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Phenotypic and molecular characterization of macrolide and streptogramin resistance in Streptococcus mitis from neutropenic patients.

OBJECTIVES: To determine the prevalence of macrolide and streptogramin resistance in Streptococcus mitis isolates from neutropenic patients and to identify mechanisms of macrolide and streptogramin resistance in resistant isolates. METHODS: MICs of erythromycin, spiramycin, lincomycin and pristinamycin were determined for S. mitis isolates. Macrolide-resistance genes were characterized by PCR and ribosomal mutations by sequencing. RESULTS: A total of 169 S. mitis isolates were recovered from 66 patients at the Tunisian Bone Marrow Transplant Centre. Of these, 120 (70%) were non-susceptible to erythromycin and one was resistant to pristinamycin; 48.5% of isolates had an MLSB phenotype with cross-resistance between erythromycin, spiramycin and lincomycin, 4% had a dissociated MLSB phenotype with resistance to erythromycin and spiramycin but apparent susceptibility to lincomycin and 47.5% displayed the M phenotype. Resistance determinants were characterized in 33 isolates. Ten of 14 isolates with the cross MLSB resistance contained an ermB-like gene and four a combination of ermB- and mefA-like genes. Four of the five isolates with a dissociated MLSB phenotype contained ermB-like and one a combination of ermB- and mefA-like genes. All the 14 isolates with an M phenotype contained mefA-like genes. The pristinamycin-resistant strain had G105 and A108 substitutions in the conserved C terminus of the L22 ribosomal protein. CONCLUSIONS: The prevalence of macrolide resistance is high in S. mitis from neutropenic patients and is due to the spread of ermB- or mefA-like genes alone or combined. Resistance to streptogramins is rare and in this case associated with ribosomal mutation.

Amino Acid Sequence↗

A case of Streptococcus mitis endocarditis successfully treated by linezolid.

We report the successful treatment of infective endocarditis caused by Streptococcus mitis with linezolid in a patient with pre-existing valvular heart disease. The patient had multiple allergies to conventional antibiotics. Linezolid may provide an oral alternative in the treatment of infective endocarditis in patients with adverse drug reactions to traditional antibiotic regimens.

Acetamides↗

Molecular analysis of age-related changes of Streptococcus anginosus group and Streptococcus mitis in saliva.

The purpose of this study was to survey the prevalence of streptococcal species, especially Streptococcus anginosus (which has been reported to be associated with cancer in the upper digestive tract), Streptococcus constellatus, and Streptococcus intermedius in the saliva of different age groups. A sequence analysis of 16S rDNA was performed and DNA quantified using real-time polymerase chain reaction. The S. anginosus level increased with age, whereas the levels of S. constellatus and S. intermedius did not change. Streptococcus mitis was the predominant species in the saliva of all the age groups but, unlike the S. anginosus, the proportion of S. mitis in the salivary bacteria decreased with age. The increase in S. anginosus with age should be carefully monitored because of its association with diseases, including cancer.

Adult↗

Clonal diversity and turnover of Streptococcus mitis bv. 1 on shedding and nonshedding oral surfaces of human infants during the first year of life.

Streptococcus mitis bv. 1 is a pioneer colonizer of the human oral cavity. Studies of its population dynamics within parents and their infants and within neonates have shown extensive diversity within and between subjects. We examined the genetic diversity and clonal turnover of S. mitis bv. 1 isolated from the cheeks, tongue, and primary incisors of four infants from birth to 1 year of age. In addition, we compared the clonotypes of S. mitis bv. 1 isolated from their mothers' saliva collected in parallel to determine whether the mother was the origin of the clones colonizing her infant. Of 859 isolates obtained from the infants, 568 were unique clones. Each of the surfaces examined, whether shedding or nonshedding, displayed the same degree of diversity. Among the four infants it was rare to detect the same clone colonizing more than one surface at a given visit. There was little evidence for persistence of clones, but when clones were isolated on multiple visits they were not always found on the same surface. A similar degree of clonal diversity of S. mitis bv. 1 was observed in the mothers' saliva as in their infants' mouths. Clones common to both infant and mothers' saliva were found infrequently suggesting that this is not the origin of the infants' clones. It is unclear whether mucosal immunity exerts the environmental pressure driving the genetic diversity and clonal turnover of S. mitis bv. 1, which may be mechanisms employed by this bacterium to evade immune elimination.

Adult↗

Streptococcus mitis. A cause of serious infection in adults.

