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PubMed2002

A XerD recombinase with unusual active site motifs in Streptococcus pneumoniae.

XerD belongs to the site specific recombinases of the integrase family of proteins that catalyze recombination events via a phosphotyrosine intermediate. Sequence alignments and crystal structure resolution of E. coli XerD and related enzymes demonstrated the importance of four conserved amino acids R-H-R-H that are spaced along the C-terminal domain in addition to a conserved K and the active site Y, all of which have been implicated in catalysis. The deduced amino acid sequence of the putative S. pneumoniae XerD contained three unique replacements at the conserved positions resulting in L-Q-R-L; moreover, the active site Y was the penultimate amino acid residue, and the extreme C-terminal region suggested to be involved in interaction of E. coli XerD with XerC was lacking. Severe growth defects in a loss-of-function xerD mutant are consistent with an important in vivo function of the S. pneumoniae XerD protein. Highly related xerD genes with similar unusual amino acid replacements were found in S. mitis, S. mutans and S. pyogenes but not in other Gram-positive bacteria, although the genetic environment was very similar in many species. There are at least another four genes in the S. pneumoniae KNR_7/87 genome encoding Xer related peptides, one of which was identified as the xerC homologue. The xerD and xerC genes were present in a sample of 20 S. pneumoniae strains whereas the other xer genes appear to be absent in some of the strains and are more closely related to integrases of phage and transposon origin.

Amino Acid Motifs↗

PubMed2003

Identification of Streptococcus sanguinis with a PCR-generated species-specific DNA probe.

The objective of the present study was to design a PCR-generated DNA probe and determine the specificity of the probe for the identification of clinical isolates of Streptococcus sanguinis. To do this, we examined over 200 arbitrarily primed PCR (AP-PCR) amplicon patterns obtained with DNA from clinical isolates of S. sanguinis. A 1.6-kb DNA amplicon that was common to all AP-PCR profiles was extracted from agarose gels and then cloned and sequenced. A search for a similar sequence in the GenBank database with the BLASTN program revealed that the 1.6-kb DNA fragment comprised an intergenic region between two housekeeping genes, uncC (proton-translocating ATPase) and murA (UDP-N-acetylglucosamine enolpyruvyl transferase). Three digoxigenin-labeled DNA probes were synthesized on the basis of the sequence of the 1.6-kb fragment: the sequence of probe SSA-1 contained the proton-translocating ATPase (uncC) and the entire intergenic region, the sequence of probe SSA-2 contained only the intergenic region, and the sequence of probe SSA-3 contained an internal region of the murA gene. Dot blot hybridization showed that the three probes displayed signals for hybridization to both S. sanguinis strain ATCC 10556 and the S. sanguinis clinical isolates. Probe SSA-1, however, hybridized to DNA from S. oralis and S. mitis. Probe SSA-3 hybridized to DNA from S. gordonii, S. mitis, S. oralis, S. parasanguinis, and S. vestibularis. The probe SSA-2-specific intergenic region appeared to be specific for S. sanguinis. The results from this study suggest that probe SSA-2 may serve as a species-specific DNA probe for the identification of clinical isolates of S. sanguinis.

Base Sequence↗

PubMed1997

The use of X-ray photoelectron spectroscopy for the study of oral streptococcal cell surfaces.

Physicochemical and structural properties of microbial cell surfaces play an important role in their adhesion to surfaces and are determined by the chemical composition of the outermost cell surface. Many traditional methods used to determine microbial cell wall composition require fractionation of the organisms and consequently do not yield information about the composition of the outermost cell surface. X-ray photoelectron spectroscopy (XPS) measures the elemental composition of the outermost cell surfaces of micro-organisms. The technique requires freeze-drying of the organisms, but, nevertheless, elemental surface concentration ratios of oral streptococcal cell surfaces with peritrichously arranged surface structures showed good relationships with physicochemical properties measured under physiological conditions, such as zeta potentials. Isoelectric points appeared to be governed by the relative abundance of oxygen- and nitrogen-containing groups on the cell surfaces. Also, the intrinsic microbial cell-surface hydrophobicity by water contact angles related to the cell-surface composition as by XPS and was highest for strains with an elevated isoelectric point. Inclusion of elemental surface compositions for tufted streptococcal strains caused deterioration of the relationships found. Interestingly, hierarchical cluster analysis on the basis of the elemental surface compositions revealed that, of 36 different streptococcal strains, only four S. rattus as well as nine S. mitis strains were located in distinct groups, well separated from the other streptococcal strains, which were all more or less mixed in one group.

