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PubMed2005

First isolation of Streptococcus downei from human dental plaques.

In this study, we isolated four bacterial strains grown on mitis-salivarius sucrose bacitracin agar. The strains had similar biochemical characteristics to biotypes I or II of mutans streptococci. The four isolates were identified as Streptococcus downei by 16S rDNA and dextranase gene (dex) sequencing as well as polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) targeting dex. To our knowledge, this is the first report of the isolation and identification of S. downei from dental plaque in humans. The results suggest that S. downei can inhabit the human oral cavity.

Culture Media↗

PubMed2001

[Typing of Streptococcus mutans (serotype C) by arbitrarily primed polymerase chain reaction].

OBJECTIVE: To study the relationship between genetic diversity within Streptococcus mutans (C), and dental caries. METHODS: One reference strain (GS5) was used to establish the suitability of primers and the initial AP-PCR conditions. Isolates of Streptococcus mutans (C) were obtained from 19 caries-free and 40 caries-active patients, and were originally isolated on mitis-salivarius-bacitracin agar. Totally 278 isolates were biochemically and serologically confirmed as Streptococcus mutans (C). Chromosomal DNA was extracted from those isolates by Chelex. One random primer, OPA-05 was used as a template in PCR reaction. All amplification reactions were conducted in a volume of 20 microliters containing 2 microliters of 10 x reaction buffer, 200 mumol/L each of dATP, dCTP, dGTP and dTTP and 2.5 units of Taq DNA polymerase. We varied the concentrations of MgCl2 (1.5-4.5 mmol/L), template DNA (2-40 ng), and primer (0.3-0.5 mumol/L) to determine the optical conditions for AP-PCR. The temperature profile in a thermocycler was 35 cycles as follows: 94 degrees C for 1 min, at 36 degrees C for 2 min, at 72 degrees C for 2 min. The initial denaturation was at 94 degrees C for 5 min and the final extension at 72 degrees C for 5 min. RESULTS: The MgCl2 concentration in 4 mmol/L and the primer concentration in 0.5 mumol/L produced clearly distinct and reproducible fingerprints. We found 105 different patterns among the 278 isolates. In high-caries persons more than one genotype isolates of Streptococcus mutans (C) were colonized. CONCLUSIONS: Isolates of Streptococcus mutans (C) exist apparent genetic diversity. The genotypes of isolates might relate to differences in caries susceptibility.

DNA, Bacterial↗

PubMed1980

Bacterial interference of Neisseria gonorrhoeae by alpha-haemolytic streptococci.

Fifty pharyngeal isolates of alpha-haemolytic streptococci were tested against 20 cervical isolates of Neisseria gonorrhoeae for bacterial interference in vitro using the lawn-spotting method. Forty-seven (94%) isolates of streptococci showed inhibitory activity toward N gonorrhoeae, although nine of these were inhibitory to only one isolate of N gonorrhoeae. Isolates of N gonorrhoeae varied widely in their sensitivity to streptococci; the most sensitive were inhibited by 40 isolates of streptococci and the least sensitive by only 14 isolates. Species of Streptococcus found to inhibit growth of N gonorrhoeae were S mitis, S MG intermedius, S sanguis II, S mutans, and S morbillorum.

Adult↗

PubMed1991

Antibiotic-resistant oral streptococci in dental patients susceptible to infective endocarditis.

The aim of this study was to determine the incidence of amoxycillin and erythromycin resistance in oral streptococci in patients at risk from infective endocarditis. Samples of gingival crevicular flora were taken from 65 patients at the site of dental treatment, prior to the prophylactic administration of amoxycillin (54 patients) or erythromycin (11 patients). Samples were also taken from 65 dental patients who were not considered to be at risk from infective endocarditis. No isolate had a minimum inhibitory concentration (MIC) of amoxycillin greater than 24 mg/L. However, erythromycin-resistant oral streptococci with MIC values greater than 3.5 mg/L were isolated from 22% of patients receiving amoxycillin prophylaxis, 9% of patients receiving amoxycillin prophylaxis, 9% of patients given erythromycin prophylaxis and 9% of patients not at risk from infective endocarditis. The antibiotic-resistant streptococci comprised mainly Streptococcus sanguis biotype II, although S. sanguis biotype I, S. mitis and S. salivarius were also frequently recovered.

