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Phylogenetic evidence for the transfer of Eubacterium suis to the genus Actinomyces as Actinomyces suis comb. nov.

The 16S rRNA primary structures of Eubacterium suis DSM 20639T (T = type strain) and Bifidobacterium bifidum DSM 20456T were determined by sequencing in vitro amplified rDNA. Sequence comparisons indicated that B. bifidum is moderately related to representatives of the genera Actinomyces and Mobiluncus. The closest relative of E. suis is Actinomyces pyogenes. E. suis and A. pyogenes are more closely related phylogenetically to one another than to the other Actinomyces species that have been investigated by using comparative 16S rRNA analysis. Therefore, we propose that E. suis should be transferred to the genus Actinomyces as Actinomyces suis comb. nov.

Actinomyces

Differential medium for detecting dental plaque bacteria resembling Actinomyces viscosus and Actinomyces naeslundii.

A medium for detecting colonies of Actinomyces viscosus and Actinomyces naeslundii in dental plaque samples was developed. The medium (CNAC-20) contains 20.0 mug of 3CdSO4-8H2O per ml of Columbia CNA agar base. Laboratory strains of A. viscosus grew on CNAC-20 in characteristic round, white, smooth, opaque colonies. Increasing the cadmium concentration impaired the growth of some A. viscosus strains. Stock strains of A. naeslundii and A. israelii grew in colonies of similar white, opaque morphology. The few strains of other gram-positive plaque bacteria that grew on CNAC-20 had colonies easily distinguished from those of A. viscosus. Most of the bacterial strains freshly isolated from Actinomyces-like colonies on CNAC-20 that had been inoculated with human dental plaque samples were found to have cultural characteristics consistent with previous descriptions of A. viscosus or A. naeslundii. CNAC-20 may facilitate investigations into the relationship of microaerophilic Actinomyces with the etiology of dental diseases.

Actinomyces

New medium for isolation of Actinomyces viscosus and Actinomyces naeslundii from dental plaque.

Metronidazole (10 microgram/ml) and cadmium sulfate (20 microgram/ml) were added to a gelatin-based medium to select for microaerophilic Actinomyces species from dental plaque samples. The new medium (GMC), when incubated anaerobically, allowed 98% recovery of seven pure cultures of Actinomyces viscosus and 73% recovery of eight pure cultures of Actinomyces naeslundii, while suppressing 76% of the total count of other organisms in dental plaque samples. In 203 plaque samples, recoveries of A. viscosus and A. naeslundii on GMC and another selective medium for oral Actinomyces (CNAC-20) were compared. Recovery of A. viscosus was comparable on the two media. Recovery of A. naeslundii was significantly higher on GMC than CNAC-20 (P is less than 0.001), and GMC allowed a more characteristic cell morphology of both organisms. GMC medium appears to be useful for the isolation and presumptive identification of A. viscosus and A. naeslundii from dental plaque.

Actinomyces

Mannose-contaminating agglutinin for Actinomyces viscosus and Actinomyces naeslundii.

Rapid agglutination of Actinomyces viscosus and Actinomyces naeslundii cells by D-mannose solutions was observed during studies of their attachment to mammalian cells in vitro. The specificity of the agglutination reaction was studied by slide agglutination tests and by measuring the rate of decrease in optical density of bacterial phosphate buffer suspensions caused by the setting of bacterial aggregates. Actinomyces cells were agglutinated by protein-containing mannose solutions of several chemical suppliers. Solutions of sugars other than D-mannose and solutions of mannitol and mannan all failed to agglutinate A. viscsus and A. naeslundii. "Mannose-enhanced" agglutination was impaired by boiling or autoclaving the mannose but was not affected by heating the bacteria, the presence of chloramphenicol, running the assay in the cold, or incorporating any of several commercially purchased sugars in the reaction mixture. During these hapten inhibition experiments, only 6-deoxy-L-talcose-containing extracts of an A. viscosus strain retarded the rate of mannose-enhanced agglutination. Protein-containing fractions of D-mannose mother liquors also agglutinated cells of A. viscosus and A. naeslundii. Other species of oral gram-positive rods were not agglutinated by mannose solutions. Together the data indicate that plant seed-derived D-mannose contains a protein-associated agglutinin for A. viscosus and A. naeslundii which may function via a "lectin-like" selective affinity for the unique cell wall sugar 6-deoxy-L-talose.

