Effects of a Berry Polyphenolic Fraction on the Pathogenic Properties of Porphyromonas gingivalis.

Porphyromonas gingivalis expresses a broad array of virulence factors that enable it to play a central role in the etiopathogenesis of periodontitis. The objective of the present study was to assess the effects of a berry polyphenolic fraction (Orophenol®) composed of extracts from cranberry, wild blueberry, and strawberry on the main pathogenic determinants of P. gingivalis. Orophenol® attenuated the growth of P. gingivalis and decreased its hemolytic activity, its adherence to a basement membrane matrix model, and its proteinase activities. The berry polyphenolic fraction also impaired the production of reactive oxygen species (ROS) by oral keratinocytes stimulated with P. gingivalis. Lastly, using an in vitro model of oral keratinocyte barrier, the fraction exerted a protective effect against the damages mediated by P. gingivalis. In conclusion, the berry polyphenolic fraction investigated in the present study attenuated several pathogenic properties of P. gingivalis. Although future clinical investigations are required, our study provided evidence that the polyphenols contained in this fraction may represent bioactive molecules of high interest for the prevention and/or treatment of periodontal disease.

A Phenolic-rich Extract of Cocoa (Theobroma cacao L.) Beans Impairs the Pathogenic Properties of Porphyromonas gingivalis and Attenuates the Activation of Nuclear Factor Kappa B in a Monocyte Model.

Periodontitis, an inflammatory disease that affects tooth-supporting tissues, is the result of a polymicrobial infection involving mainly Gram negative anaerobic bacteria. The aim of the present study was to investigate the effects of a phenolic-rich extract of cocoa (Theobroma cacao L.) beans on the pathogenic properties of Porphyromonas gingivalis, which is well-known as a keystone pathogen in the development of periodontitis. The effect of the cocoa extract on P. gingivalis-induced activation of the nuclear factor kappa B (NF-κB) transcription factor in a monocyte model was also assessed. The cocoa extract, whose major phenolic compound was epicatechin, inhibited the growth, hemolytic activity, proteolytic activities, and adherence properties (basement membrane matrix, erythrocytes) of P. gingivalis in a dose-dependent manner. It also protected the barrier function of a keratinocyte model against the deleterious effects mediated by P. gingivalis, and attenuated reactive oxygen species (ROS) production by oral keratinocytes treated with P. gingivalis. Lastly, the cocoa extract showed an anti-inflammatory property by preventing P. gingivalis-induced NF-κB activation in monocytes. In conclusion, this in vitro study highlighted the potential value of an epicatechin-rich extract of cocoa beans for preventing and/or treating periodontal diseases.

Identification and Characterization of a Two-Peptide Class IIb Bacteriocin in Streptococcus pluranimalium Isolated from the Nasal Cavity of a Healthy Pig.

In addition to be an important zoonotic agent, Streptococcus suis serotype 2 causes severe infections in pigs. In this study, we characterized a new bacteriocin produced by Streptococcus pluranimalium 2N12 isolated from a pig nasal sample. The bacteriocin, termed pluranimalicin 2N12, was a two-peptide class IIb bacteriocin active against S. suis. The gene cluster responsible for the biosynthesis of pluranimalicin 2N12 by S. pluranimalium contained seven open reading frames, including putative genes for peptides (pluα, pluß), export (pluA, pluB), and regulation (pluC, pluD, pluE). The deduced amino acid sequences of the peptides Pluα (33 amino acids) and Pluß (29 amino acids) showed 73% and 69% identity in amino acid residues, respectively, with the peptides SthA and SthB of the streptocin produced by Streptococcus gordonii. The antibacterial activity of pluranimalicin 2N12 against S. suis was dependent on the presence of the two peptides Pluα and Pluß that exhibited a membrane permeabilization effect. No activity was found against the other swine pathogens tested. Depending on the concentrations used, Pluα and Pluß displayed no or low toxicity towards swine tracheal epithelial cells. The pluranimalicin peptides Pluα and Pluß, either individually or in combination, exhibited anti-inflammatory activity since they attenuated IL-6 and TNF-α production by macrophages challenged with lipopolysaccharide. Given its dual action (antibacterial and anti-inflammatory), pluranimalicin 2N12 holds promise as a potential therapeutic agent for controlling S. suis infections.