Twenty strains of Streptococcus mitis were isolated from blood or body fluids at the Cleveland Veterans Administration Medical Center from Jan 1, 1981, to April 30, 1984. Fifteen (75%) isolates were considered contaminants. Five (25%) were clinically important and associated with a serious infection of the oropharynx or gastrointestinal tract (three of five), endovascular system (one of five), or a prosthetic hip. Four of five patients required surgical intervention for treatment. Two of five died; one death was directly attributable to S mitis infection. Eighteen strains were available for detailed bacteriologic study. Three strains had a minimum inhibitory concentration of greater than 0.1 micrograms/mL of penicillin and six other strains were tolerant to penicillin. This review suggests that S mitis can be an important pathogen in adults and may cause infections other than endocarditis.

Adult↗

Inhibitory activity of Streptococcus mitis against oral bacteria.

The antagonistic properties of three strains of Streptococcus mitis were investigated. They were found to inhibit a wide range of oral bacteria; Gram-positive and Gram-negative, facultative and anaerobic species being susceptible. The S. mitis strains were shown to be producing hydrogen peroxide, this being partially responsible for the aerobic inhibitory activity. A second inhibitory factor(s) was also produced, aerobically and anaerobically, although this could not be isolated. A limited characterization of this factor was undertaken using plate cultures.

Actinomycetaceae↗

Inhibitory effects of human salivary histatins and lysozyme on coaggregation between Porphyromonas gingivalis and Streptococcus mitis.

The effects of histatins on coaggregation between Porphyromonas gingivalis 381 and Streptococcus mitis ATCC 9811 were investigated by using a turbidimetric assay. The coaggregation activity was significantly inhibited by histatins 5 and 8 and strongly by lysozyme. Tritium-labeled histatin 8 bound to P. gingivalis cells but not to S. mitis cells.

Amino Acid Sequence↗

Identification of streptococcus mitis group species by RFLP of the PCR-amplified 16S-23S rDNA intergenic spacer.

Mitis group streptococci are pioneer colonizers of tooth surfaces and are implicated in various pathologies. Thus, accurate identification of oral mitis group strains would be valuable for studies of plaque ecology and dental caries and for diagnostic use in endocarditis or sepsis patients. The aim of this study was to evaluate the usefulness of PCR-RFLP analysis of the 16S-23S intergenic spacer for differentiating and identifying streptococcus mitis group species. The 16S-23S rDNA spacer regions of 27 type and reference Streptococcus strains, representing 8 species, were studied by PCR-mediated amplification by using oligonucleotide primers FGPS 1490-72 and FGPL 132'-38. PCR products were digested, independently, with 14 restriction enzymes. Only AluI, MboI, CfoI, HinfI and MaeII distinguished some species, particularly AluI and CfoI, but not all the species. Eight clusters were clearly generated, corresponding to currently recognized species, but only with the addition of five ITS restriction patterns, generated by AluI + MboI + CfoI + HinfI + MaeII, then clustered by UPGMA, on a distance consensus matrix. The combination of these five ITS RFLP tests allowed a relatively conclusive genomic group differentiation of mitis group species. Despite this observation, more strains of each species will need to be analyzed, particularly clinical isolates, before arriving at general conclusions about the utility of ITS restrictions for identification of strains at the species level. An ITS PCR-RFLP-based identifying method for streptococcus mitis group species would provide significant advantages over other molecular taxonomic methods which require DNA extraction and DNA-DNA hybridization.

DNA, Ribosomal Spacer↗

Structures of two cell wall-associated polysaccharides of a Streptococcus mitis biovar 1 strain. A unique teichoic acid-like polysaccharide and the group O antigen which is a C-polysaccharide in common with pneumococci.

The cell wall of Streptococcus mitis biovar 1 strain SK137 contains the C-polysaccharide known as the common antigen of a closely related species Streptococcus pneumoniae, and a teichoic acid-like polysaccharide with a unique structure. The two polysaccharides are different entities and could be partially separated by gel chromatography. The structures of the two polysaccharides were determined by chemical methods and by NMR spectroscopy. The teichoic acid-like polymer has a heptasaccharide phosphate repeating unit with the following structure: The structure neither contains ribitol nor glycerol phosphate as classical teichoic acids do, thus we have used the expression teichoic acid-like for this polysaccharide. The following structure of the C-polysaccharide repeating unit was established: where AAT is 2-acetamido-4-amino-2,4, 6-trideoxy-D-galactose. It has a carbohydrate backbone identical to that of one of the two structures of C-polysaccharide previously identified in S. pneumoniae. C-polysaccharide of S. mitis is characterized by the presence, in each repeating unit, of two residues of phosphocholine and both galactosamine residues in the N-acetylated form. Immunochemical analysis showed that C-polysaccharide constitutes the Lancefield group O antigen. Studies using mAbs directed against the backbone and against the phosphocholine moiety of the C-polysaccharide revealed several different patterns of these epitopes among 95 S. mitis and Streptococcus oralis strains tested and the exclusive presence of the group O antigen in the majority of S. mitis biovar 1 strains.