Actinomyces

Conservation of an Actinomyces viscosus T14V type 1 fimbrial subunit homolog among divergent groups of Actinomyces spp.

The type 1 fimbrial subunit gene of the human Actinomyces viscosus T14V was used as a DNA probe in Southern analyses to detect related DNA sequences in 16 of 30 strains of Actinomyces spp. under conditions of high stringency. The organisms with homology to the DNA probe included two human and six nonhuman A. viscosus, three human and three nonhuman A. naeslundii, and two A. bovis isolates. Homologous DNA sequences were not detected in strains of A. odontolyticus and A. israelii examined in this study. Northern (RNA) blot analysis revealed expression of a transcript from each of the A. viscosus and A. naeslundii strains and from one A. bovis strain that was comparable in size to that detected from A. viscosus T14V. Cell surface fimbriae were observed on a majority of the strains that expressed the transcript. Various degrees of cross-immunoreactivities between these strains and antibodies specific for type 1 fimbriae of A. viscosus T14V were also observed by colony immunoassay. Thus, the data clearly demonstrate the existence in, and expression by, divergent Actinomyces groups of genomic sequences that are closely related to the type 1 fimbriae of A. viscosus T14V.

Actinomyces

Establishment and distribution of Actinomyces viscosus and Actinomyces naeslundii in the human oral cavity.

The intraoral establishment and proportional distribution of suspected periodontal pathogens Actinomyces viscosus and Actinomyces naeslundii were studied using a recently developed differential plating medium, CNAC-20. Saliva and dental plaque samples were collected from 108 subjects ranging in age from infants to young adults; tongue and buccal mucosa samples were collected from only the adult subjects. Catalase-negative A. naeslundii was isolated from 40% of the predentate infants' and almost all other subjects' saliva samples. It predominated among CNAC-20 isolates in the saliva of subjects of all age groups, in the plaques of young children, and in the adult tongue samples. In contrast, catalase-positive A. viscosus was not isolated from predentate infant samples, and its frequency of isolation increased slowly with age (greater than 50% detection by age 7). A. viscosus was isolated in highest relative proportions from dental plaque and buccal mucosa samples. The two closely related species A. viscosus and A. naeslundii apparently differ in respect to factors determining the host age at which they colonize and their relative intraoral distribution in humans.

Actinomyces

[Population composition of Actinomyces tumemacerans and Actinomyces albus var. fungatus].

The population analysis of Actinomyces tumemacerans and Actinomyces albus var. fungatus has revealed identical homologous series of spontaneous variants, this suggesting the genetical relationship of the cultures. Variants of the same type (basic, oligosporous, asporogenous and proactinomycete-like) are identical not only according to their individual properties (Kuznetsov, 1973) but also in total characteristics typical of the variants of a given species. Populations of the studied cultures comprise variants synthesizing several antibiotics, i.e. albofungin, albonursin and tetraene (basic and oligosporous variants), albofungin and tetraene (asprogenous green variants), as well as variants producing only one tetraene antibiotic (proactinomycete-like variants). Therefore, the population analysis based on the law of homologous series in hereditary variability of actinomycetes can be used as a new approach to the taxonomy of these microorganisms.

Anti-Bacterial Agents

Specificity of coaggregation reactions between human oral streptococci and strains of Actinomyces viscosus or Actinomyces naeslundii.