Streptococcus pluranimalium 2N12 Exerts an Antagonistic Effect Against the Swine Pathogen Actinobacillus pleuropneumoniae by Producing Hydrogen Peroxide.

Actinobacillus pleuropneumoniae is the causal agent of porcine pleuropneumonia, a highly contagious and often deadly respiratory disease that causes major economic losses in the swine industry worldwide. The aim of the present study was to investigate the hydrogen peroxide (H2O2)-dependent antagonistic activity of Streptococcus pluranimalium 2N12 (pig nasal isolate) against A. pleuropneumoniae. A fluorimetric assay showed that S. pluranimalium produces H2O2 dose- and time-dependently. The production of H2O2 increased in the presence of exogenous lactate, suggesting the involvement of lactate oxidase. All 20 strains of A. pleuropneumoniae tested, belonging to 18 different serovars, were susceptible to H2O2, with minimal inhibitory concentrations and minimal bactericidal concentrations ranging from 0.57 to 2.3 mM. H2O2, as well as a culture supernatant of S. pluranimalium, killed planktonic cells of A. pleuropneumoniae. Treating the culture supernatant with catalase abolished its bactericidal property. H2O2 was also active against a pre-formed biofilm-like structure of A. pleuropneumoniae albeit to a lesser extent. A checkerboard assay was used to show that there were antibacterial synergistic interactions between H2O2 and conventional antibiotics, more particularly ceftiofur. Based on our results and within the limitations of this in vitro study, the production of H2O2 by S. pluranimalium could be regarded as a potential protective mechanism of the upper respiratory tract against H2O2-sensitive pathogens such as A. pleuropneumoniae.

A green tea extract and epigallocatechin-3-gallate attenuate the deleterious effects of irinotecan in an oral epithelial cell model.

OBJECTIVE: To investigate the ability of a green tea extract and epigallocatechin-3-gallate (EGCG) to protect oral epithelial cells against the deleterious effects of the chemotherapeutic agent irinotecan, with respect to cytotoxicity; reactive oxygen species (ROS) generation; cytokine and matrix metalloproteinase (MMP) production; and cell proliferation and migration. METHODS: The B11 oral keratinocyte and GMSM-K oral epithelial cell lines were used in this study. Cell viability was determined using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) colorimetric assay. A fluorometric assay was used to quantify ROS production. Cell proliferation was assessed using a fluorescent cell tracker dye, while a migration assay kit was used to monitor cell migration. Cytokine and MMP secretion was quantified by an enzyme-linked immunosorbent assay. RESULTS: The green tea extract and EGCG reduced the cytotoxicity of irinotecan toward oral keratinocyte and epithelial cell lines. Irinotecan-induced intracellular ROS generation by oral keratinocytes was reduced by the green tea extract and EGCG. Irinotecan negatively affected the proliferation and migration of oral keratinocytes in a dose-dependent manner. However, these effects were not neutralized by the green tea extract, while EGCG showed a trend to attenuate the irinotecan-induced decrease in cell migration. The green tea extract and EGCG also had a dose-dependent inhibitory effect on irinotecan-induced secretion of interleukin-6 and interleukin-8 by oral epithelial cells. Lastly, the irinotecan-induced decrease in the secretion of MMP-2 and MMP-9 by oral epithelial cells was partially restored by the green tea extract and EGCG. CONCLUSIONS: The green tea extract and EGCG, through anti-cytotoxic, anti-oxidative, and anti-inflammatory properties, may protect the oral mucosa against the deleterious effects of the chemotherapeutic agent irinotecan and may be of interest for treating oral mucositis.