Carbohydrate Conformation↗

Development of aggregating ability in cells of Streptococcus mitis ATCC 903 grown under glucose-limiting conditions in continuous culture.

Streptococcus mitis ATCC 903 grown under glucose-limiting conditions in continuous culture did not aggregate upon incubation in 10 mM phosphate buffer at pH 5--7 unless a metabolizable sugar was added. Aggregation started 45--60 min after the addition of glucose or sucrose whereas slowly metabolized sugars as galactose and lactose required several hours to cause aggregation. Active metabolism of the carbohydrate was a prerequisite for aggregation as indicated by acid formation. Chloramphenicol inhibited the development of aggregating ability in the presence of glucose or sucrose. The addition of a source of nitrogen (peptides and amino acids) enhanced aggregation and shortened the time for development of aggregating ability. No aggregation occurred at at 0 degrees C and the ability to aggregate was markedly delayed at 20 degrees C as compared to 30 degrees C and 37 degrees C. Trypsin treatment of the bacteria abolished aggregation, indicating that surface components of protein or glycoprotein nature contributed to the capacity to aggregate.

Carbohydrate Metabolism↗

Endocarditis caused by penicillin-resistant Streptococcus mitis in a 12-year-old boy.

A case of congenital coronary arteriovenous fistula with infective endocarditis caused by penicillin-resistant Streptococcus mitis is reported. Lack of prophylactic antibiotics during dental procedure may cause the development of endocarditis. Bactericidal test (Schlichter test) was performed to guide the therapy for this case of bacterial endocarditis caused by penicillin-resistant viridans streptococci. This case highlights the importance of antibiotic prophylaxis in patients with underlying heart disease undergoing dental procedures.

Ampicillin↗

Characterization of LytA-like N-acetylmuramoyl-L-alanine amidases from two new Streptococcus mitis bacteriophages provides insights into the properties of the major pneumococcal autolysin.

Two new temperate bacteriophages exhibiting a Myoviridae (phiB6) and a Siphoviridae (phiHER) morphology have been isolated from Streptococcus mitis strains B6 and HER 1055, respectively, and partially characterized. The lytic phage genes were overexpressed in Escherichia coli, and their encoded proteins were purified. The lytAHER and lytAB6 genes are very similar (87% identity) and appeared to belong to the group of the so-called typical LytA amidases (atypical LytA displays a characteristic two-amino-acid deletion signature). although they exhibited several differential biochemical properties with respect to the pneumococcal LytA, e.g., they were inhibited in vitro by sodium deoxycholate and showed a more acidic pH for optimal activity. However, and in sharp contrast with the pneumococcal LytA, a short dialysis of LytAHER or LytAB6 resulted in reversible deconversion to the low-activity state (E-form) of the fully active phage amidases (C-form). Comparison of the amino acid sequences of LytAHER and LytAB6 with that of the pneumococcal amidase suggested that Val317 might be responsible for at least some of the peculiar properties of S. mitis phage enzymes. Site-directed mutagenesis that changed Val317 in the pneumococcal LytA amidase to a Thr residue (characteristic of LytAB6 and LytAHER) produced a fully active pneumococcal enzyme that differs from the parental one only in that the mutant amidase can reversibly recover the low-activity E-form upon dialysis. This is the first report showing that a single amino acid residue is involved in the conversion process of the major S. pneumoniae autolysin. Our results also showed that some lysogenic S. mitis strains possess a lytA-like gene, something that was previously thought to be exclusive to Streptococcus pneumoniae. Moreover, the newly discovered phage lysins constitute a missing link between the typical and atypical pneumococcal amidases known previously.

Amino Acid Sequence↗

Neonatal meningitis due to Streptococcus mitis.