Animals↗

PubMed2003

Invasion and killing of human endothelial cells by viridans group streptococci.

Colonization of the cardiovascular endothelium by viridans group streptococci can result in infective endocarditis and possibly atherosclerosis; however, the mechanisms of pathogenesis are poorly understood. We investigated the ability of selected oral streptococci to infect monolayers of human umbilical vein endothelial cells (HUVEC) in 50% human plasma and to produce cytotoxicity. Planktonic Streptococcus gordonii CH1 killed HUVEC over a 5-h period by peroxidogenesis (alpha-hemolysin) and by acidogenesis but not by production of protein exotoxins. HUVEC were protected fully by addition of supplemental buffers and bovine liver catalase to the culture medium. Streptococci were also found to invade HUVEC by an endocytic mechanism that was dependent on polymerization of actin microfilaments and on a functional cytoskeleton, as indicated by inhibition with cytochalasin D and nocodazole. Electron microscopy revealed streptococci attached to HUVEC surfaces via numerous fibrillar structures and bacteria in membrane-encased cytoplasmic vacuoles. Following invasion by S. gordonii CH1, HUVEC monolayers showed 63% cell lysis over 4 h, releasing 64% of the total intracellular bacteria into the culture medium; however, the bacteria did not multiply during this time. The ability to invade HUVEC was exhibited by selected strains of S. gordonii, S. sanguis, S. mutans, S. mitis, and S. oralis but only weakly by S. salivarius. Comparison of isogenic pairs of S. gordonii revealed a requirement for several surface proteins for maximum host cell invasion: glucosyltransferase, the sialic acid-binding protein Hsa, and the hydrophobicity/coaggregation proteins CshA and CshB. Deletion of genes for the antigen I/II adhesins, SspA and SspB, did not affect invasion. We hypothesize that peroxidogenesis and invasion of the cardiovascular endothelium by viridans group streptococci are integral events in the pathogenesis of infective endocarditis and atherosclerosis.

Cells, Cultured↗

PubMed1994

Site-related streptococcal attachment to buccocervical tooth surfaces. A correlative micromorphologic and microbiologic study.

Scanning electron (SEM) microscopy of epoxy replicas made from dental impressions has shown that in buccal gingival recession the root surfaces are devoid of cementum, leaving the dentin exposed. In this study replication techniques were applied to correlate the micromorphology of the buccocervical region with early streptococcal attachment. The subjects were 27 healthy young adults. The buccocervical surfaces of all the premolars were meticulously cleaned. The subjects fasted for 2 h before impression-taking. Replicas were made from impressions in hydrophilic A-silicone, and streptococcal attachment was visualized by light microscopy of mitis-salivarius agar replicas incubated anaerobically for 48 h. The surface micromorphology was documented by SEM of corresponding epoxy replicas. Colonization only 2 h after cleaning was very sparse. Sites with healthy or inflamed gingivae had markedly different colonization patterns in the sulcular region. In 4 subjects with a total of 12 sites where gingival recession, undetected clinically, was disclosed by SEM, representative colonies were retrieved and identified microbiologically to species level. Two healthy sites per subject were also sampled. Streptococcus mutans and S. sobrinus were identified from eight sites with exposed root dentin. S. oralis predominated on the enamel surfaces. The method offers a valuable complement to in situ and in vitro microbiologic studies of exposed dentin and a novel technique for sampling clinical isolates of streptococci.

Adolescent↗

PubMed1990

Effect of chlorhexidine-sodium fluoride gel applied by tray or by toothbrush on salivary mutans streptococci.

