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Strains of Streptococcus mutans and Streptococcus sobrinus attach to different pellicle receptors.

We compared the levels of adsorption of Streptococcus mutans JBP and Streptococcus sobrinus 6715 to experimental pellicles formed from unsupplemented and glucosyltransferase (GTF)-supplemented saliva. Pellicles formed on hydroxyapatite beads from GTF or from saliva-GTF mixtures possessed detectable GTF activity. Low levels of GTF activity were also detected in clarified whole human saliva, but not in samples of submandibular saliva. The adsorptive behavior of S. mutans JBP to pellicles formed from saliva or saliva-GTF mixtures was strikingly different from that of S. sobrinus 6715. S. mutans JBP adsorbed in higher numbers to pellicles formed from whole or submandibular saliva than to buffer-treated hydroxyapatite under the assay conditions used, in which blocking with albumin was used. In contrast, S. sobrinus 6715 attached in lower numbers and did not show enhanced adsorption to pellicles prepared from saliva. Pellicles prepared from the high-molecular-weight mucin fraction of submandibular saliva effectively promoted adsorption of S. mutans JBP, but none of the saliva fractions tested enhanced the attachment of S. sobrinus 6715 above the levels of buffer controls. Exposure of pellicles which contained GTF to sucrose to permit in situ synthesis of glucan markedly enhanced attachment of S. sobrinus 6715 but not attachment of S. mutans JBP. Also, the presence of sucrose throughout the adsorption period did not enhance attachment of S. mutans JBP. Both organisms possessed cell-associated GTF, and GTF preparations derived from S. sobrinus 6715 and Streptococcus sanguis FC-1 behaved like GTF derived from S. mutans JBP. S. sobrinus 6715 attached in high numbers to dextran-treated hydroxyapatite, whereas S. mutans JBP did not. These observations suggest that S. mutans JBP cells possess an adhesin which binds to salivary components in the pellicles. In contrast, S. sobrinus 6715 cells appear to possess an adhesin which binds to glucan in the pellicles. Four additional strains of S. mutans and four additional strains of S. sobrinus behaved qualitatively like strains JBP and 6715, respectively, and thus the differences observed appear to be representative of these species. Collectively, our data indicate that S. mutans and S. sobrinus attach to different receptors in experimental pellicles.

Adhesiveness↗

Characterization of a P1-deficient strain of Streptococcus mutans that expresses the SpaA protein of Streptococcus sobrinus.

The Streptococcus sobrinus SpaA protein and the Streptococcus mutans P1 protein share 66% sequence homology at the amino acid level. To determine if the SpaA protein can be expressed in S. mutans and functionally replace the P1 protein, the spaA gene of S. sobrinus 6715 was isolated from plasmid pX1303 and inserted into the Escherichia coli-Streptococcus shuttle vector pVA838. The resulting plasmid pX1600 was transformed into the P1-deficient strain S. mutans 834 that has defects in saliva-mediated aggregation and in the ability to adhere to saliva-coated hydroxyapatite surfaces. Western blot (immunoblot) analysis of cellular protein fractions of S. mutans 834 (pX1600) detected in mutanolysin-solubilized cell walls a major protein of 210 kDa with an electrophoretic mobility similar to that of S. sobrinus SpaA protein and a minor 210-kDa protein and a major 64-kDa protein in the extracellular protein fraction. Analysis of virulence traits showed that expression of SpaA protein by S. mutans 834(pX1600) cells had restored the ability of the S. mutans 834 cells to aggregate in the presence of saliva or salivary agglutinin but not to adhere to saliva-coated hydroxyapatite. This cell aggregation was inhibited specifically by antisera to S. sobrinus SpaA protein. These results indicate that SpaA plays a role in the virulence of S. sobrinus by specifically interacting with fluid-phase salivary agglutinin to mediate cell aggregation.

Agglutinins↗

Co-induction of beta-galactosidase and the lactose-P-enolpyruvate phosphotransferase system in Streptococcus salivarius and Streptococcus mutans.