Effects of the Licorice Isoflavans Licoricidin and Glabridin on the Growth, Adherence Properties, and Acid Production of Streptococcus mutans, and Assessment of Their Biocompatibility.

Pharmacological studies have linked a number of human health benefits with licorice due to its anticancer, anti-inflammatory, anti-oxidant, and antimicrobial properties. The aim of this study was to investigate the effects of licoricidin and glabridin, two major licorice isoflavans, on growth and virulence properties (biofilm formation, acid production, dextran production, adherence) of the cariogenic bacterium Streptococcus mutans. Moreover, the biocompatibility of these licorice compounds was assessed in an in vitro model of oral keratinocytes. We used a broth microdilution assay to show that licoricidin and glabridin exhibit a marked antibacterial activity against S. mutans. Glabridin and, to a lesser extent, licoricidin reduced the biofilm viability of S. mutans. In addition, glabridin decreased the production of dextran by S. mutans. The two licorice isoflavans attenuated the adherence of S. mutans to a saliva-coated hydroxylapatite surface, and reduced acid production from glucose. Lastly, depending on the concentrations tested, the two licorice isoflavans showed no or low toxicity toward oral keratinocytes. Within the limitations of this study, our data suggest that licoricidin and glabridin may be promising agents for controlling dental caries.

A polyphenolic cinnamon fraction exhibits anti-inflammatory properties in a monocyte/macrophage model.

Periodontal diseases are bacteria-induced inflammatory disorders that lead to the destruction of the tooth-supporting tissues. Active compounds endowed with a capacity to regulate the inflammatory response are regarded as potential therapeutic agents for the treatment of periodontal diseases. The aim of this study was to characterize the anti-inflammatory properties of a polyphenolic cinnamon fraction. Chromatographic and mass spectrometry analyses of the polyphenolic composition of the cinnamon fraction revealed that phenolic acids, flavonoids (flavonols, anthocyanins, flavan-3-ols), and procyanidins make up 9.22%, 0.72%, and 10.63% of the cinnamon fraction, respectively. We used a macrophage model stimulated with lipopolysaccharides (LPS) from either Aggregatibacter actinomycetemcomitans or Escherichia coli to show that the cinnamon fraction dose-dependently reduced IL-6, IL-8, and TNF-α secretion. Evidence was brought that this inhibition of cytokine secretion may result from the ability of the fraction to prevent LPS-induced NF-κB activation. We also showed that the cinnamon fraction reduces LPS binding to monocytes, which may contribute to its anti-inflammatory properties. Lastly, using a competitor assay, it was found that the cinnamon fraction may represent a natural PPAR-γ ligand. Within the limitations of this in vitro study, the cinnamon fraction was shown to exhibit a therapeutic potential for the treatment of periodontal diseases due to its anti-inflammatory properties.

Effects of a tart cherry (Prunus cerasus L.) phenolic extract on Porphyromonas gingivalis and its ability to impair the oral epithelial barrier.

Periodontal diseases, including gingivitis and periodontitis, are a global oral health problem. Porphyromonas gingivalis, a key pathogen involved in the onset of periodontitis, is able to colonize the subgingival epithelium and invade the underlying connective tissue due to the contribution of cysteine proteases known as gingipains. In this study, we investigated the effects of a phenolic extract prepared from tart cherry (Prunus cerasus L.) juice on the growth, adherence, and protease activity of P. gingivalis. We also assessed the protective effect of the tart cherry extract on the disruption of the oral epithelial barrier induced by P. gingivalis. The tart cherry extract that contains procyanidins and quercetin and its derivatives (rutinoside, glucoside) as the most important phenolic compounds attenuated P. gingivalis growth, reduced adherence to an experimental basement membrane matrix model, and decreased the protease activities of P. gingivalis. The tart cherry extract also exerted a protective effect on the integrity of the oral epithelial barrier in an in vitro model infected with P. gingivalis. More specifically, the extract prevented a decrease in transepithelial electrical resistance as well as the destruction of tight junction proteins (zonula occludens-1 and occludin). These results suggest that the tart cherry phenolic extract may be a promising natural product for the treatment of periodontitis through its ability to attenuate the virulence properties of P. gingivalis and curtail the ability of this pathogen to impair the oral epithelial barrier.