A chlorhexidine (1%)-sodium fluoride (0.2%) gel applied using a dental tray six times during two consecutive days significantly decreased salivary levels of mutans streptococci but had only a slight effect on salivary levels of lactobacilli. The same gel used as a dentifrice for one week (twice a day) also significantly reduced salivary levels of mutans streptococci. However, this treatment was somewhat less effective than that using trays, on the basis of the ratio of mutans streptococci to total aerobes. With both treatments baseline salivary levels of mutans streptococci were re-attained after 70 days. One day after the gel treatment, mutans streptococci were unable to adhere to plastic strips (Dentocult SM Strip mutans) but detectable growth was found on mitis-salivarius-bacitracin agar. This suggests that not only are numbers of mutans streptococci reduced after chlorhexidine-sodium fluoride gel treatment but also mutans streptococci adherence.

Administration, Topical↗

PubMed2001

Oral colonization and cariogenicity of Streptococcus gordonii in specific pathogen-free TAN:SPFOM(OM)BR rats consuming starch or sucrose diets.

The significance of Streptococcus gordonii in dental caries is undefined, as is that of other alpha-amylase-binding bacteria (ABB) commonly found in the mouth. To clarify the ecological and cariological roles of S. gordonii our specific pathogen-free Osborne-Mendel rats, TAN:SPFOM(OM)BR, were fed either diet 2000 (containing 56% confectioner's sugar, most of which is sucrose) or diet 2000CS (containing 56% cornstarch, in lieu of confectioner's sugar) and inoculated with S. gordonii strains. Uninoculated rats were free of both indigenous mutans streptococci (MS) and ABB, including S. gordonii, as shown by culture on mitis salivarius and blood agars of swabs and sonicates of dentitions after weanlings had consumed these diets for 26 days. ABB were detected by radiochemical assay using [125I]-amylase reactive to alpha-amylase-binding protein characteristic of the surface of S. gordonii and other ABB. No ABB were detected (detection limit < 1 colony-forming units in 10(6) colony-forming units). Thus the TAN:SPFOM(OM)BR colony presents a 'clean animal model' for subsequent study. Consequently, S. gordonii strains Challis or G9B were used to inoculate weanling rat groups consuming either the high-sucrose diet 2000 or the cornstarch diet 2000CS. Two additional groups fed each of these diets remained unioculated. Recoveries of inoculants were tested 12 and 26 days later by oral swabs and sonication of the molars of one hemimandible of each animal, respectively. Uninoculated animals were reconfirmed to be free of ABB and mutans streptococci, but inoculated ones eating diet 2000CS had S. gordonii recoveries of 1-10% or, if eating diet 2000, 10-30% of total colony-farming units in sonicates. There were no statistically significant differences among the inoculated and uninoculated animal groups' caries scores when they ate the cornstarch diet. Lesion scores for sucrose-eating rats were, however, from 2.4-5.1-fold higher than for cornstarch-eating rats, P < 0.001, and were still higher if animals had been inoculated with either Challis (1.41-fold) or G9B (1.64-fold), than if uninoculated, both P < 0.001, so long as the rats ate the sucrose diet. Therefore, TAN:SPFOM(OM)BR rats do not harbour ABB or S. gordonii but can be colonized by S. gordonii. Colonization levels of S. gordonii on the teeth are higher in the presence of high sucrose than with high starch-containing diets. Caries scores are augmented by sucrose compared with starch, and are further augmented by S gordonii colonization. S. gordonii is thus cariologically significant in the presence of sucrose, at least in this rat.

Amylases↗

PubMed1993

The resistance and adaptation of selected oral bacteria to mercury and its impact on their growth.

Selected strains of oral Streptococcus and Actinomyces have been tested for their ability to grow in the presence of mercury. Strains were tested for growth on a semi-defined medium with low mercury-binding characteristics. Sensitivities were initially measured on agar plates, and subsequently, selected strains were grown in broth so that the impact of mercury on the growth characteristics could be determined. Streptococci were more resistant to mercury (5 micrograms/mL to 40 micrograms/mL) than Actinomyces (< 5 micrograms/mL to 30 micrograms/mL). The most resistant streptococci included S. mitis biovar 1, S. salivarius, S. sobrinus, and one strain of S. mutans, all of which grew on agar with 40 micrograms/mL of mercury. Two other S. mutans strains were more sensitive, being inhibited by 10 and 20 micrograms/mL mercury. The most resistant Actinomyces was A. naeslundii genospecies 1 (ATCC12104), which grew on medium with 30 micrograms/mL mercury; two strains of Actinomyces were completely inhibited by 5 micrograms/mL. Mercury caused increased lag times and reduced cell density in broth cultures. Enrichment cultures of samples of human dental plaque showed that streptococci were the most resistant organisms that could be cultured on the medium and that these strains could adapt to relatively high mercury concentrations. S. oralis and S. mitis biovar 1 were the most resistant organisms isolated from enriched cultures, growing in broth media with 65 micrograms/mL mercury. Mercury was bound to cell walls and cell cytoplasm of streptococci grown in the presence of mercury.(ABSTRACT TRUNCATED AT 250 WORDS)

