Effects of Diluents, Saliva and Other Organics on the Microbicidal Activity of Cetylpyridinium Chloride and Povidone-iodine.

Povidone-iodine (PVP-I) is used for infection control and preoperative sterilization of the oral and pharyngeal regions. Marketed preparations containing cetylpyridinium chloride (CPC) are used to inhibit growth of oral bacteria. We conducted an in vitro study of the sterilizing effects of these microbicides on 10 oral bacterial strains and fungi related to pneumonia and periodontal disease, after dilution with phosphate-buffered saline (PBS), saliva, and components in saliva. The CPC solution was evaluated at 50 mg/100 mL, which is the concentration used in products. CPC sterilized all strains within 1 minute. Prolongation of the sterilization time associated with dilution was more gradual in comparison to PVP-I solution. CPC sterilized 7 of 10 microbial strains within 3 minutes at 3 mg/100 mL. At 500 mg/100 mL, which is near the upper limit of the concentration that is actually used, PVP-I solution sterilized 7 microbial strains within 3 minutes. However, PVP-I had no sterilization effect when diluted to 100 mg/100 mL or lower. With addition of saliva, PVP-I sterilized 2 microbial strains within 3 minutes at 500 mg/100 mL, whereas CPC solution sterilized 9 microbial strains within 1 minute at 50 mg/100 mL. Our results show that in use influenced by dilution with saliva, CPC is likely to maintain a strong sterilization effect, whereas PVP-I may have a reduced effect.

Comparative inhibitory effects of 4-allylpyrocatechol isolated from Piper betle on Streptococcus intermedius, Streptococcus mutans, and Candida albicans.

Streptococcus intermedius, Streptococcus mutans, and Candida albicans are harmful oral pathogens and prone to resist chemical antimicrobial agents. Active ingredients from plants are of increasing interest as an alternative. This study aims to compare antimicrobial effects of 4-allylpyrocatechol (APC) extracted from Piper betle on these oral pathogens. Minimum concentration of APC against the tested pathogens was determined using a broth microdilution assay. Killing kinetic study of APC was carried out within 24 h. Morphology of the pathogenic cells was assessed using scanning electron microscopy (SEM). Anti-biofilm was investigated using crystal violet assay and confocal laser scanning microscopy (CLSM). The results showed that the mechanism of inhibition of APC was bactericidal and fungicidal effects. APC at minimum concentration of 400 µg/mL could completely kill Streptococcus and Candida spp., however, the killing rate on S. intermedius and C. albicans was significantly faster than on S. mutans. APC inhibited biofilm formation of C. albicans more efficiently than that of the bacterial cells. Cell morphology from SEM indicated that APC caused bacterial cell membrane destruction and inhibited fungal budding or tubing formation. CLSM images confirmed the killing potential of APC and suggested that bacterial dead cells could be easier washed out than the fungal dead cells. It is concluded that APC potentially inhibits growth and biofilms of oral Streptococcus and Candida spp. in different mechanism of action and killing rate. APC can be considered as a promising agent for preventing and treating dental disorders caused by S. intermedius, S. mutans, and C. albicans.

Effects of Piper betle fractionated extracts on inhibition of Streptococcus mutans and Streptococcus intermedius.