Effects of Labrador Tea, Peppermint, and Winter Savory Essential Oils on Fusobacterium nucleatum.

Bad breath or halitosis is an oral condition caused by volatile sulfur compounds (VSC) produced by bacteria found in the dental and tongue biofilms. Fusobacterium nucleatum is a Gram-negative anaerobic bacterium that has been strongly associated with halitosis. In this study, essential oils (EO) from three plants, Labrador tea (Rhododendron groenlandicum [Oeder] Kron & Judd), peppermint (Mentha x piperita L.), and winter savory (Satureja montana L.), were investigated for their effects on growth, biofilm formation and killing, and VSC production by F. nucleatum. Moreover, their biocompatibility with oral keratinocytes was investigated. Using a broth microdilution assay, winter savory EO and to a lesser extent Labrador tea and peppermint EO showed antibacterial activity against F. nucleatum. A treatment of pre-formed biofilms of F. nucleatum with EO also significantly decreased bacterial viability as determined by a luminescence assay monitoring adenosine triphosphate production. The EO were found to permeabilize the bacterial cell membrane, suggesting that it represents the target of the tested EO. The three EO under investigation were able to dose-dependently reduce VSC production by F. nucleatum. Lastly, no significant loss of cell viability was observed when oral keratinocytes were treated with the EO at concentrations effective against F. nucleatum. This study supports the potential of Labrador tea, peppermint, and winter savory EO as promising agents to control halitosis and promote oral health.

Antimicrobial Activity, Biocompatibility and Anti-inflammatory Properties of Cetylpyridinium Chloride-based Mouthwash Containing Sodium Fluoride and Xylitol: An In Vitro Study.

PURPOSE: The use of a mouthwash as an adjunct to mechanical plaque removal may be useful to improve oral hygiene. In this study, cetylpyridinium chloride (CPC)-based mouthwashes containing sodium fluoride and xylitol (X-PUR Opti-Rinse 0.05% NaF and X-PUR Opti-Rinse 0.2% NaF) were evaluated for their antimicrobial activity against important oral pathogens associated with dental caries, periodontal disease, and candidiasis. Moreover, their biocompatibility and anti-inflammatory properties were assessed.
Materials and Methods: Antimicrobial activity was determined using a disk-diffusion assay, a microplate dilution assay, and the European standard protocols for antiseptics. Microbicidal properties were assessed against both planktonic and biofilm cultures. An oral epithelial cell model was used to evaluate the biocompatibility of mouthwashes and their ability to attenuate cytokine secretion.
Results: Using three different antimicrobial assays, the CPC-based mouthwashes were found to be highly active against the tested microorganisms. More specifically, the mouthwashes met the European Standard NF EN 1040 and NF EN 1275 defined as a log10 reduction ≥ 5 (≥ 99.999% killing) for bacteria and log10 reduction ≥ 4 (≥ 99.99% killing) for fungi, respectively. The CPC-based mouthwashes were also bactericidal against biofilms of S. mutans, S. sobrinus, and P. gingivalis. Using an oral epithelial cell model, the CPC-based mouthwashes were found to be less cytotoxic than a chlorhexidine-containing mouthwash used as control. Lastly, the CPC-based mouthwashes decreased the secretion of interleukin-6 and interleukin-8 by lipopolysaccharide-stimulated oral epithelial cells.
Conclusion: The CPC-based mouthwashes supplemented with sodium fluoride (0.05% or 0.2%) and xylitol (10%) were highly active against important oral pathogens. Moreover, using an oral epithelial cell model, these mouthwashes were found to be biocompatible and to exhibit anti-inflammatory activity.