Coaggregation reactions between actinomycete and streptococcal cells occurred frequently when human strains of Actinomyces viscosus or A. naeslundii were mixed with human isolates of Streptococcus sanguis or S. mitis, but were infrequent with other oral actinomycetes and streptococci. Two groups of actinomycetes and four groups of streptococci were defined by the patterns of their coaggregation reactions and by the ability of beta-linked galactosides (i.e., lactose) to reverse these reactions. Coaggregations occurred by one of the following three kinds to cell-cell interactions: (i) coaggregation that was blocked by heating the streptococcus but not the actinomycete and was not reversed by lactose; (ii) coaggregation that was blocked by heating the actinomycete but not the streptococcus and was reversed by lactose; and (iii) coaggregation that was blocked only by heating both cell types. The latter reaction was a combination of the first two since lactose reversed coaggregation between heated streptococci and unheated actinomycetes but did not reverse coaggregations between unheated streptococci and heated actinomycetes. Cells that could be heat inactivated also were inactivated by amino group acetylation or protease digestion, whereas cells that were unaffected by heat were not inactivated by these treatments. Coaggregation reactions of each kind were Ca2+ dependent and insensitive to dextranase treatment. These findings are consistent with the hypothesis that human strains of A. viscosus and A. naeslundii coaggregate with strains of S. sanguis and S. mitis by a system of specific cell surface interactions between protein or glycoprotein receptors on one cell type and carbohydrates on the other type.

Acetylation

Immune labeling of certain strains of Actinomyces naeslundii and Actinomyces viscosus by fluorescence and electron microscopy.

A total of 12 well-characterized strains of Actinomyces viscosus and A. naeslundii grown on Trypticase soy agar plates supplemented with sheep erythrocytes were examined by light microscopy and transmission electron microscopy after treatment with appropriately labeled antisera to homologous and heterologous strains. Cells incubated with homologous rabbit antisera followed by fluorescein-isothiocyanate (FITC)-conjugated goat anti-rabbit immunoglobulin G (IgG) exhibited a completely smooth fluorescent outline in the case of A. naeslundii and and interrupted, irregular fluorescent outline in the case of human strains of A. viscosus. The different labeling patterns appeared to be related to the presence at the ultrastructural level of long, unevenly distributed strands of "fuzz" on the surface of human A. viscosus cells, whereas A. naeslundii cells had a narrower layer of fuzz, or more even thickness. The immunocoating reaction revealed homologous antibody binding to the irregular strands of fuzz on the surface of human A. viscosus cells, whereas homologous antisera to A. naeslundii coated A. naeslundii cells with a moderately electron-dense coating of antibody of even thickness. Human strains of A. viscosus incubated with heterologous antiserum to A. naeslundii followed by FITC-labeled goat anti-rabbit IgG exhibited a segmented fluorescent outline, which differed from that produced with homologous antisera. A. naeslundii incubated with heterologous rabbit antisera to human A. viscosus strains and FITC-labeled anti-rabbit IgG exhibited a completely smooth fluorescent outline similar to that produced with homologous antiserum. A. viscosus strains of hamster origin differed from A. viscosus strains of human origin by the absence of a surface fuzz and the comparatively smooth, even fluorescence produced by incubating these cells with homologous rabbit antiserum followed by FITC-labeled goat anti-rabbit IgG. Antiserum to a hamster strain did not cross-react with A. naeslundii or human strains of A. viscosus. Under the growth conditions of this experiment, ultrastructural features and labeling patterns with the indirect fluorescent technique may be useful in differentiating these serotypes from one another.

Actinomyces

Neuraminidase-dependent hamagglutination of human erythrocytes by human strains of Actinomyces viscosus and Actinomyces naeslundii.