The overgrowth of certain strains of normal flora in oral cavity can cause many kinds of oral infections or diseases such as carries, periodontitis, and gingivitis. Prevention and treatment of these diseases are usually achieved by chemical antiseptics. However, these chemicals are found as negative impacts of human health hazards and accession of microbial resistance. The present study explores the potential of Piper betle extracts on inhibition of two oral pathogenic bacteria; Streptococcus mutans DMST 41283 and Streptococcus intermedius DMST 42700. P. betle demonstrated significantly higher inhibitory activity against both pathogenic strains than Acacia catechu, Camellia sinensis, Coccinia grandis, Solanum indicum, and Streblus asper. Among fractionated extracts of P. betle from several solvents, the extract from ethyl acetate (Pb-EtOAc) possessed the widest inhibition zone of 11.0 ± 0.1 and 11.3 ± 0.4 mm against both bacterial strains, respectively. Pb-EtOAc showed the same minimum inhibitory concentration of 0.5 mg/mL against both strains, whereas its minimum bactericidal concentrations were 2.0 and 0.5 mg/mL against S. mutans and S. intermedius, respectively. HPLC analysis demonstrated that the major active compound of Pb-EtOAc was 4-allylpyrocatechol. It was found that the killing kinetics of Pb-EtOAc against both test strains were time and dose dependent. Scanning electron microscopy micrographs showed the morphological changes and depletion of the tested pathogens indicating cell destruction after exposure to Pb-EtOAc. It is confirmed that Pb-EtOAc is potentially effective against both oral pathogens and might be used as natural alternative agents in prevention and treatment of oral infections caused by oral pathogenic bacteria.

Antibacterial Efficacy and Discoloration Potential of Endodontic Topical Antibiotics.

INTRODUCTION: The optimal concentration for the use of endodontic topical antibiotics is not known. The aims of this study were to determine the minimum bactericidal concentrations (MBCs) and minimum inhibitory concentrations (MICs) of metronidazole, ciprofloxacin, minocycline, Augmentin (GlaxoSmithKline, Research Triangle Park, NC), and tigecycline against common endodontic pathogens and to evaluate ex vivo the antibacterial efficacy and discoloration effect of triple antibiotic paste (TAP), Augmentin, and tigecycline at different concentrations using a slow-release hydrogel scaffold. METHODS: Using the Epsilometer test method (Etest; bioMérieux USA, St Louis, MO), MICs and MBCs of selected antibiotics were determined against Fusobacterium nucleatum, Porphyromonas gingivalis, Streptococcus intermedius, and Enterococcus faecalis. Biofilms of these bacterial species were then grown in extracted single-rooted teeth anaerobically for 3 weeks. Root canals were filled with TAP, Augmentin, and tigecycline at concentrations of 1 or 0.1 mg/mL in a degradable hydrogel scaffold or pure TAP at 1 g/mL for 7 days. Coronal discoloration was evaluated spectrophotometrically at 1, 2, and 3 weeks after dressing. RESULTS: MIC/MBC data showed significant efficacy of tigecycline, Augmentin, and minocycline compared with the other antibiotics (P < .05). Significant differences were found when comparing the log10 colony-forming units of all experimental groups (P < .05). TAP at 1 g/mL had no bacterial growth but caused the greatest discoloration. Hydrogel mixtures with TAP, Augmentin, or tigecycline at 1 mg/mL significantly reduced bacterial growth and the number of positive samples compared with those at 0.1 mg/mL (P < .05) with minimal discoloration. CONCLUSIONS: TAP, Augmentin, and tigecycline in a hydrogel at 1 mg/mL reduced bacterial growth significantly with minimal color change.

Sites of infection associated with Streptococcus anginosus group among children.

Streptococcus anginosus group (SAG) are parts of normal flora of the oral cavity and associated with abscess forming in various sites on the body. Although the clinical features of infections caused by each member of the SAG in adults has been reported, it has not well been known in children. The aim of this study was to clarify the site of infections associated with individual SAG species among children. Medical records from March 2010 to July 2016 were reviewed at Tokyo Metropolitan Children's Medical Center. Any SAG species (S. anginosus, S. constellatus, or S. intermedius) isolated from clinical samples and recorded in the microbiological database were included for analysis. Analysis of 52 infectious episodes found that S. anginosus was most frequently isolated from the genitourinary tract, and 73% of genitourinary tract infection was balanoposthitis. All genitourinary tract infections were associated with S. anginosus. These findings were different from those of a previous study of adults. Of all the patients, 45 patients (87%) had polymicrobial infections. More than 70% of patients infected by S. anginosus and S. constellatus were co-infected by obligate anaerobes, in comparison with only 21% of S. intermedius cases. Among the obligate anaerobes species, Bacteroides spp. was significantly accompanied with S. anginosus. Susceptibility to penicillin, ampicillin, cefotaxime, erythromycin, clindamycin, levofloxacin, and vancomycin was 100%, 100%, 100%, 77%, 89%, 97% and 100%, respectively. S. anginosus was often isolated from balanoposthitis among children.