The bacteriocin from the prophylactic candidate Streptococcus suis 90-1330 is widely distributed across S. suis isolates and appears encoded in an integrative and conjugative element.

The Gram-positive α-hemolytic Streptococcus suis is a major pathogen in the swine industry and an emerging zoonotic agent that can cause several systemic issues in both pigs and humans. A total of 35 S. suis serotypes (SS) have been identified and genotyped into > 700 sequence types (ST) by multilocus sequence typing (MLST). Eurasian ST1 isolates are the most virulent of all S. suis SS2 strains while North American ST25 and ST28 strains display moderate to low/no virulence phenotypes, respectively. Notably, S. suis 90-1330 is an avirulent Canadian SS2-ST28 isolate producing a lantibiotic bacteriocin with potential prophylactic applications. To investigate the suitability of this strain for such purposes, we sequenced its complete genome using the Illumina and PacBio platforms. The S. suis 90-1330 bacteriocin was found encoded in a locus cargoed in what appears to be an integrative and conjugative element (ICE). This bacteriocin locus was also found to be widely distributed across several streptococcal species and in a few Staphylococcus aureus strains. Because the locus also confers protection from the bacteriocin, the potential prophylactic benefits of using this strain may prove limited due to the spread of the resistance to its effects. Furthermore, the S. suis 90-1330 genome was found to code for genes involved in blood survival, suggesting that strain may not be a benign as previously thought.

Antibacterial activity against porcine respiratory bacterial pathogens and in vitro biocompatibility of essential oils.

Bacterial respiratory infections affecting pigs such as pneumonia, pleuropneumonia, and pleurisy, are a major health concern in the swine industry and are associated with important economic losses. This study aimed to investigate the antibacterial activities of essential oils against major swine respiratory pathogens with a view to developing a potential alternative to antibiotics. Their synergistic interactions with the bacteriocin nisin was also examined. Lastly, we assessed the in vitro biocompatibility of the most efficient essential oils using a pig tracheal epithelial cell line. Of the nine essential oils tested, those from cinnamon, thyme, and winter savory were the most active against Streptococcus suis, Actinobacillus pleuropneumoniae, Actinobacillus suis, Bordetella bronchiseptica, Haemophilus parasuis, and Pasteurella multocida, with minimum inhibitory concentrations and minimum bactericidal concentrations ranging from 0.01 to 0.156% (v/v). The main component found in cinnamon, thyme, and winter savory oils were cinnamaldehyde, thymol, and carvacrol, respectively. Treating pre-formed S. suis and A. pleuropneumoniae biofilms with thyme or winter savory oils significantly decreased biofilm viability. We also observed a synergistic growth inhibition of S. suis with mixtures of nisin and essential oils from thyme and winter savory. Concentrations of nisin and cinnamon, thyme and winter savory essential oils that were effective against bacterial pathogens had no effect on the viability of pig tracheal epithelial cells. The present study brought evidence that essential oils are potential antimicrobial agents against bacteria associated with porcine respiratory infections.

Lobaric acid and pseudodepsidones inhibit NF-κB signaling pathway by activation of PPAR-γ.

Herein we report the anti-inflammatory activity of lobaric acid and pseudodepsidones isolated from the nordic lichen Stereocaulon paschale. Lobaric acid (1) and three compounds (2, 7 and 9) were found to inhibit the NF-κB activation and the secretion of pro-inflammatory cytokines (IL-1ß and TNF-α) in LPS-stimulated macrophages. Inhibition and docking simulation experiments provided evidence that lobaric acid and pseudodepsidones bind to PPAR-γ between helix H3 and the beta sheet, similarly to partial PPAR-γ agonists. These findings suggest that lobaric acid and pseudodepsidones reduce the expression of pro-inflammatory cytokines by blocking the NF-κB pathway via the activation of PPAR-γ.