Amoxicillin↗

PubMed1991

A scheme for the identification of viridans streptococci.

A collection of strains representing all the currently recognised species of oral or viridans streptococci (Streptococcus sanguis, S. parasanguis, S. gordonii, S. oralis, S. mitis, S. salivarius, S. vestibularis, S. anginosus, S. constellatus, s. intermedius, S. mutans and S. sobrinus) were tested for the production of a range of glycosidase activities with 4-methylumbelliferyl-linked fluorogenic substrates, and for reactions in a range of conventional fermentation and hydrolytic tests. The resulting biochemical scheme, consisting of 14 tests, enabled the differentiation of all these species and distinguished three biotypes within S. sanguis. The scheme reported here represents an improvement over currently available schemes for the identification of viridans streptococci.

Culture Media↗

PubMed1980

Changes in some biological properties of Streptococcus salivarius isolates from infected rats.

Isolation of colonies with altered colonial morphology was frequently observed following infection of specific pathogen-free rats fed a caries-inducing high sucrose diet with two human strains of Streptococcus salivarius. These isolates produced unique rough colonies on mitis salivarius agar, in contrast to the smooth, mucoid colonial morphology of the parental strain. The rough variants were characterized by a marked increase in the synthesis of polysaccharide from sucrose, especially cell-associated water-insoluble glucan, whereas the smooth parental strains mainly produced cell-free fructan. Glucose-grown cells of rough variants but not smooth parental strains were aggregated upon addition of sucrose. Such variants were not detected during laboratory transfers of the human strains of S. salivarius nor by mutagenesis of the cultures.

Animals↗

PubMed1976

Comparative ultrastructure of selected oral streptococci: thin-sectioning and freeze-etching studies.

The ultrastructure of Streptococcus mutans, serotypes a-e, S. sanguis, S. mitis, and S. salivarius HHT, were examined by the techniques of thin-sectioning and freeze-etching. The cell walls varied in width between 15 and 46 nm and were covered with an electron-dense fibrillar or fuzz layer. The cytoplasmic membrane was in close association with numerous mesosomes which were, in turn, either closely associated or in contact with the bacterial chromosome. In freeze-etch replicas, the outermost layer of the cell wall was fibrous, and the cytoplasmic membrane was covered with particles composed of several subunits. Both particle-clusters and particle-free areas occurred on the surfaces of the cytoplasmic membrane, as well as a crystalline array in the ground plasm of the cell.

Cell Membrane↗

PubMed2004

Identification of clinically relevant viridans group streptococci by sequence analysis of the 16S-23S ribosomal DNA spacer region.

The feasibility of sequence analysis of the 16S-23S ribosomal DNA (rDNA) intergenic spacer (ITS) for the identification of clinically relevant viridans group streptococci (VS) was evaluated. The ITS regions of 29 reference strains (11 species) of VS were amplified by PCR and sequenced. These 11 species were Streptococcus anginosus, S. constellatus, S. gordonii, S. intermedius, S. mitis, S. mutans, S. oralis, S. parasanguinis, S. salivarius, S. sanguinis, and S. uberis. The ITS lengths (246 to 391 bp) and sequences were highly conserved among strains within a species. The intraspecies similarity scores for the ITS sequences ranged from 0.98 to 1.0, except for the score for S. gordonii strains. The interspecies similarity scores for the ITS sequences varied from 0.31 to 0.93. Phylogenetic analysis of the ITS regions revealed that evolution of the regions of some species of VS is not parallel to that of the 16S rRNA genes. One hundred six clinical isolates of VS were identified by the Rapid ID 32 STREP system (bioMérieux Vitek, Marcy l'Etoile, France) and by ITS sequencing, and the level of disagreement between the two methods was 18% (19 isolates). Most isolates producing discrepant results could be unambiguously assigned to a specific species by their ITS sequences. The accuracy of using ITS sequencing for identification of VS was verified by 16S rDNA sequencing for all strains except strains of S. oralis and S. mitis, which were difficult to differentiate by their 16S rDNA sequences. In conclusion, identification of species of VS by ITS sequencing is reliable and could be used as an alternative accurate method for identification of VS.