Positive- and Negative-Control Pathways by Blood Components for Intermedilysin Production in Streptococcus intermedius.

Streptococcus intermedius is an opportunistic bacterial pathogen secreting a human-specific cytolysin called intermedilysin (ILY) as a major pathogenic factor. This bacterium can degrade glycans into monosaccharides using two glycosidases, multisubstrate glycosidase A (MsgA) and neuraminidase (NanA). Here, we detected a stronger hemolytic activity mediated by ILY when S. intermedius PC574 was cultured in fetal bovine serum (FBS) than when it was grown in the standard culture medium. FBS-cultured cells also showed higher MsgA and NanA activity, although overproduction of ILY in FBS was undetectable in mutants nanA-null and msgA-null. Addition of purified MsgA and NanA to the FBS resulted in a release of 2.8 mM galactose and 4.3 mM N-acetylneuraminic acid; these sugar concentrations were sufficient to upregulate the expression of ILY, MsgA, and NanA. Conversely, when strain PC574 was cultured in human plasma, no similar increase in hemolytic activity was observed. Moreover, addition of human plasma to the culture in FBS appeared to inhibit the stimulatory effect of FBS on ILY, MsgA, and NanA, although there were individual differences among the plasma samples. We confirmed that human plasma contains immunoglobulins that can neutralize ILY, MsgA, and NanA activities. In addition, human plasma had a neutralizing effect on cytotoxicity of S. intermedius toward HepG2 cells in FBS, and a higher concentration of human plasma was necessary to reduce the cytotoxicity of an ILY-high-producing strain than an ILY-low-producing strain. Overall, our data show that blood contains factors that stimulate and inhibit ILY expression and activity, which may affect pathogenicity of S. intermedius.

Effects of Caesalpinia sappan on pathogenic bacteria causing dental caries and gingivitis.

The present study explores antimicrobial activities of Caesalpinia sappan extracts against three strains of oral pathogenic bacteria; Streptococcus mutans DMST9567 (Smu9), Streptococcus mutans DMST41283 (Smu4), and Streptococcus intermedius DMST42700 (Si). Ethanol crude extract of C. sappan (Cs-EtOH) was firstly compared to that of other medicinal plants using disc diffusion method. Cs-EtOH showed significantly higher effective inhibition against all tested strains than other extracts and 0.12% chlorhexidine with the inhibition zone of 17.5 ± 0.5, 18.5 ± 0.0, and 17.0 ± 0.0 mm against Smu9, Smu4, and Si, respectively. Three fractionated extracts of C. sappan using hexane, ethyl acetate, and ethanol, respectively, were further investigated. The fractionated extract from ethanol (F-EtOH) presented the strongest activities with the minimum bactericidal concentration (MBC) of 125-250 µg/mL. Killing kinetics of F-EtOH was depended on the bacterial species and the concentration of F-EtOH. Two-fold MBC of F-EtOH could kill all tested strains within 12 h whereas its 4-fold MBC showed killing effect against Si within 6 h. Separation of F-EtOH by column chromatography using chloroform/methanol mixture as an eluent yielded 11 fractions (F1-F11). The fingerprints of these fractions by high-performance liquid chromatography at 280 nm revealed that F-EtOH consisted of at least 5 compounds. F6 possessed the significantly highest antimicrobial activity among 11 fractions, however less than F-EtOH. It is considered that F-EtOH is the promising extract of C. sappan for inhibiting oral pathogenic bacteria and appropriate as natural antiseptic for further develop of oral hygiene products.

Identification of Anion Channels Responsible for Fluoride Resistance in Oral Streptococci.