The addition of lactose, galactose, or isopropyl-beta-D-thiogalactoside (IPTG) to glucose-grown cells of Streptococcus salivarius 25975 resulted in the co-induction of both the lactose-P-enolpyruvate phosphotransferase system (lactose-PTS) and beta-galactosidase, with the latter the predominant metabolic system. With various strains of Streptococcus mutans and Streptococcus sanguis 10556, on the other hand, the lactose-PTS was the major metabolic pathway with beta-galactosidase induced either to low or negligible levels. In all cases, induction of the lactose-PTS resulted in the concomitant induction of 6-P-beta-galactosidase. The induction by lactose of both the lactose-PTS and beta-galactosidase in all strains was repressed by glucose and other catabolites, notably, fructose. Induction of beta-galactosidase in S. salivarius 25975 by IPTG was, however, relatively resistant to glucose repression. Induction experiments with IPTG and lactose suggested that a cellular metabolite of lactose metabolism was a repressor of enzyme activity. Exogenous cAMP was shown to reverse the transient repression by glucose of beta-galactosidase induction in cells of S. salivarius 25975 receiving lactose, provided the cells were grown with small amounts of toluene to overcome the permeability barrier to this nucleotide, cAMP, was however, unable to overcome the permanent repression of beta-galactosidase activity to a significant extent under these conditions.

Cyclic AMP↗

IDENTIFICATION OF STIMULATORY FACTOR INVOLVED IN SYMBIOTIC GROWTH OF STREPTOCOCCUS LACTIS AND STREPTOCOCCUS CREMORIS.

Dahiya, R. S. (North Carolina State College, Raleigh) and M. L. Speck. Identification of stimulatory factor involved in symbiotic growth of Streptococcus lactis and Streptococcus cremoris. J. Bacteriol. 85:585-589. 1963.-Single-strain cultures of Streptococcus lactis and S. cremoris isolated from a commercial starter culture showed symbiotic growth in milk. This study dealt with identification of the main component responsible for high acid production resulting from the combined growth of these cultures. Paper chromatography and bioautography were adapted to isolate the main stimulatory factor excreted by the slower-growing culture (S. lactis) in the culture media. It was identified as adenine on the basis of ultraviolet absorption spectra and paper chromatographic R(F) value. Pure adenine possessed the same properties and was also stimulatory to the faster-growing culture (S. cremoris) when added to milk.

Adenine↗

FATTY ACID COMPOSITION OF LIPIDS FROM STREPTOCOCCUS CREMORIS AND STREPTOCOCCUS LACTIS VAR. MALTIGENES.

MacLeod, Patricia (University of Connecticut, Storrs) and Joseph P. Brown. Fatty acid composition of lipids from Streptococcus cremoris and Streptococcus lactis var. maltigenes. J. Bacteriol. 85:1056-1060. 1963.-A method for the extraction of lipids from bacterial cells, involving grinding in a ball mill, is described. Cells of Streptococcus lactis var. maltigenes and S. cremoris yielded 5.3 and 5.5%, respectively, of their dry cellular weights as lipid. The lipids of both organisms were separated into three fractions by thin-layer chromatography. Two of these fractions have been classified: one fraction containing phospho- and perhaps glycolipids and the other containing triglycerides. The unidentified fraction was probably either mono- or diglyceride in nature. The fatty acid composition of the total lipid as well as that of each of the separated lipid components from the two lactic strains were compared.

Chemical Phenomena↗

A simple biochemical scheme for the differentiation of Streptococcus mutans and Streptococcus sobrinus.

A collection of wild-type and reference strains of Streptococcus mutans and Streptococcus sobrinus was compared using a set of fermentation and enzymic tests in order to establish a simple scheme for the differentiation of these two species isolated from selective media. Tests for the production of acid from N-acetylglucosamine, arbutin and melibiose, and the presence of alpha-galactosidase and alpha-glucosidase activities were useful in differentiating these species. Strains of Streptococcus mutans unable to ferment melibiose were also frequently unable to ferment other sugars; a consideration of the genetic control and organization of the genes involved in melibiose metabolism indicates that certain of the conventional characterization tests are not measurements of independent biochemical capabilities.

Adolescent↗

Bacterial interference of penicillin-sensitive and -resistant Streptococcus pneumoniae by Streptococcus oralis in an adenoid organ culture: implications for the treatment of recurrent upper respiratory tract infections in children and adults.