Amylases↗

PubMed1997

An outbreak of penicillin resistant Streptococcus pneumoniae investigated by a polymerase chain reaction based genotyping method.

AIMS: To characterise the genotypes of penicillin resistant Streptococcus pneumoniae infecting patients in a care of the elderly ward and to study its transmission in a hospital environment. METHODS: Isolates of S pneumoniae were cultured from specimens obtained from patients who had been admitted to a care of the elderly ward where an outbreak had occurred. Penicillin resistant S pneumoniae were also obtained from a series of surveillance throat swabs taken from patients in the same ward. In addition, all penicillin resistant S pneumoniae isolated from specimens submitted for culture at the time of the outbreak were included. Four sensitive strains isolated from a routine microbiology laboratory were included as controls. A simple polymerase chain reaction (PCR) based genotyping method for the penicillin binding protein (PBP) genes 1a, 2x, and 2b was used to characterise the genotypes. RESULTS: Nine patients were infected with serotype 9 S pneumoniae. Four of these patients died; two deaths were directly attributable to the infection. Tested against a battery of haemolytic streptococci and other organisms found in the respiratory tract, only two false positive reactions for PBP 2x were found among S mitis. The method demonstrated that the outbreak strain had altered PBP 1a, 2b, and 2x genes, a pattern clearly distinguishable from other penicillin resistant strains isolated at the same time. CONCLUSIONS: This method is simple to perform and would enable many laboratories to characterise the genotype of penicillin resistant S pneumoniae and investigate transmission in their hospitals.

Aged↗

PubMed1975

Increased selectivity of Mitis-Salivarius agar containing polymyxin.

Incorporation of the antibiotic polymixin B sulfate in plates of Mitis-Salivarius medium at a level of 100 U per ml effectively suppressed the growth of gram-negative spreading organisms which occasionally overgrow Mitis-Salivarius plates of oral samples and make them unsuitable for enumeration and differentiation of streptococci. At the indicated level the antibiotic did not affect the growth or typical colonial characteristics of the oral streptococci.

Agar↗

PubMed1991

Oxygen and the sugar metabolism in oral streptococci.

Streptococci have several ways of adapting themselves to the constantly changing environment of the human oral cavity. This paper discusses the adaptation of sugar metabolism to variations in oxygen levels. In all streptococci the Embden-Meyerhof pathway of glycolysis works under aerobic as well as anaerobic conditions, but pyruvate is converted into different metabolic end products depending on the oxygen levels. Under anaerobic conditions all streptococci form formate, acetate, and ethanol by a pyruvate formate-lyase pathway. If sugar is in excess, they also form lactate using a lactate dehydrogenase. Under aerobic conditions pyruvate formate-lyase is inactivated. This enzyme is then replaced by a pyruvate oxidase in some streptococci and by a pyruvate dehydrogenase in others. The characteristics of these enzymes help streptococci like S. sanguis, S. oralis, S. gordonii, and S. mitis to compete successfully with other bacteria in those sites of the oral cavity that are freely exposed to saliva, while mutans streptococci have to colonize anaerobic sites such as those in-between the teeth and in the occlusal fissures of the teeth.

Glucose↗

PubMed1987

The growth of oral bacteria on saliva.