Recently, it has been reported that eriC and crcB are involved in bacterial fluoride resistance. However, the fluoride-resistance mechanism in oral streptococci remains unclear. BLAST studies showed that two types of eriCs (eriC1 and eriC2) and two types of crcBs (crcB1 and crcB2) are present across 18 oral streptococci, which were identified in ≥ 10% of 166 orally healthy subjects with ≥ 0.01% of the mean relative abundance. They were divided into three groups based on the distribution of these four genes: group I, only eriC1; group II, eriC1 and eriC2; and group III, eriC2, crcB1, and crcB2. Group I consisted of Streptococcus mutans, in which one of the two eriC1s predominantly affected fluoride resistance. Group II consisted of eight species, and eriC1 was responsible for fluoride resistance, but eriC2 was not, in Streptococcus anginosus as a representative species. Group III consisted of nine species, and both crcB1 and crcB2 were crucial for fluoride resistance, but eriC2 was not, in Streptococcus sanguinis as a representative species. Based on these results, either EriC1 or CrcBs play a role in fluoride resistance in oral streptococci. Complementation between S. mutans EriC1 and S. sanguinis CrcB1/CrcB2 was confirmed in both S. mutans and S. sanguinis. However, neither transfer of S. sanguinis CrcB1/CrcB2 into wild-type S. mutans nor S. mutans EriC1 into wild-type S. sanguinis increased the fluoride resistance of the wild-type strain. Co-existence of different F- channels (EriC and CrcB) did not cause the additive effect on fluoride resistance in oral Streptococcus species.

Hydroalcoholic extract of Carum carvi L. in oral mucositis: a clinical trial in male golden hamsters.

OBJECTIVES: Several studies have attempted to prevent or improve oral mucositis (OM) but have not produced a qualified treatment yet. This study evaluates the effects of Carum carvi L. (caraway) hydroalcoholic extract (CHE) as one of the traditional medicinal plants in 5-fluorouracil (5-FU)-induced OM in golden hamsters. MATERIALS AND METHODS: OM was induced in 54 male golden hamsters by 5-FU and cheek pouch scratching. Starting from day 12, 500 and 1000 mg kg(-1) per day topical CHE were administered. Pouch histopathology score, malondialdehyde and reduced glutathione contents, and activity of myeloperoxidase plus microbial cultures of cheek pouch, antimicrobial properties of CHE, and essential oil constituents were evaluated. RESULTS: Lower histopathology score (0, 1, and 2) and malondialdehyde level, higher reduced glutathione level and activities of myeloperoxidase were detected in 1000 and 500 mg kg(-1) per day topical CHE and control groups, respectively (P < 0.001). The CHE was more potent against Staphylococcus epidermidis and Streptococcus intermedius. γ-Terpinene (37.2%) was identified as the main constituent of essential oil. CONCLUSION: The use of CHE in topical form may be associated with reduced intensity of OM. This may be due to appropriate antibacterial activity and terpinene contents.

Antibiotic susceptibility of periodontal Streptococcus constellatus and Streptococcus intermedius clinical isolates.

BACKGROUND: Streptococcus constellatus and Streptococcus intermedius in subgingival dental plaque biofilms may contribute to forms of periodontitis that resist treatment with conventional mechanical root debridement/surgical procedures and may additionally participate in some extraoral infections. Because systemic antibiotics are often used in these clinical situations, and little is known of the antibiotic susceptibility of subgingival isolates of these two bacterial species, this study determined the in vitro susceptibility to six antibiotics of fresh S. constellatus and S. intermedius clinical isolates from human periodontitis lesions. METHODS: A total of 33 S. constellatus and 17 S. intermedius subgingival strains, each recovered from separate patients with severe chronic periodontitis (n = 50) before treatment, were subjected to antibiotic gradient strip susceptibility testing with amoxicillin, azithromycin, clindamycin, ciprofloxacin, and doxycycline on blood-supplemented Mueller-Hinton agar and to the inhibitory effects of metronidazole at 16 mg/L in an enriched Brucella blood agar dilution assay. Clinical and Laboratory Standards Institute and European Committee on Antimicrobial Susceptibility Testing interpretative standards were used to assess the results. RESULTS: Clindamycin was the most active antibiotic against S. constellatus (minimum inhibitory concentration at 90% [MIC90] 0.25 mg/L), and amoxicillin was most active against S. intermedius (MIC90 0.125 mg/L). A total of 30% of the S. constellatus and S. intermedius clinical isolates were resistant in vitro to doxycycline, 98% were only intermediate in susceptibility to ciprofloxacin, and 90% were resistant to metronidazole at 16 mg/L. CONCLUSION: Subgingival S. constellatus and S. intermedius exhibited variable antibiotic susceptibility profiles, potentially complicating empirical selection of periodontitis antibiotic therapy in patients who are species positive.