Human A, B, and O erythrocytes (RBC) were agglutinated by many human strains of Actinomyces viscosus and A. naeslundii. At 37 degrees C, these bacterium-mediated hemagglutination reactions required the action of bacterial neuraminidase upon the RBC; however, at 4 degrees C, the requirement for neuraminidase was not as striking. Bacterial cell suspensions which caused hemagglutination at 37 degrees C contained both soluble extracellular and cell-associated neuraminidase activities as shown by enzyme assays using a soluble substrate (i.e., alpha 1-acid glycoprotein). Bacterium-mediated hemagglutination occurred only in the presence of soluble neuraminidase activity, and the rate of hemagglutination could be inhibited by 2-deoxy-2,3-dehydro-N-acetylneuraminic acid, a competitive inhibitor of purified soluble neuraminidase from A. viscosus T14V. Suspensions of bacteria which contained only cell-associated neuraminidase activity were unable to initiate hemagglutination, but they caused immediate hemagglutination when mixed with neuraminidase-treated RBC. All hemagglutination reactions were reversible in the presence of 0.02 M lactose and were abolished by heating (85 degrees C for 30 min) the actinomycete cells but not the RBC. The proposed mechanism of hemagglutination involves two sequential steps: (i) the action of neuraminidase to unmask galactose-containing receptors on the RBC and (ii) the multivalent binding of these receptors by many low-affinity lection sites on the bacterial surface.

Actinomyces

[Comparative study of the effect of aeration conditions on the biosynthesis of oxytetracycline by an actinomyces rimosus culture and of lincomycin by an Actinomyces roesolus culture].

The effect of aeration conditions on growth of the oxytetracycline- and lincomycin-producing organisms and antibiotic biosynthesis was studied. It was shown that the lincomycin-producing organism respiration rate was much higher and required better aeration condition-than the oxytetracycline-producing organism. The highest respiration rate of the young myces lium was observed in the growth phase. During the period of the antibiotic biosynthesis the rate of oxygen consumption somewhat decreased though remained sufficiently high. Decreased productivity of the mycelium at the end of the process was accompanied by a drop in the respiration rate. The lack of oxgen lowered the mycelium productivity with respect to the antibiotic formation to a much much greater extent than the culture growth rate. The limiting of the antibiotic biosynthesis process by the lack of dissolved oxygen was accompanied by changes in the culture metabolism evident from production of organic acids: ketoacids and volatile acids by Act. rimosus and volatile and lactic acids by Act. roseolus.

Aerobiosis

Numerical taxonomy and laboratory identification of Actinomyces and Arachnia and some related bacteria.

A numerical taxonomic study was made on 49 facultative anaerobic Gram-positive filamentous and/or diphtheroidal organisms isolated from dental plaques, carious dentin and faeces, together with 63 reference strains belonging to the genera Actinomyces, Arachnia, Bifidobacterium, Actinobacterium, Propionibacterium, Eubacterium and Lactobacillus. They were examined for 90 unit characters covering a wide range of tests and properties. The data were subjected to computer analysis in which the simple matching coefficient (SSM) and the similarity index (SJ) were calculated, and the results of single linkage techniques and an unweighted average linkage cluster analysis technique were compared. The strains fell into six major groups (phena). The Actinomyces strains were recovered in two phena; the first contained Actinomyces israelii and the other facultative anaerobic Actinomyces, including subclusters equal to taxospecies of A. odontolyticus and A. viscosus/A. naeslundii, while the other phenon corresponded to the genera Arachnia, Actinobacterium, Bifidobacterium and Propionibacterium. The groups of Arachnia and Actinobacterium each contained one species, representing taxospecies of Arachnia propionica and Actinobacterium meyerii. Taxonomic criteria, both constant and discriminative, were selected to form a diagnostic table useful for laboratory identification of this group of organisms. Immunofluorescence supported the numerical data.

Actinomyces

Pelvic colonization with Actinomyces in women using intrauterine contraceptive devices.

Recent reports suggest a relationship between intrauterine contraceptive device (IUD) use and colonization or infection of the genital tract with Actinomyces species. This prospective, case-controlled study was designed to determine the incidence of colonization or infection with Actinomyces in IUD users. None of 50 control patients and four of 50 study patients (8%) had Actinomyces identified. None of the patients with positive results had symptomatic pelvic infection. Cervical cytology was effective in detecting each of the four study patients with positive results. It appears that the presence of any type of IUD is a major predisposing factor to colonization with Actinomyces.