Dipeptidylpeptidase IV of Streptococcus suis degrades the porcine antimicrobial peptide PR-39 and neutralizes its biological properties.

Streptococcus suis is a major swine pathogen causing pathologies such as meningitis, sepsis, endocarditis, and arthritis. Several surface-bound and secreted proteases produced by S. suis have been identified and proposed as virulence factors. PR-39 is a proline/arginine-rich antimicrobial peptide produced by porcine leucocytes. In addition to play a role in innate immunity, this peptide possesses immunomodulatory properties. In this study, we hypothesized that proteases produced by S. suis inactivate PR-39. Most strains of S. suis tested were relatively resistant to PR-39, with minimal inhibitory concentration (MIC) values ≥ 200 µg/ml. The proteolytic cleavage of PR-39 by recombinant subtilisin-like protease and dipeptidylpeptidase IV (DPPIV) of S. suis was assessed by SDS-PAGE. While PR-39 was not cleaved by the subtilisin-like protease, it was time-dependently degraded by DPPIV. Whole cells of S. suis also degraded PR-39. When S. suis was grown in a culture medium supplemented with recombinant DPPIV, its susceptibility to PR-39 was decreased. Activation of brain microvascular endothelial cells with PR-39 resulted in an increased secretion of the chemokine interleukin-8 (IL-8) thus confirming the immunomodulatory activity of this porcine antimicrobial peptide. However, a pre-treatment of PR-39 with DPPIV completely neutralized the increased IL-8 secretion. In this study, we showed that DPPIV produced by S. suis can degrade PR-39 and neutralize its antibacterial and immunomodulatory properties. This may allow survival of S. suis in the central nervous system by resisting to killing by this antimicrobial peptide and delaying the recruitment of phagocytic cells such as neutrophils to the site of infection.

In vitro antibacterial activity of plant essential oils against Staphylococcus hyicus and Staphylococcus aureus, the causative agents of exudative epidermitis in pigs.

Greasy pig disease or exudative epidermitis, a generalized or localized skin disease affecting piglets, is mainly caused by Staphylococcus hyicus, although other staphylococcal species such as Staphylococcus aureus may also induce disease. Piglets with skin lesions can be treated systemically with antibiotics. However, antimicrobial resistance to ß-lactam antibiotics are now frequently observed in S. hyicus and S. aureus isolates. In this study, the antibacterial activity of plant essential oils as well as their ability to potentiate the effect of several antimicrobial compounds against S. hyicus and S. aureus were investigated with a view to a potential use as skin disinfectants. Among ten essential oils tested, those from cinnamon, thyme, and winter savory were the most active with minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) values ranging from 0.078 to 0.313% (v/v). Using a fluorescent probe with DNA affinity, it was found that thyme and winter savory oils act, at least in part, by disturbing the bacterial membrane integrity. At concentrations below the MIC, thyme and winter savory oils reduced biofilm formation by S. hyicus. Moreover, a treatment of pre-formed biofilms of S. hyicus with cinnamon or thyme oils significantly decreases its viability. Synergistic interactions between essential oils, more particularly from thyme and winter savory, and penicillin G, chlorhexidine or nisin, were observed. This study supports the therapeutic potential of essential oils as topical therapeutic agents against exudative epidermitis.

Distribution of Suicin Gene Clusters in Streptococcus suis Serotype 2 Belonging to Sequence Types 25 and 28.