DNA, Ribosomal Spacer↗

PubMed2005

Identification of clinically relevant viridans streptococci by an oligonucleotide array.

Viridans streptococci (VS) are common etiologic agents of subacute infective endocarditis and are capable of causing a variety of pyogenic infections. Many species of VS are difficult to differentiate by phenotypic traits. An oligonucleotide array based on 16S-23S rRNA gene intergenic spacer (ITS) sequences was developed to identify 11 clinically relevant VS. These 11 species were Streptococcus anginosus, S. constellatus, S. gordonii, S. intermedius, S. mitis, S. mutans, S. oralis, S. parasanguinis, S. salivarius, S. sanguinis, and S. uberis. The method consisted of PCR amplification of the ITS regions by using a pair of universal primers, followed by hybridization of the digoxigenin-labeled PCR products to a panel of species-specific oligonucleotides immobilized on a nylon membrane. After 120 strains of the 11 species of VG and 91 strains of other bacteria were tested, the sensitivity and specificity of the oligonucleotide array were found to be 100% (120 of 120 strains) and 95.6% (87 of 91 strains), respectively. S. pneumoniae cross-hybridized to the probes used for the identification of S. mitis, and simple biochemical tests such as optochin susceptibility or bile solubility should be used to differentiate S. pneumoniae from S. mitis. In conclusion, identification of species of VS by use of the present oligonucleotide array is accurate and could be used as an alternative reliable method for species identification of strains of VS.

DNA, Bacterial↗

PubMed1983

Inhibition by the antimicrobial agent chlorhexidine of acid production and sugar transport in oral streptococcal bacteria.

Oral streptococci transport sugars via the phosphoenolpyruvate-phosphotransferase (PEP-PTS) system. In a specific assay of this system, low concentrations of chlorhexidine abolished the activity of the glucose and sucrose PTS in batch-grown cells of Streptococcus mutans Ingbritt and B13, Strep. sanguis NCTC 7865, Strep. mitis ATCC 903, Strep. milleri NCTC 10709 and Strep. salivarius NCTC 8606. Intact cells and cells made permeable to the assay reagents with toluene were used. Toluenized cells were more sensitive to chlorhexidine than intact cells (0.09 and 0.25 mM, respectively). This PTS-inhibitory concentration of chlorhexidine reduced acid production from glucose in pH fall experiments to values higher than are obtained solely from endogenous metabolism. The effect of chlorhexidine on rates of acid production was determined at pH 7.0 using cells washed with either 135 mM NaCl or 135 mM KCl. In general, faster rates of acid production from the metabolism of glucose and sucrose were obtained with potassium-treated cells. Addition of the PTS-inhibitory concentration of chlorhexidine markedly reduced or totally abolished acid production by NaCl-treated cells; a greater residual-activity was detected in the same cells washed with KCl (except with Strep. mutans B13 and Strep. mitis ATCC 903). The PTS-inhibitory concentration of chlorhexidine had little or no effect on the viability of cells. The results confirm the existence of sugar uptake systems in oral streptococci additional to the PTS and provide an explanation for the additive anti-caries effect of mouth-rinses containing both fluoride and chlorhexidine.

Acids↗

PubMed2004

Accuracy of phenotypic and genotypic testing for identification of Streptococcus pneumoniae and description of Streptococcus pseudopneumoniae sp. nov.

We have identified an unusual group of viridans group streptococci that resemble Streptococcus pneumoniae. DNA-DNA homology studies suggested that a subset of these isolates represent a novel species that may be included in the S. oralis-S. mitis group of viridans group streptococci. We suggest that this novel species be termed Streptococcus pseudopneumoniae. A combination of phenotypic and genetic reactions allows its identification. S. pseudopneumoniae strains do not have pneumococcal capsules, are resistant to optochin (inhibition zones, less than 14 mm) when they are incubated under an atmosphere of increased CO2 but are susceptible to optochin (inhibition zones, >14 mm) when they are incubated in ambient atmospheres, are not soluble in bile, and are positive by the GenProbe AccuProbe Pneumococcus test. The bile solubility test is more specific than the optochin test for identification of S. pneumoniae. Genetic tests for pneumolysin (ply) and manganese-dependent superoxide dismutase (sodA) and identification tests with a commercial probe, AccuProbe Pneumococcus, do not discriminate between the new species and S. pneumoniae.