Actinomycosis

Actinomyces tissue specificity may depend on differences in receptor specificity for GalNAc beta-containing glycoconjugates.

Actinomyces naeslundii 12104 and A. viscosus LY7 were compared for receptor specificities and adherence properties because these relate to their oral colonization sites. Both strains bind GalNAc beta-containing glycosphingolipids (GSLs) in a GalNAc beta 1-3Gal alpha Oethyl-sensitive fashion but differ with respect to the number of cells bound to GSLs and the effect of neighboring sugar groups on the binding. Their hemagglutination and saccharide inhibition profiles confirms the existence of two receptor specificities (for example, when GalNAc beta 1-3Gal alpha Oethyl is multivalently conjugated to albumin, its inhibitory activity increases fourfold toward strain 12104 but decreases fourfold toward strain LY7). Trypsin or chymotrypsin treatment of human erythrocytes, which possess receptor GSLs, improves their hemagglutination with strain 12104. In contrast, the same treatment of chicken erythrocytes, which lack receptor GSLs, abolishes their hemagglutination. These findings suggest that both GSLs and glycoproteins act as functional receptors on eukaryotic cells. The strains also differ with respect to the following GalNAc beta 1-3Gal alpha Oethyl-sensitive adherence properties: (i) strain LY7 adheres somewhat better than does strain 12104 to buccal epithelial cells; (ii) in spite of their similar overall coaggregation patterns with streptococci, strain 12104 coaggregates with Streptococcus oralis MPB1 but strain LY7 does not; (iii) strain 12104 alone shows GalNAc beta-sensitive saliva aggregation and adherence to saliva-coated hydroxyapatite. The GSL binding patterns of fresh Actinomyces isolates reveal a high prevalence of LY7-like specificities among buccal isolates, whereas 12104-like specificities are most prevalent among plaque isolates. These findings strongly suggest that fresh Actinomyces isolates use fine specificity for GalNAc beta-containing glycoconjugates in recognition and subsequent colonization of specific oral surfaces.

Acetylgalactosamine

Salivary receptors for GalNAc beta-sensitive adherence of Actinomyces spp.: evidence for heterogeneous GalNAc beta and proline-rich protein receptor properties.

The receptors for GalNAc beta 1-3Gal alpha Oethyl (GalNAc beta)-sensitive adherence of Actinomyces strains to salivary pellicles were investigated. Parotid and submaxillary saliva from one individual was size fractionated and utilized in hydroxyapatite adherence assays with Actinomyces naeslundii 12104 and A. viscosus 19246 and LY7 with and without GalNAc beta. Three parotid salivary fractions, the high-molecular-weight, acidic proline-rich protein (PRP), and statherin fractions, promote GalNAc beta-sensitive adherence of strain 12104, whereas only the high-molecular-weight fraction of submaxillary saliva promotes such adherence. In contrast, strain LY7, possessing a variant GalNAc beta specificity, shows GalNAc beta-sensitive adherence to the leading and trailing regions of the submaxillary PRP fractions but less distinct adherence to the parotid and submaxillary high-molecular-weight fractions. In addition, the PRP and statherin fractions promote adherence of strains LY7 and 19246 that is not inhibited by GalNAc beta. However, whereas strain LY7 binds more strongly to the PRP fraction than to the statherin fraction, strain 19246 binds preferentially to the statherin fractions of parotid and submaxillary saliva. These salivary protein fractions were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunostained to detect glycosylated proteins. The different salivary receptor properties are paralleled by different glycosylation patterns. The variable GalNAc beta specificities may have evolved to match different salivary glycosylation patterns, and PRP and statherin binding properties seem to be heterogeneous among the Actinomyces strains.

Acetylgalactosamine