The present experiments were aimed at studying the degradation of salivary glycoproteins by the oral microflora. To this end, S. sanguis I strain Ny476 and S. sanguis II (S. mitior) strain Ny581 were grown continuously in human-whole saliva. Under these conditions, the strains produced a variety of cell-associated hydrolytic activities, including glycosidases, exo- and endopeptidases, and esterases. S. sanguis II generally exhibited higher levels of enzyme activity than did S. sanguis I, in particular of neuraminidase that was produced only by S. sanguis II. In accordance, S. sanguis II had a higher cell yield and consumed a higher proportion of the sugars and sialic acid in the glycoproteins than did S. sanguis I. Interestingly, S. sanguis I, which is devoid of neuraminidase, is known to have a lectin with specificity for sialic acid, whereas S. sanguis II has affinity for galactose residues in the glycoproteins. We propose that specific binding of glycoproteins by oral bacteria constitutes a mechanism to collect nutrients in the vicinity of the cell. The special ability of S. sanguis II to utilize saliva for growth was further exemplified by its selection in batch-wise enrichments of dental plaque on saliva. The microflora in these enrichment cultures always consisted of Peptostreptococcus micros, S. sanguis II, and Fusobacterium nucleatum as the dominant organisms. Further, S. mitis and Gemella haemolysans were generally found to be present. The enrichment cultures produced a wide variety of mainly cell-bound hydrolytic enzymes. This resulted in almost complete breakdown of salivary glycoproteins in the culture.

Bacteriological Techniques↗

PubMedDate not supplied

[The classification of streptococci and human streptococcal diseases].

The modern nomenclature, phenotypic, medical, ecological and phylogenetic classification of streptococci and different classification of streptococcal human diseases are presented. All phylogenetic groups of streptococci have been shown to contain species causing diseases in man. The most medically significant groups are the phylogenetic groups Pyogenes and Mitis. Directions of the improvement of the classification of streptococci and streptococcal human diseases on the basis of modern concepts on the taxonomy of streptococci, the biological properties and ecology of the infective agents, as well as the genesis and clinical picture of diseases induced by them, have been determined.

Humans↗

PubMed1989

Speciation of viridans streptococci & their significance in human infections.

Sixty strains of viridans streptococci recovered, from various infections, encountered in a large referral Hospital in Tamil Nadu (south India) from December 1983 to May 1985 were characterised by conventional tests. Of these, 57 (95%) belonged to four species viz., Sanguis II, Mitis, Intermedius and Constellatus. Of the 40 strains that were assessed for their clinical significance, 23 (57.5%) were found to be either significant or of suggestive significance. The study suggested viridans streptococci are not particularly virulent pathogens. But local/systemic factors were found to predispose the subjects to this infection.

Humans↗

PubMed2000

Effect of an essential oil-containing antiseptic mouthrinse on plaque and salivary Streptococcus mutans levels.

BACKGROUND: Clinical studies in which antimicrobial mouthrinses were shown to have significant antiplaque activity most frequently have used gingivitis as the clinically relevant endpoint. However, there is evidence to suggest that mouthrinses containing active agents effective against Streptococcus mutans, such as chlorhexidine, may also have a role in inhibiting dental caries. This clinical study was conducted to determine the effect of 2x-daily rinsing with an essential oil-containing antiseptic mouthrinse (Listerine Antiseptic) on levels of recoverable S. mutans and total streptococci in supragingival interproximal plaque and in saliva. Additionally, a follow-up in vitro study is reported which determined whether a differential susceptibility to the antiseptic mouthrinse exists among different strains of streptococci. METHOD: Following baseline saliva and plaque sampling for quantification of recoverable S. mutans and total streptococci, 29 qualifying subjects were randomly assigned either the essential oil mouthrinse or a sterile water control. They rinsed with 20 ml for 30s 2 x daily for 11 days and once on the 12th day, in addition to their usual oral hygiene procedures. On day 12, saliva and plaque samples were again collected and microbiological quantification performed. The procedures were repeated with the alternate rinse after a 1-week washout period. RESULTS: The essential oil mouthrinse produced respective reductions of 69.9% and 75.4% in total recoverable streptococci and in S. mutans in plaque, and corresponding reductions of 50.8% and 39.2% in saliva. The in vitro study revealed that streptococci from the mutans group were more susceptible to the bactericidal activity of the essential oil mouthrinse than streptococci from the mitis group. CONCLUSIONS: As antimicrobial mouthrinses are most frequently recommended to patients whose mechanical oral hygiene procedures are not adequate for the control of supragingival plaque and gingivitis, this study provides an additional rationale for the inclusion of the essential-oil mouthrinse as an adjunct to daily oral hygiene procedures.