Bacterial Typing Techniques↗

PubMed2001

Nationwide German multicenter study on the prevalence of antibiotic resistance in streptococcal blood isolates from neutropenic patients and comparative in vitro activities of quinupristin-dalfopristin and eight other antimicrobials.

In a prospective multicenter study (1996 to 1999), 156 episodes of bacteremic streptococcal infections of neutropenic patients were evaluated. Streptococcus oralis (26.3%), S. pneumoniae (26.3%), S. agalactiae (11.5%), S. mitis (9%), and S. pyogenes (5.8%) were the predominant species. Four strains (2.6%) were found to be intermediately resistant to penicillin. One strain (0.6%) was found to be highly resistant to penicillin (MIC, 8 mg/liter). Reduced susceptibility to penicillin was detected among S. oralis (14.6%), S. mitis (7.1%), and S. pneumoniae (4.9%) isolates but was not recorded among S. agalactiae and S. pyogenes. Resistance rates and intermediate resistance rates for other antimicrobials were as follows (all species): amoxicillin, 1.3 and 3.2%; erythromycin, 16 and 2.6%; clindamycin, 5.8 and 0%; ciprofloxacin, 1.9 and 7.7%. Quinupristin-dalfopristin showed good in vitro activity against most streptococcal isolates (MIC at which 50% of the isolates were inhibited [MIC(50)], 0.5 mg/liter; MIC(90), 1 mg/liter, MIC range, 0.25 to 4 mg/liter).

Adolescent↗

PubMed1990

Sialidase activity of the "Streptococcus milleri group" and other viridans group streptococci.

Viridans group streptococci were examined for the production of sialidase (neuraminidase) activity, using the fluorescent substrate 4-methylumbelliferyl-alpha-D-N-acetylneuraminic acid in a simple and rapid (15-min) assay. Sialidase was produced by all strains of Streptococcus oralis and S. intermedius and by a majority of S. mitis strains. S. mutans, S. sobrinus, S. gordonii, S. sanguis, S. vestibularis, S. salivarius, S. anginosus, S. constellatus, "S. parasanguis," and the "tufted fibril group" were uniformly negative. Sialidase production may be a useful characteristic to assist in the identification of viridans group streptococci.

Humans↗

PubMed1995

A study on prevention of hospital infection control caused by tooth preparation dust in the dental clinic. Part 1. Preventive measures against environmental pollution in the dental clinic caused by microbial particles.

Tooth preparation dust, an indispensable part of dental treatment, contains bacteria and viruses. In order to examine environmental pollution in the dental clinic during tooth preparation, we monitored microorganism spread by counting colony forming units (CFU) with the "Andersen Microbe Sampler." This test was conducted in a "Clean Booth." Mitis-salivarius medium was used to count the oral-streptococcus species. The ability of the "Extra-Oral Vacuum Aspirator (EOVA)" to effectively eliminate contamination was also tested, and the EOVA was found to reduce the spread of oral-streptococci, significantly. The EOVA was therefore viewed as an effective method for reducing air pollution in the dental clinic, and should be used when treating patients with certain infectious diseases.

Air Microbiology↗

PubMed1980

Comparison of selective broth and plating methods for isolation of Streptococcus mutans from root surface dental plaques.

Approximately 150 sound root surfaces in 44 subjects prone to root surface caries were sampled longitudinally to determine the microbial flora associated with root caries initiation. During the first 16 months of the study, a comparison of Streptococcus mutans recovery was made by using three bacteriological media: mitis-salivarius agar (MSA), mitis-salivarius-bacitracin-sucrose agar (MSB), and a partially selective mannitol-containing broth. Total streptococcal and S. mutans populations were found to be much lower than in previous reports. MSB was more selective; S. mutans was detected in many samples when its numbers were too low for isolation on MSA. However, recovery of S. mutans was greater on MSA than on MSB for most samples yielding colonies on both media. Mannitol-containing broth used as an enrichment medium yielded the highest frequency of S. mutans isolation among the three media.

Aged↗

PubMed1975

Membrane filter technique for enumeration of enterococci in marine waters.

