomes of the selected Cluster A isolates, DppIV and mrp genes were not expressed. These results revealed the genetic differences between virulent and low-virulence SS2 isolates from China and provide a better understanding of the SS2 pathogenic mechanism. [ABSTRACT FROM AUTHOR]

Streptococcus suis ( S. suis ) is a swine pathogen and also a zoonotic agent. Biofilms of S. suis may cause persistent infections by the host immune system and antibiotics. Sub-minimal inhibitory concentration (sub-MIC) of erythromycin can inhibit biofilm formation in bacteria. Here, we performed comparative proteomic analyses of cells at two different conditions: sub-MIC erythromycin treated and nontreated cells. Using iTRAQ strategy, we found some novel proteins that involved in biofilm formation. 79 differentially expressed proteins were identified in sub-MIC erythromycin inhibiting planktonic cell when the protein had both a fold-change of more that a ratio > 1.2 or < 0.8 (p-value < 0.05). Several cell surface proteins (such as Primosomal protein N′, l -fucose isomerase, and ABC superfamily ATP binding cassette transporter, membrane protein), as well as those involved in Quorum-sensing, were found to be implicated in biofilm formation. Overall, our results indicated that cell surface proteins played an important role in biofilm formation. Quorum-sensing played a crucial role leading to biofilm formation. ABC superfamily ATP binding cassette transporter, membrane protein and comD might act as channels for erythromycin uptake in Quorum-sensing system. Thus, our data analyzed rough regulatory pathways of biofilm formation that might potentially be exploited to deal with biofilm infections of S. suis . This article is part of a Special Issue entitled: Microbial Proteomics. Biological significance In this study, we identified many proteins involved in cell transport, biological regulation and signal transduction, stress responses and other metabolic processes that were not previously known to be associated with biofilm formation of S. suis and target spot of erythromycin. Therefore, our manuscript represents the most comprehensive analysis of protein profiles of biofilm formation of S. suis inhibited by sub-MIC erythromycin and provides new proteomic information about biofilm formation. [ABSTRACT FROM AUTHOR]

Invasive Streptococcus suis (S. suis) infections in pigs are often associated with serotypes 2 and 9. Mucosal sites of healthy pigs can be colonized with these serotypes, often multiple serotypes per pig. To unravel the contribution of these serotypes in pathogenesis and epidemiology, simultaneous quantification of serotypes is needed. A quantitative real-time PCR (qPCR) targeting cps2J (serotypes 2 and 1/2) and cps9H (serotype 9) was evaluated with nasal and tonsillar samples from S. suis exposed pigs. qPCR specifically detected serotypes in all pig samples. The serotypes loads in pig samples estimated by qPCR showed, except for serotype 9 in tonsillar samples (correlation coefficient = 0.25), moderate to strong correlation with loads detected by culture (correlation coefficient > 0.65), and also in pigs exposed to both serotypes (correlation coefficient > 0.75). This qPCR is suitable for simultaneous differentiation and quantification of important S. suis serotypes.

Streptococcus (S.) suis infections are the most common cause of meningitis in pigs. Moreover, S. suis is a zoonotic pathogen, which can lead to meningitis in humans, mainly in adults. We assume that glial cells may play a crucial role in host-pathogen interactions during S. suis infection of the central nervous system. Glial cells are considered to possess important functions during inflammation and injury of the brain in bacterial meningitis. In the present study, we established primary astrocyte-microglial cell co-cultures to investigate interactions of S. suis with glial cells. For this purpose, microglial cells and astrocytes were isolated from new-born mouse brains and characterized by flow cytometry, followed by the establishment of astrocyte and microglial cell mono-cultures as well as astrocyte-microglial cell co-cultures. In addition, we prepared microglial cell mono-cultures co-incubated with uninfected astrocyte mono-culture supernatants and astrocyte mono-cultures co-incubated with uninfected microglial cell mono-culture supernatants. After infection of the different cell cultures with S. suis, bacteria-cell association was mainly observed with microglial cells and most prominently with a non-encapsulated mutant of S. suis. A time-dependent induction of NO release was found only in the co-cultures and after co-incubation of microglial cells with uninfected supernatants of astrocyte mono-cultures mainly after infection with the capsular mutant. Only moderate cytotoxic effects were found in co-cultured glial cells after infection with S. suis. Taken together, astrocytes and astrocyte supernatants increased interaction of microglial cells with S. suis. Astrocyte-microglial cell co-cultures are suitable to study S. suis infections and bacteria-cell association as well as NO release by microglial cells was enhanced in the presence of astrocytes.