Recently, we reported the purification and characterization of three distinct lantibiotics (named suicin 90-1330, suicin 3908, and suicin 65) produced by Streptococcus suis. In this study, we investigated the distribution of the three suicin lantibiotic gene clusters among serotype 2 S. suis strains belonging to sequence type (ST) 25 and ST28, the two dominant STs identified in North America. The genomes of 102 strains were interrogated for the presence of suicin gene clusters encoding suicins 90-1330, 3908, and 65. The gene cluster encoding suicin 65 was the most prevalent and mainly found among ST25 strains. In contrast, none of the genes related to suicin 90-1330 production were identified in 51 ST25 strains nor in 35/51 ST28 strains. However, the complete suicin 90-1330 gene cluster was found in ten ST28 strains, although some genes in the cluster were truncated in three of these isolates. The vast majority (101/102) of S. suis strains did not possess any of the genes encoding suicin 3908. In conclusion, this study indicates heterogeneous distribution of suicin genes in S. suis.

Purification and Characterization of Suicin 65, a Novel Class I Type B Lantibiotic Produced by Streptococcus suis.

Bacteriocins are antimicrobial peptides of bacterial origin that are considered as a promising alternative to the use of conventional antibiotics. Recently, our laboratory reported the purification and characterization of two lantibiotics, suicin 90-1330 and suicin 3908, produced by the swine pathogen and zoonotic agent Streptococcus suis (serotype 2). In this study, a novel bacteriocin produced by S. suis has been identified and characterized. The producing strain S. suis 65 (serotype 2) was found to belong to the sequence type 28, that includes strains known to be weakly or avirulent in a mouse model. The bacteriocin, whose production was only possible following growth on solid culture medium, was purified to homogeneity by cationic exchange and reversed-phase high-pressure liquid chromatography. The bacteriocin, named suicin 65, was heat, pH and protease resistant. Suicin 65 was active against all S. suis isolates tested, including antibiotic resistant strains. Amino acid sequencing of the purified bacteriocin by Edman degradation revealed the presence of modified amino acids suggesting a lantibiotic. Using the partial sequence obtained, a blast was performed against published genomes of S. suis and allowed to identify a putative lantibiotic locus in the genome of S. suis 89-1591. From this genome, primers were designed and the gene cluster involved in the production of suicin 65 by S. suis 65 was amplified by PCR. Sequence analysis revealed the presence of ten open reading frames, including a duplicate of the structural gene. The structural genes (sssA and sssA') of suicin 65 encodes a 25-amino acid residue leader peptide and a 26-amino acid residue mature peptide yielding an active bacteriocin with a deducted molecular mass of 3,005 Da. Mature suicin 65 showed a high degree of identity with class I type B lantibiotics (globular structure) produced by Streptococcus pyogenes (streptococcin FF22; 84.6%), Streptococcus macedonicus (macedocin ACA-DC 198; 84.6%), and Lactococcus lactis subsp. lactis (lacticin 481; 74.1%). Further studies will evaluate the ability of suicin 65 or the producing strain to prevent experimental S. suis infections in pigs.

Hyaluronate lyase activity of Streptococcus suis serotype 2 and modulatory effects of hyaluronic acid on the bacterium's virulence properties.

BACKGROUND: Streptococcus suis serotype 2 is a major swine pathogen and zoonotic agent worldwide causing mainly meningitis and septicemia. Hyaluronate lyases are enzymes that degrade hyaluronic acid, a major constituent of animal tissues, and have been reported as virulence factors in various bacterial species. Since the hyaluronate lyase of S. suis has been considered ambiguously as a virulence factor, we screened 50 isolates from the three major clonal complexes found in North America (sequence type [ST] 1, ST25, and ST28) known to differ in their degree of virulence in order to link the presence or absence of this activity with the degree of virulence. Moreover, the effect of exogenous hyaluronic acid on S. suis virulence factor gene expression and the pro-inflammatory response of brain macrovascular endothelial cells (BMEC) was also investigated. RESULTS: We found that all but one ST1 isolates (high virulence) were devoid of hyaluronate lyase activity whereas all ST25 (intermediate virulence) and ST28 (low virulence) isolates possessed the activity. A 2 bp insertion was responsible for the lack of activity in ST1 strains. Since the most virulent isolates did not degrade hyaluronic acid, this tissue component may be found during the infectious process. Therefore, we investigated its effect on S. suis and host cells. Hyaluronic acid was found to modulate S. suis adhesion to BMEC, to increase S. suis virulence factor expression, and to enhance pro-inflammatory cytokine secretion by BMEC. CONCLUSIONS: These findings suggest that S. suis hyaluronate lyase does not represent a critical virulence factor in its active form. However, exogenous hyaluronic acid that is likely to interact with S. suis and host cells during the course of infection appears to modulate several virulence determinants of the bacterium, in addition to promote inflammation.