A baby girl became ill 2 days after a normal delivery. Streptococcus mitis was isolated from the cerebrospinal fluid, and she recovered uneventfully on treatment with penicillin and, for the first 2 days, netilmicin. The organism was penicillin-tolerant in vitro. Non-groupable alpha-hemolytic streptococci are an uncommon cause of meningitis; three cases have been ascribed to S. mitis.

Adult↗

Aggregation of enzymatically modified Streptococcus mitis indicating involvement of lectin-ligand type interaction.

The aggregation properties of Streptococcus mitis ATCC 903 cells modified by treatment with heat or different enzymes was investigated. Bacteria that had the ability to aggregate spontaneously lost this capacity by treatment with proteolytic enzymes, beta-galactosidase or heat. Cells subjected to different types of modification were mixed in various proportions and their aggregation properties were recorded. To discriminate between the two kinds of cells in the suspension, one partner in the aggregation reaction was labelled with 14C-palmitic acid. Bacteria treated with beta-galactosidase co-aggregated with spontaneously aggregating cells (not modified) and with cells treated with heat. Heat-treated cells co-aggregated with spontaneously aggregating cells and with cells treated with beta galactosidase. Cells treated with trypsin did not co-aggregate either with spontaneously aggregating cells or cells treated with heat or beta-galactosidase. These findings are consistent with the hypothesis that two surface components are required for specific aggregation of S. mitis cells. We suggest that both components are degraded or released from the bacterial surface by treatment with trypsin (and other proteolytic enzymes) as shown by the inability of these cells to take part in any co-aggregation with spontaneously aggregating cells. Treatment with beta-galactosidase degrades a carbohydrate receptor constituting the terminal part of a glycoprotein. Heat treatment inactivates a protein lectin. The fact that heat-treated bacteria and bacteria treated with beta-galactosidase aggregate when mixed supports the assumption that two components take part in the aggregation reaction.

Dental Plaque↗

Regulation of synthesis of beta-fructofuranosidase (invertase) in Streptococcus mitis.

beta-Fructofuranosidase activity was found to be cell-bound in Streptococcus mitis ATCC 903. The following evidence suggests that induction functions as a regulatory mechanism for beta-fructofuranosidase in S. mitis: (1) on transfer of glucose-grown exponential phase bacteria to sucrose medium, the specific activity of beta-fructofuranosidase increased fourfold in the course of one generation; (2) other sugars had no stimulatory effect on the rate of synthesis of beta-fructofuranosidase; (3) the effect of sucrose on the rate of synthesis of beta-fructofuranosidase could be measured within a few minutes. Glucose, fructose and mannose repressed beta-fructofuranosidase. The addition of glucose to bacteria growing on sucrose repressed beta-fructofuranosidase for about one generation. The intracellular concentration of glucose was considerably increased during repression, while the intracellular concentration of glycolytic intermediates did not vary significantly.

Carbohydrate Metabolism↗

Detection of penicillin-binding protein 2b gene alteration in Streptococcus mitis by polymerase chain reaction.

Three isolates of beta-lactam-resistant streptococci from the saliva of healthy adults were identified as Streptococcus mitis. Minimum inhibitory concentrations (MICs) were 2 to 4 micro g/ml for ampicillin (ABPC) and 64 to more than 128 micro g/ml for cefaclor (CCL). To determine the position of base alterations of the penicillin-binding protein 2b ( pbp2b) gene, upstream primers containing possible mutation points were designed, and used for polymerase chain reaction (PCR), together with a downstream primer. Alterations adjacent to the conserved motifs of the pbp2b gene were apparent. DNA sequencing data indicated replacements in deduced amino acid sequences in all resistant isolates: from threonine to alanine just after the serine-serine-asparagine (SSN) motif, and from alanine to glycine two residues downstream of the lysine-threonine-glycine (KTG) motif. These changes were the same as those in penicillin-resistant Streptococcus pneumoniae (PRSP), suggesting importance for the enzymatic activity of the protein. Thus, Beta-lactam susceptibility of S. mitis may be partially predicted by PCR using our primer set for pbp2b.

Aminoacyltransferases↗

[Streptococcus mitis endocarditis. Description of a clinical case].

The diagnostic iter of a case of Streptococcus mitis endocarditis is reported. Bacterial endocarditis was diagnosed in a 32-year-old patient following ultrasound cardiography and microbiological tests. The paper stresses the importance of ultrasound cardiography in the diagnosis of bacterial endocarditis.

Adult↗