Adult↗

PubMed2005

Identification of non-mutans streptococci organisms in dental plaques recovering on mitis-salivarius bacitracin agar medium.

The objective of this study was to both isolate and identify non-mutans streptococci organisms (non-MSO) from dental plaques recovered on mitis-salivarius sucrose bacitracin agar (MSB) plates. The dental plaque samples, which had been collected from 63 human subjects, were diluted and plated on MSB. The bacteria growing on the MSB plates were then identified with biochemical tests, as well as with 16S rDNA cloning and sequencing techniques. Our data indicated that bacteria from 30 subjects had been recovered on the MSB plates. Among the 21 typical colonies selected from the 30 subjects, 12 colonies, derived from 10 subjects, were identified as non-MSO. These 12 colonies were determined to be Streptococcus anginosus (8 colonies), S. sanguinis (1 colony), and Pantoea agglomerans (3 colonies). These results strongly suggest that a new selective medium will be required for the reliable isolation of mutans streptococci.

Adolescent↗

PubMed1999

Antiphosphorylcholine antibody levels are elevated in humans with periodontal diseases.

Human immunoglobulin G2 (IgG2) serum concentrations and the IgG2 antibody response to Actinobacillus actinomycetemcomitans can be influenced by genes, by environmental factors such as smoking, and by periodontal disease status. Examination of the IgG2 response to phosphorylcholine (PC), a response thought to be mainly induced by the C polysaccharide of Streptococcus pneumoniae, suggested that periodontal disease status was also associated with this response. This prompted the hypothesis that PC is an important oral antigen associated with organisms in the periodontal flora and that anti-PC antibody is elevated as a consequence of periodontal disease. Subjects in various periodontal disease diagnostic categories in which attachment loss is exhibited were tested for anti-PC in serum. Those with adult periodontitis, localized juvenile periodontitis, generalized early-onset periodontitis, and gingival recession all had similar levels of anti-PC IgG2 serum antibody which were significantly greater than in the group of subjects with no attachment loss. Analysis of plaque samples from subgingival and supragingival sites in all diseases categories for reactivity with the anti-PC specific monoclonal antibody TEPC-15 revealed that a substantial proportion of the bacteria in dental plaque (30 to 40%) bear PC antigen; this antigen was not restricted to morphotypes resembling only cocci but was also present on rods and branched filamentous organisms. We found that S. mitis, S. oralis, and S. sanguis, as well as oral actinomycetes, including A. viscosus, A. odontolyticus, and A. israelii, incorporated substantial amounts of [(3)H]choline from culture media. Further analysis of antigens derived from these organisms by Western blot indicated that S. oralis, S. sanguis, A. viscosus, A. odontolyticus, and A. israelii contained TEPC-15-reactive antigens. The data show that many commonly occurring bacterial species found in dental plaque contain PC antigen and that immunization with plaque-derived PC antigens as a consequence of inflammation and periodontal attachment loss may influence systemic anti-PC antibody concentrations.

Actinomycetales↗

PubMed1993

Antibacterial activity of dentinal bonding agents.

The susceptibility of five bacterial species to seven dentinal bonding agents was examined in vitro. Agar diffusion tests using filterpaper disks containing 10 microL each of conditioner, primer, or resin were performed on blood agar and mitis salivarius bacitracin agar. Chlorhexidine (0.2%) was used as a positive control. After incubation, zones of inhibited bacterial growth were measured. Of all the compounds tested, Gluma cleanser and Gluma etchant showed the strongest growth inhibition for all bacterial strains. No antibacterial effect was noted for Prisma Universal Bond 2 and Superlux Universal Bond 2 systems. The primers of Gluma, Denthesive, and Scotchbond 2 displayed antibacterial activity that, in some cases, was comparable to that of 0.2% chlorhexidine. Zones of inhibition were seen for the resin materials of Scotchbond 2 and Tripton with Streptococcus mutans, Streptococcus sanguis and Actinomyces viscosus. No inhibition was seen after these resins were cured, whereas the antibacterial effect of XR-Bond on S sanguis and A viscosus was not affected by light curing.

Actinomyces viscosus↗