Effective oral health in infective endocarditis: efficacy of high-street mouthwashes against the viridans group streptococci.

AIM: Recent UK National Institute for Health and Clinical Excellence guidelines state that there is no longer a need for oral antibiotic prophylaxis in patients undergoing dental procedures who are at risk of infective endocarditis (IE), and advocate the importance of maintaining good oral health. As viridans group streptococci (VGS) are common etiological agents of IE and inhabitants of the mouth, the purpose of this study was to examine the efficacy of common high-street mouthwashes against four classes of VGS organisms (salivarius, mitis, anginosus, and mutans groupings). METHODS: The survival of VGS, Streptococcus gordonii (National Collection of Type Cultures [NCTC] 7865), Streptococcus intermedius (NCTC 11324), Streptococcus mutans (NCTC 10449), Streptococcus oralis (NCTC 11427), Streptococcus pneumoniae (NCTC 7465, NCTC 7978, & American Type Culture Collection 49619) and Streptococcus salivarius (NCTC 8618) was assessed in vitro following treatment of approximately 10(7) c.f.u. in planktonic state with four mouthwashes. RESULTS: No organisms were culturable following 1-min exposure, and were not recovered following non-selective enrichment following incubation in Brain Heart Infusion broth supplemented with 0.8% (w/v) yeast extract. CONCLUSIONS: These data indicate that such mouthwashes are able to completely kill VGS organisms tested in planktonic solution, where their use would promote good oral hygiene in patients at risk of IE.

Effects of extracellular DNA and DNA-binding protein on the development of a Streptococcus intermedius biofilm.

AIMS: The aim of this study was to clarify the effects of homologous and heterologous extracellular DNAs (eDNAs) and histone-like DNA-binding protein (HLP) on Streptococcus intermedius biofilm development and rigidity. METHODS AND RESULTS: Formed biofilm mass was measured with 0·1% crystal violet staining method and observed with a scanning electron microscope. The localizations of eDNA and extracellular HLP (eHLP) in formed biofilm were detected by staining with 7-hydoxyl-9H-(1,3-dichloro-9,9-dimethylacridin-2-one) and anti-HLP antibody without fixation, respectively. DNase I treatment (200 U ml(-1)) markedly decreased biofilm formation and cell density in biofilms. Colocalization of eHLP and eDNA in biofilm was confirmed. The addition of eDNA (up to 1 µg ml(-1)) purified from Strep. intermedius, other Gram-positive bacteria, Gram-negative bacteria, or human KB cells into the Strep. intermedius culture increased the biofilm mass of all tested strains of Strep. intermedius, wild-type, HLP-downregulated strain and control strains. In contrast, the addition of eDNA (>1 µg ml(-1)) decreased the biofilm mass of all Strep. intermedius strains. CONCLUSIONS: These findings demonstrated that eDNA and eHLP play crucial roles in biofilm development and its rigidity. SIGNIFICANCE AND IMPACT OF THE STUDY: eDNA- and HLP-targeting strategies may be applicable to novel treatments for bacterial biofilm-related infectious diseases.

Comparison of the antibacterial effect of modified 3-mix paste versus Ultrapex over anaerobic microorganisms from infected root canals of primary teeth: an in vitro study.