Streptococcus suis is a pathogen of zoonotic diseases. Moreover, the emergence of fluoroquinolones (FQs) resistance in this pathogen has severe consequences for pigs and human health. In this study, the molecular mechanism of FQs resistance in S. suis type 2 (SS2) sensitive strains isolated from pigs was assessed after in vitro induction of resistance against the most frequently used FQs: ciprofloxacin, norfloxacin, and enrofloxacin. Proteome analysis, sequencing and real-time RT-PCR results strongly established an overexpression of an ABC transporter protein (other than SatAB) and topoisomerase mutations in GyrA (Ser81Arg), GyrB (Glu354Lys), and ParC (Ser79Phe) in contributing to high level ciprofloxacin resistance in SS2. Due to the overexpression of the ABC transporter, intracellular ciprofloxacin concentrations were significantly lower in the resistant strains than those of sensitive strains after 20, 35, and 60 min exposures to ciprofloxacin (p < 0.05). It was concluded that improper use of FQs is one of the main causes of the emergence of this zoonotic pathogen as a multiresistant organism against commonly used antibiotics. The existence of an efflux-like protein is an incentive to find new drug targets to avoid the spread of FQs-resistant S. suis isolates in pigs and the human population. [ABSTRACT FROM AUTHOR]

The article discusses a study on the antimicrobial sensitivity of Escherichia coli, Salmonella spp., Pasteurella multocida, Streptococcus suis, and Actinobacillus pleropneumoniae isolated from diagnostic samples from large pig breeding farms in Croatia. The researchers found that S. suis isolates were most sensitive to cefotaxime and florfenicol but had the highest resistance to streptomycin.

Emergence of fatal Streptococcus suis serotype 2 ( S. suis 2, SS2) in China, 2005, has greatly challenged the public health, raising serious concerns about its global circulation. Whole genome sequencing of the representative strains has identified a potential pathogenicity island (PAI), named 89K. Here, we reported the recurrence of human S. suis meningitis cases (two in Shenzhen City and one in Sichuan Province) in China, in 2007. Multiple lines of evidence supported that these human cases are caused by highly pathogenic SS2 isolates. Pulsed-field gel electrophoresis analysis indicated that these re-emerging SS2 isolates (07CQH01, 07SZH01 and 07SZH02) can be featured with different genotypes. Molecular dissection demonstrated that they all share 89K, the newly proposed PAI that is common to 2005 prevalent strains in China. Further mapping of 89K revealed that gene losses/deletions occur in 07CQH01 and 07SZH01, except 07SZH02. These findings suggested that heterogeneous SS2 circulates in China, with the capability of causing endemic with life-threatening danger. [ABSTRACT FROM AUTHOR]

Abstract: Streptococcus suis was isolated from 6.1% of raw pork meat from 3 of the 6 wet markets in 6 districts in Hong Kong. S. suis was particularly isolated in sites from the tongue, tonsil, bone, and tail, but not from lean meat/minced pork or internal organs. Isolates were confirmed by polymerase chain reaction using S. suis-specific primers, did not belong to serotype 2 using serotype 2-specific antiserum, and were clustered closely with other known serotypes by phylogenetic analysis. Ten strains from patients admitted to Hong Kong hospitals with sepsis or meningitis in the past 10 years all belonged to type 2, with closely related pulsed-field gel electrophoresis types that were distinct from the S. suis strains isolated from pork in this study. These methods may serve as useful tools in studying and enhancing our understanding of these infections in Hong Kong. [Copyright &y& Elsevier]

The synthesis of a selection of multivalent arrays of mannose mono- and disaccharides, that are of potential use as anti-infective agents against enterobacteria infections, is described. [Copyright &y& Elsevier]

Summary: The intergenic spacer region (ISR) between the 16S and 23S rRNA genes was tested as a tool for differentiating lactococci commonly isolated in a dairy environment. 17 reference strains, representing 11 different species belonging to the genera Lactococcus, Streptococcus, Lactobacillus, Enterococcus and Leuconostoc, and 127 wild streptococcal strains isolated during the whole fermentation process of “Fior di Latte” cheese were analyzed. After 16S–23S rDNA ISR amplification by PCR, species or genus-specific patterns were obtained for most of the reference strains tested. Moreover, results obtained after nucleotide analysis show that the 16S–23S rDNA ISR sequences vary greatly, in size and sequence, among Lactococcus garvieae, Lactococcus raffinolactis, Lactococcus lactis as well as other streptococci from dairy environments. Because of the high degree of inter-specific polymorphism observed, 16S–23S rDNA ISR can be considered a good potential target for selecting species-specific molecular assays, such as PCR primer or probes, for a rapid and extremely reliable differentiation of dairy lactococcal isolates. [Copyright &y& Elsevier]

The one-pot synthesis of multivalent arrays of mannose mono- and disaccharides, of potential use as anti-infective agents against enterobacteria infections, is described. [Copyright &y& Elsevier]