OBJECTIVES: The role of the viridans group of streptococci (Streptococcus oralis) in the prevention of colonization with Streptococcus pneumoniae was investigated in an adenoid organ culture system. METHODS: The adenoids from 10 patients who were undergoing adenoidectomy for either hypertrophy or recurrent otitis media were used. RESULTS: Streptococcus oralis Parker and S. oralis Booth (two organisms isolated from the nasopharynges of patients undergoing adenoidectomy only and patients undergoing adenoidectomy and bilateral tympanostomy with tubes, respectively) uniformly inhibited both penicillin-sensitive and penicillin-resistant S. pneumoniae. Although both strains of S. oralis inhibited the growth of both S. pneumoniae strains, strain Parker provided more complete inhibition than did strain Booth. CONCLUSIONS: The results indicate that some strains of S. oralis may inhibit the growth of the most serious pathogens in the nasopharynx. It is therefore possible that colonization of inhibitory strains of viridans streptococci may be used in the nasopharynx as a relatively safe and inexpensive approach to prevention of recurrent otitis media in some children and of recurrent suppurative sinusitis in both children and adults.

Adenoids↗

Membrane-associated and solubilized ATPases of Streptococcus mutans and Streptococcus sanguis.

The proton-translocating, membrane ATPases of oral streptococci have been implicated in cytoplasmic pH regulation, acidurance, and cariogenicity. Membranes were isolated from Streptococcus mutans GS-5 and Streptococcus sanguis NCTC 10904 following salt-induced lysis of cells treated with lysozyme and mutanolysin. The ATPase activities of these membranes were 1.8 and 1.1 units per mg membrane protein, respectively. F1 ATPases were washed free from the membranes and purified by fast protein liquid chromatography (FPLC). Hydrolytic activities of the F1 ATPases were maximal at pH values between 6.0 and 6.6, whereas the membrane-bound enzymes had pH maxima of 7.5 (S. sanguis) and 6.0 (S. mutans). The F1 ATPases of the streptococci were similar to the well-characterized enzyme of Escherichia coli; they consisted of five different polypeptides and had apparent, aggregate molecular weights of from 335 to 350 Kd. The membrane-bound ATPases were characterized biochemically and found to be similar to those of proton-translocating ATPases of E. coli and Streptococcus faecalis. Km values for the membranes with respect to ATP were found to be 0.9 and 1.0 mmol/L for S. mutans and S. sanguis, respectively. Both enzymes had specificities for purine triphosphates and were active with a variety of divalent cations, although optimal activity occurred with ATP and Mg. The membrane-associated enzymes were sensitive to the inhibitors dicyclohexylcarbodiimide (DCCD) and azide, but insensitive to ouabain and vanadate.(ABSTRACT TRUNCATED AT 250 WORDS)

Cell Membrane↗

Associations between salivary levels of Streptococcus mutans, Streptococcus sobrinus, lactobacilli, and caries experience in Kenyan adolescents.

Salivary levels of mutans streptococci (S. mutans and S. sobrinus) and lactobacilli were determined in a random sample of rural Kenyans between 15 and 19 years of age (n = 149). It is possible for the natural history of dental caries in this population to be studied since it is characterized by a limited access to conventional dental treatment. Using a short set of biochemical tests, we identified from seven to ten presumptive mutans streptococcus colonies--cultured from the saliva of each individual--to differentiate between S. mutans and S. sobrinus. No colonies resembling S. rattus (S. mutans serotype b) were isolated. Lactobacilli were identified as Gram-positive, catalase-negative rods. The mean D1-4MFS and D3-4MFS were 7.03 +/- 6.43 and 1.46 +/- 3.44, respectively. The mean mutans streptococcus and lactobacillus levels were 8.7 x 10(4) and 6.7 x 10(4), respectively. The salivary mutans streptococcus and lactobacillus levels were significantly correlated (p less than 0.01). Of the subjects, 64% harbored only S. mutans, 4% only S. sobrinus, 30% both species, and 2% neither. Lactobacilli were ubiquitous. The caries experience of the group was significantly (p less than 0.001) correlated with both the total salivary level of mutans streptococci and the salivary S. mutans levels, but not with the salivary S. sobrinus level.