OBJECTIVE: The aim of this study was to evaluate in vitro the antimicrobial efficacy of a modified 3-mix paste and to compare it with an iodoform paste (Ultrapex) against anaerobic microorganisms isolated from root canals of infected or necrotic primary teeth. STUDY DESIGN: An in vitro experimental assay was performed over isolated and identified anaerobic microorganisms of 21 samples, in order to compare the antimicrobial ability of both root canal filling materials, using a disc-diffusion method. RESULTS: A total of 21 microbial samples (15 polymicrobial and 6 monomicrobial) were obtained, from which 19 different strains were identified. Modified 3-mix paste showed an excellent antimicrobial effect against most of both kinds of microbial samples, although some of them exhibited resistance; on the other hand, Ultrapex showed only minimal antimicrobial ability (null or low categories). Clostridium ramosum exhibited the most resistance to both materials. CONCLUSION: The bactericidal effect of the modified 3-mix paste was superior to Ultrapex, with a statistically significant difference, against anaerobic microorganisms isolated from infected root canals of primary teeth.

Clinical and microbiologic effects of commercially available dentifrice containing aloe vera: a randomized controlled clinical trial.

BACKGROUND: Certain plants used in folk medicine serve as a source of therapeutic agents that have antimicrobial and other multipotential effects. This prospective, randomized, placebo, and positively controlled clinical trial was designed to evaluate the clinical and microbiologic effects of a commercially available dentifrice containing aloe vera on the reduction of plaque and gingival inflammation in patients with gingivitis. METHODS: Ninety patients diagnosed with chronic generalized gingivitis were selected and randomly divided into three groups: group 1, placebo toothpaste; group 2, toothpaste containing aloe vera; and group 3, toothpaste with polymer and fluoride containing triclosan. Clinical evaluation was undertaken using a gingival index, plaque was assessed using a modification of the Quigley-Hein index, and microbiologic counts were assessed at baseline, 6 weeks, 12 weeks, and 24 weeks. A subjective evaluation was also undertaken by questionnaire. RESULTS: Toothpaste containing aloe vera showed significant improvement in gingival and plaque index scores as well as microbiologic counts compared with placebo dentifrice. These improvements were comparable to those achieved with toothpaste containing triclosan. CONCLUSION: Toothpaste containing aloe vera may be a useful herbal formulation for chemical plaque control agents and improvement in plaque and gingival status.

A lingual abscess caused by Streptococcus intermedius.

Lingual abscesses are rare. We describe a case in a healthy female with no recent history of trauma. The organism recovered by culture of drainage material collected prior to antibiotic treatment was Streptococcus intermedius, an organism recognized as flora of the oropharynx and associated with abscess formation. The isolate was resistant to clindamycin, which was the antibiotic therapy that the patient received.

Effect of saliva viscosity on the co-aggregation between oral streptococci and Actinomyces naeslundii.

BACKGROUND: The co-aggregation of oral bacteria leads to their clearance from the oral cavity. Poor oral hygiene and high saliva viscosity are common amongst the elderly; thus, they frequently suffer from pneumonia caused by the aspiration of oral microorganisms. OBJECTIVES: To examine the direct effect of saliva viscosity on the co-aggregation of oral streptococci with actinomyces. MATERIALS AND METHODS: Fifteen oral streptococcal and a single actinomyces strain were used. Co-aggregation was assessed by a visual assay in phosphate buffer and a spectrophotometric assay in the same buffer containing 0-60% glycerol or whole saliva. RESULTS: Nine oral streptococci co-aggregated with Actinomyces naeslundii ATCC12104 in the visual assay and were subsequently used for the spectrophotometric analysis. All tested strains displayed a decrease in co-aggregation with increasing amounts of glycerol in the buffer. The co-aggregation of Streptococcus oralis with A. naeslundii recovered to baseline level following the removal of glycerol. The per cent co-aggregation of S. oralis with A. naeslundii was significantly correlated with the viscosity in unstimulated and stimulated whole saliva samples (correlation coefficients: -0.52 and -0.48, respectively). CONCLUSION: This study suggests that saliva viscosity affects the co-aggregation of oral streptococci with actinomyces and that bacterial co-aggregation decreases with increasing saliva viscosity.