Suicin 3908, a new lantibiotic produced by a strain of Streptococcus suis serotype 2 isolated from a healthy carrier pig.

While Streptococcus suis serotype 2 is known to cause severe infections in pigs, it can also be isolated from the tonsils of healthy animals that do not develop infections. We hypothesized that S. suis strains in healthy carrier pigs may have the ability to produce bacteriocins, which may contribute to preventing infections by pathogenic S. suis strains. Two of ten S. suis serotype 2 strains isolated from healthy carrier pigs exhibited antibacterial activity against pathogenic S. suis isolates. The bacteriocin produced by S. suis 3908 was purified to homogeneity using a three-step procedure: ammonium sulfate precipitation, cationic exchange HPLC, and reversed-phase HPLC. The bacteriocin, called suicin 3908, had a low molecular mass; was resistant to heat, pH, and protease treatments; and possessed membrane permeabilization activity. Additive effects were obtained when suicin 3908 was used in combination with penicillin G or amoxicillin. The amino acid sequence of suicin 3908 suggested that it is lantibiotic-related and made it possible to identify a bacteriocin locus in the genome of S. suis D12. The putative gene cluster involved in suicin production by S. suis 3908 was amplified by PCR, and the sequence analysis revealed the presence of nine open reading frames (ORFs), including the structural gene and those required for the modification of amino acids, export, regulation, and immunity. Suicin 3908, which is encoded by the suiA gene, exhibited approximately 50% identity with bovicin HJ50 (Streptococcus bovis), thermophilin 1277 (Streptococcus thermophilus), and macedovicin (Streptococcus macedonicus). Given that S. suis 3908 cannot cause infections in animal models, that it is susceptible to conventional antibiotics, and that it produces a bacteriocin with antibacterial activity against all pathogenic S. suis strains tested, it could potentially be used to prevent infections and to reduce antibiotic use by the swine industry.

Characterization of DNase activity and gene in Streptococcus suis and evidence for a role as virulence factor.

BACKGROUND: The Gram-positive bacterium Streptococcus suis serotype 2 is an important swine pathogen and emerging zoonotic agent. Multilocus sequence typing allowed dividing S. suis serotype 2 into sequence types (STs). The three major STs of S. suis serotype 2 from North America are 1 (most virulent), 25 (intermediate virulence) and 28 (less virulent). Although the presence of DNase activity in S. suis has been previously reported, little data is available. The aim of this study was to investigate DNase activity in S. suis according to STs, to characterize the activity and gene, and to provide evidence for a potential role in virulence. RESULTS: We showed that ST1 and ST28 strains exhibited DNase activity that was absent in ST25 strains. The lack of activity in ST25 isolates was associated with a 14-bp deletion resulting in a shifted reading frame and a premature stop codon. The DNase of S. suis P1/7 (ST1) was cell-associated and active on linear DNA. A DNase-deficient mutant of S. suis P1/7 was found to be less virulent in an amoeba model. Stimulation of macrophages with the DNase mutant showed a decreased secretion of pro-inflammatory cytokines and matrix metalloproteinase-9 compared to the parental strain. CONCLUSIONS: This study further expands our knowledge of S. suis DNase and its potential role in virulence.