Adolescent↗

Acute glomerulonephritis associated with streptococcus pyogenes with concomitant spread of streptococcus constellatus in four rural families.

We studied history, renal histopathology and microbiology of an epidemic of acute glomerulonephritis associated with throat infections and uncommon culture results in four neighbour families. A 40-year-old man (index patient) was referred to a university hospital for dialysis and kidney biopsy due to a suspected acute glomerulonephritis. An acute tonsillitis had preceded the condition. Penicillin treatment had been started four days before the discovery of renal failure. Throat swabs were positive for beta-hemolytic streptococci, group C (GCS). GCS were also found in throat cultures from his wife and two of their children. The bacteria were typed as Streptococcus constellatus. A third child had S. constellatus expressing Lancefield antigen group G. A neighbour and two of his children fell ill the following week with renal involvement. Throat swabs from both these children were positive for S. constellatus. His third child had erythema multiforme and S. constellatus in the throat while a fourth child had beta-hemolytic streptococci group A; Streptococcus pyogenes. Kidney biopsies on the index patient and his neighbour showed an acute diffuse prolipherative glomerulonephritis compatible with acute post-streptococcal nephritis and microbiological analysis of renal tissue revealed in both cases S. pyogenes and S. constellatus. The families had had much contact and had consumed unpasteurized milk from our index patient's farm. In four of seven persons in two additional neighbouring families S. constellatus was found in throat swabs during the same month while two persons carried Streptococcus anginosus expressing the Lancefield C antigen. In conclusion spread of S. constellatus coincided with the occurrence of four cases of acute glomerulonephritis. The two biopsied patients had both S. pyogenes and S. constellatus present in renal tissue. The epidemic either suggested that the outbreak of glomerulonephritis was due to S. pyogenes but coincided with the transmission and colonization of S. constellatus or that the S. constellatus strains were highly pathogenic or nephritogenic and that this organism can be transmitted in such cases.

Acute Disease↗

[MGB probes detect Streptococcus mutans and Streptococcus sobrinus in real time].

OBJECTIVE: To detect and distinguish Streptococcus mutans (S. mutans) and Streptococcus sobrinus (S. sobrinus) quickly in epidemiology and investigate the distribution of S. mutans in the oral of children with rampant caries. METHODS: Designed minor groove binder (MGB) probes according to the gtf gene of S. mutans and S. sobrinus. Detected 9 reference strains of Streptococcus mutans group by MGB probes in real time and after cultivation. Evaluated the results of these two methods. 92 dental plaques from pre-school children with rampant caries were detected in real time with MGB probes. RESULTS: The primers could amplify the target sequences specificity and distinguished S. mutans and S. sobrinus from each other using MGB probes. Though the fluorescence occurred earlier in S. mutans than in S. sobrinus, they had the same results in nature. In 92 children with rampant caries, the detective ratio of S. mutans was 96.7% and that of S. sobrinus was 32.6%. All the samples which could detect S. sobrinus were positive for S. mutans. CONCLUSION: The primers and probe designed from gtf genes of S. mutans and S. sobrinus can amplify the target sequence and distinguish them from each other in real time.

Child↗

Immunoelectrophoretic study of cell surface antigens from different Streptococcus mutans serotypes and Streptococcus sanguis.

Antigens prepared from culture supernatants or whole cells of several cariogenic strains were examined by immunoelectrophoresis for their crossed antigenicity, with reference to Streptococcus mutans OMZ175, serotype f. Crossed immunoelectrophoresis revealed a crossreactivity between soluble extracellular and wall associated antigens of six strains of Streptococcus mutans and one strain of Streptococcus sanguis. Protease destroyed the immunoreactivity of crossreactive antigens. One of them was shown to be localized on the bacterial surface.

Antigens, Surface↗

Plaque formation in vitro by Actinomyces viscosus in the presence of Streptococcus sanguis or Streptococcus mutans.

Actinomyces viscosus, growing on a tooth in the presence of sucrose, slowly produced a loosely-attached plaque, the pH being 6.1 after 120 h. When the tooth was inoculated simultaneously with A. viscosus and either Streptococcus sanguis or Streptococcus mutans, firmly-adherent plaque was quickly formed and the pH fell below 5 after 33 h with the former Streptococcus and 24 h with the latter. A. viscosus disappeared from each mixed plaque by 120 h.