A membrane filter procedure is described for the enumeration of enterococci in marine waters. The procedure utilizes a highly selective and somewhat differential primary isolation medium followed by an in situ substrate test for identifying colonies of those organisms capable of hydrolyzing esculin. The procedure (mE) was evaluated with known streptococci strains and field samples with regard to its accuracy, sensitivity, selectivity, precision, and comparability to existing methods. Essentially quantitative recovery was obtained with seawater-stressed cells of Streptococcus faecalis and S. faecium. Neither S. bovis, S. equinus, S. mitis, nor S. salivarius grew on the medium. The selectivity of the medium was such that a 10,000-fold reduction in background organisms was obtained relative to a medium which contained no inhibitors and was incubated at 35 C. About 90% of those typical colonies designated as enterococci confirmed as such and about 12% of the colonies not so designated were, in fact, identified as enterococci. Plate to plate variability across samples approximated that expected by chance alone. Verified recoveries of enterococci from natural samples by the mE procedure, on the average, exceeded those by the KF method by one order of magnitude.

Cell Count↗

PubMed1985

Cross-reactions between alpha-streptococci and Omniserum, a polyvalent pneumococcal serum, demonstrated by direct immunofluorescence, immunoelectroosmophoresis, and latex agglutination.

In recent years several groups have used serological methods to demonstrate pneumococcal capsular antigens in sputum. In the present study 123 strains of alpha-hemolytic streptococci (including 97 strains from sputum or pharyngeal specimens) were tested for cross-reactions with a polyvalent antipneumococcal serum (Omniserum). Representatives of the following species were included: Streptococcus bovis, S. equinus, S. intermedius, S. lactis, S. milleri, S. mitis, S. mutans, S. sobrinus, S. salivarius, S. sanguis, S. suis, and Aerococcus viridans. Serological reactions were detected by direct immunofluorescence, immunoelectroosmophoresis, and latex agglutination. Fifteen (12%) of the strains gave positive reactions by all three methods. Positive reactions were also observed with another 32 strains (26%) with two of the methods, whereas 37 strains (30%) gave positive reactions by just one technique. Altogether 84 (68%) strains gave positive reactions with one or more of the methods. Latex agglutination gave positive reactions with 26 (21%) strains compared with 57 (46%) in immunofluorescence and 63 (51%) in immunoelectroosmophoresis. Absorption of the antiserum with one alpha-hemolytic strain reduced but did not entirely eliminate the cross-reactions with five tested strains. These findings indicate a potential risk of cross-reactions with polyvalent antipneumococcal serum in tests carried out on sputa or other specimens which may be contaminated with alpha-hemolytic streptococci.

Cross Reactions↗

PubMed1994

Conservation of salivary glycoprotein-interacting and human immunoglobulin G-cross-reactive domains of antigen I/II in oral streptococci.

In this study we localized more precisely the salivary glycoprotein-interacting and the human immunoglobulin G (hIgG)-cross-reacting domains on the SR molecule, an antigen I/II-related protein from S. mutans serotype f. Mapping of the SR molecule with polypeptides expressed by subclones covering the entire molecule and with synthetic peptides demonstrates that the salivary glycoprotein-binding domain is located in the N-terminal alanine-rich repeats of the SR molecule. In order to investigate the degree of conservation of both regions in various oral streptococci, we tested the reactivity of 8 representative strains of the mutans group and 11 nonmutans oral Streptococcus strains (S. anginosus, S. milleri, S. constellatus, S. intermedius, S. mitis, S. sanguis, S. gordonii, S. salivarius, and S. mitis strains) with antipeptide antibodies in a whole-cell enzyme linked immunosorbent assay together with colony hybridization analysis using DNA probes designed to map these two regions. All the mutans group strains except S. rattus and the 11 nonmutans streptococcal strains showed a high conservation of the C-terminal part of the SR molecule, especially the hIgG-cross-reacting domain, and less homology for the N-terminal salivary glycoprotein-binding region. Almost all of the sera from patients with rheumatic disease reacted strongly with SR from S. mutans serotype f, P1 from S. mutans serotype c, and four peptides located in the hIgG-cross-reacting region and not with peptides located at the C and N termini and in the proline-rich repeats. These results confirm that epitopes located within this region are immunogenic in humans and could lead to the synthesis of natural anti-IgG antibodies.

Animals↗