Actinomyces↗

16S-23S and 23S-5S intergenic spacer regions of Streptococcus thermophilus and Streptococcus salivarius, primary and secondary structure.

The 16S-23S intergenic spacer region (spacer region 1) of Streptococcus salivarius, S. thermophilus, and Lactococcus lactis subsp. cremoris and the 23S-5S intergenic spacer region (spacer region 2) of S. salivarius and L. lactis subsp. cremoris were sequenced and compared with the spacer regions 1 and 2 of other streptococci. A high degree of intraspecific conservation was observed for S. thermophilus and L. lactis, and very similar sequences were found for S. salivarius and S. thermophilus. Whereas spacer region 1 is highly conserved in the genus Streptococcus sensu-stricto, only the tRNA gene and the rRNA processing stems are highly conserved in the three genera: Streptococcus sensu-stricto, Lactococcus, and Enterococcus. The presence of a unique tRNA(Ala) gene without the 3' terminal CCA sequence seems to be a general feature of the streptococci spacer region 1. A secondary structure model was built to show the interaction between the spacer regions 1 and 2 of S. thermophilus and S. salivarius. The rapid evolution of spacer region 1 in streptococci is in part due to insertions and deletions of small RNA stem/loop structures.

Base Sequence↗

Modified latex agglutination test for rapid detection of Streptococcus pneumoniae and haemophilus influenzae in cerebrospinal fluid and direct serotyping of Streptococcus pneumoniae.

A modified latex agglutination test was designed and evaluated for the rapid detection of Streptococcus pneumoniae and haemophilus influenzae type b capsular antigens, and for direct serotyping of Streptococcus pneumoniae in the cerebrospinal fluid. Reagents were prepared by sensitizing latex particles with Omniserum (against 83 capsular serotypes of pneumococci) and Haemophilus influenzae type b burro antiserum. For serotyping reagents, latex particles were similarly coated with nine pneumococcal pool (a to I) antisera and 46 individual pneumococcal serogroup/serotype specific antisera. The test was performed on cerebrospinal fluid from 298 patients with suspected meningitis. Serotyping was done directly on untreated cerebrospinal fluid samples showing positive reactions with the Omniserum reagent. Pneumococcal or Haemophilus influenzae type b antigens were detected in 41 patients; in 32 of these the etiology was established by culture and in 2 by smear examination. Five of the remaining seven cases were judged clinically and by cytological examination of cerebrospinal fluid to have partially treated bacterial meningitis. In two cases the test was false positive. The overall sensitivity and specificity of the latex agglutination test for the detection of Streptococcus pneumoniae and Haemophilus influenzae type b antigens was 100% and 96.8% respectively. The commonest pneumococcal serotypes were type 1 (30%), types 6 and 19 (10% each). The latex agglutination test is rapid and simple to perform, yielding serotype data directly by testing of cerebrospinal fluid.

Adult↗

Characterization of fluoroquinolone-resistant beta-hemolytic Streptococcus spp. isolated in North America and Europe including the first report of fluoroquinolone-resistant Streptococcus dysgalactiae subspecies equisimilis: report from the SENTRY Antimicrobial Surveillance Program (1997-2004).

Beta-hemolytic streptococci are common bacterial pathogens that can cause serious invasive disease, and although this group of species has remained susceptible to the fluoroquinolone class, resistant strains have been reported. This multicenter investigation determined the rate of fluoroquinolone-resistant beta-hemolytic streptococci using the SENTRY Antimicrobial Surveillance Program network data (1997-2004). Forty-seven surveillance culture isolates of beta-hemolytic streptococci from North America and Europe with elevated levofloxacin MIC results (2 to >32) microg/mL were tested for susceptibility to other fluoroquinolones including ciprofloxacin, garenoxacin, gatifloxacin, gemifloxacin, and moxifloxacin using reference broth microdilution and Etest (BIODISK, Solna, Sweden) methods. Strains were characterized using polymerase chain reaction and sequencing to detect mutations in the quinolone-resistance determining region (QRDR). The beta-hemolytic streptococci isolates with reduced fluoroquinolone susceptibility included the following Lancefield groups: A (Streptococcus pyogenes; 9 strains), B (Streptococcus agalactiae; 24 strains), C and G (14 strains). Vitek and API 20 strep (bioMerieux, Hazelwood, MO) identification systems, as well as conventional biochemical methods and colony morphology, were used to confirm the organism identifications. The overall potency (MIC90 in microg/mL) for the fluoroquinolones against all tested beta-hemolytic streptococci showed the following rank order: gemifloxacin (0.06) > garenoxacin (0.12) > moxifloxacin (0.25) > gatifloxacin (0.5) > levofloxacin = ciprofloxacin (1). The rate of levofloxacin-resistant beta-hemolytic streptococci in the SENTRY program was 0.14% (Europe) and 0.51% (North America) during the study period. All levofloxacin-resistant strains tested by molecular methods had significant mutations in either parC (position 79 or 83) and/or gyrA (position 81 or 85). All but 2 isolates with high-level resistance to levofloxacin (>32 microg/mL) had gyrA mutations. Strains with lower MIC values to levofloxacin (2-4 microg/mL) only had mutations in parC. The increasing rate of fluoroquinolone-resistant streptococci including Streptococcus pneumoniae, viridans group streptococci, and the more recently reported beta-hemolytic streptococci, is becoming a clinical concern due to the morbidity and mortality caused by these pathogens.

Adolescent↗

Unique poly(glycerophosphate) lipoteichoic acid and the glycolipids of a Streptococcus sp. closely related to Streptococcus pneumoniae.

The Streptococcus sp. studied here is closely related to Streptococcus pneumoniae with 98.6% 16S rRNA similarity and 65% DNA/DNA homology. We isolated the lipoteichoic acid and the membrane glycolipids whose structures were established using conventional procedures and NMR spectroscopy. The lipoteichoic acid contains a linear 1,3-linked poly(glycerophosphate) chain which is partly substituted with D-alanine ester and is phosphodiester-linked to O6 of beta-D-Galf(1-->3)acyl2Gro. This lipoteichoic acid is the first example in which a monohexosylglycerol serves as the glycolipid anchor; and with an average chain length of 10 glycerophosphate residues it is the shortest known to date. MS analysis, applied for the first time to a native acylated lipoteichoic acid, revealed a continuous increase in chain length from seven to 17 glycerophosphate residues with a maximum at 10, and allowed identification of the fatty acid combinations. Membrane glycolipids consisted of beta-D-Galf(1-->3)acyl2Gro (9%), alpha-D-Glcp(1-->3)acyl2Gro (22%), alpha-D-Galp(1-->2)-alpha-D-Glcp(1-->3)acyl2Gro (64%) and alpha-D-Galp(1-->2)-(6-O-acyl)-alpha-D-Glcp(1-->3)acyl2Gro (5%). It is noteworthy that in lipoteichoic acid biosynthesis, Galfacyl2Gro, a less abundant membrane glycolipid, is selected as the lipid anchor. Despite the genetic relatedness to Streptococcus pneumoniae, the lipoteichoic acid structure is quite different to the complex structure of pneumococcal lipoteichoic acid [T. Behr et al. (1992) Eur. J. Biochem. 207, 1063-1075], thus providing an example that minor differences in DNA sequence exert major changes in macromolecular structure.

Chromatography, High Pressure Liquid↗

Acute pansinusitis with bacteremia due to a beta-hemolytic group C streptococcus: Streptococcus milleri.

A 41-year-old truck driver had acute onset of weakness, severe headache and pain over the left side of the face, forehead, and orbital area. He was found to have acute pansinusitis. Blood cultures and culture of the sinus drainage yielded beta-hemolytic group C streptococcus: Streptococcus milleri. He recovered completely after treatment with cefazolin, surgical drainage and debridement, and outpatient cephalexin therapy. Beta-hemolytic streptococci are uncommon causes, and bacteremia is rare in acute sinusitis. Speciation of the streptococcus is important in determining the epidemiology and clinical spectrum of streptococcal infections.

Acute Disease↗