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PubMed1991

Lactose-sensitive and -insensitive cell surface interactions of oral Streptococcus milleri strains and actinomyces.

Of 158 oral Streptococcus milleri strains, 46 exhibited cellular coaggregation with the reagent strains of the actinomyces coaggregation groups A, B, and/or E. All but 1 of the 33 serotype b, e, f/F, and k/G strains belonged to streptococcus coaggregation group 2, and only 14 strains of limited seroclasses (g, i, Lancefield group F, or untypeable) appeared to be members of group 5, 3, or 4 (10, 3, and 1 strain, respectively). Thus, S. milleri infrequently exhibits lactose-inhibitable coaggregation with actinomyces.

Actinomyces↗
PubMed1991

Nucleotide sequence of a gene coding for a saliva-binding protein (SsaB) from Streptococcus sanguis 12 and possible role of the protein in coaggregation with actinomyces.

The nucleotide sequence of a 2.9-kb streptococcal DNA fragment which codes for two proteins with MrS of 36,000 (Streptococcus sanguis adhesin B [SsaB]) and 20,000 has been determined. The ssaB gene is 927 bp and codes for a 34,684-Da protein. The open reading frame coding for the 20-kDa protein is 489 bp and codes for a protein of 17,885 Da. The SsaB protein has a putative hydrophobic 19-amino-acid signal sequence resulting in a 32,620-Mr secreted protein, whereas the 20-kDa protein has no signal sequence. Both proteins are hydrophilic, and neither appears to have a hydrophobic membrane anchor sequence in the carboxy-terminal region. A DNA sequence homology of 73% exists between the cloned fragment containing the ssaB gene from S. sanguis 12 and the cloned fragment containing the type 1 fimbrial gene of S. sanguis FW213 (J.C. Fenno, D.J. LeBlanc, and P. Fives-Taylor, Infect. Immun. 57:3527-3533, 1989). Amino acid comparisons of the SsaB and type 1 fimbrial proteins show 87% homology, indicating a close similarity of the two proteins. Antiserum raised against the cloned SsaB protein cross-reacts with a 38-kDa protein identified from Streptococcus gordonii (S. sanguis) PK488 which was proposed to mediate coaggregation with Actinomyces naeslundii PK606 (P.E. Kolenbrander and R.N. Andersen, Infect. Immun. 58:3064-3072, 1990). The SsaB adhesion may play a role in oral colonization by binding either to a receptor on saliva or to a receptor on actinomyces.

Actinomyces↗
PubMed1993

Cloning of the Streptococcus gordonii PK488 gene, encoding an adhesin which mediates coaggregation with Actinomyces naeslundii PK606.

Coaggregation between Streptococcus gordonii PK488 and Actinomyces naeslundii PK606 is mediated by a 38-kDa streptococcal protein, designated ScaA. The gene, scaA, which encodes this protein has been cloned into Escherichia coli. A genomic S. gordonii PK488 library (in Lambda ZAP II) was screened with anti-S. gordonii immunoglobulin G absorbed with S. gordonii PK1804, an isogenic coaggregation-defective mutant of strain PK488. A positive recombinant phage was isolated, and a phagemid designated pRA1 was obtained which contained a 6.6-kb insert. Expression of scaA from pRA1 and from a subcloned internal 2.1-kb fragment was observed. The absorbed antiserum cross-reacted with a 34.7-kDa protein, SsaB, from S. sanguis 12, also a coaggregation partner of A. naeslundii PK606. Absorbed antiserum to S. gordonii PK488 and antiserum to SsaB both reacted with 38-kDa proteins in supernatants from mildly sonicated preparations from 11 other coaggregation partners of A. naeslundii PK606. Putative adhesin genes were identified in each of these coaggregation partners by Southern analysis of their genomic DNA with the cloned 2.1-kb fragment as a probe. A 30-base oligonucleotide probe based on the sequence of ssaB of S. sanguis 12 hybridized in an identical manner. These data extend the notion that most of the viridans streptococci that coaggregate with actinomyces are capable of expressing ScaA-related proteins.

Actinomyces↗
PubMed1995

Construction and use of integration plasmids to generate site-specific mutations in the Actinomyces viscosus T14V chromosome.

Stable transformants of Actinomyces viscosus T14V carrying heterologous DNA were obtained with the aid of integration plasmids. These plasmids contained a kanamycin resistance (Kmr) gene flanked by A. viscosus T14V genomic DNA, including parts of the type 1 structural fimbrial subunit gene (fimP) on one or both sides of the antibiotic marker. Significantly more Kmr transformants were obtained with a plasmid carrying longer segments of homologous strain T14V DNA. Integration of this plasmid into the A. viscosus T14V genome affected the expression and function of type 1 fimbriae in the transformants. In the transformant strain designated A. viscosus MY50D, the inactivated fimP replaced the wild-type fimP via allelic replacement. A. viscosus MY51S and MY52S each contained a copy of the plasmid integrated into the genome by a Campbell-like insertion mechanism. A. viscosus MY50D and MY51S lacked type 1 fimbriae and did not bind to proline-rich proteins (the fimbrial receptors) immobilized on nitrocellulose. In contrast, strain MY52S synthesized the structural subunit protein, as detected by immunostaining with anti-A. viscosus T14V type 1 fimbria antibodies. However, the high-molecular-weight proteins observed in sodium dodecyl sulfate-polyacrylamide gels of fimbriae from the cell wall of the wild-type strain T14V were absent in cell wall preparations of this strain. Moreover, A. viscosus MY52S failed to bind, in vitro, to proline-rich proteins. Thus, these results demonstrate that insertion of heterologous DNA at specific sites of the Actinomyces genome can be facilitated with integratable plasmids and that the transformants and mutants generated will aid in the delineation of the roles and contributions of specific genes to the structure and function of any macromolecule produced by these organisms.

Actinomyces↗
PubMed1997

Synthesis and function of Actinomyces naeslundii T14V type 1 fimbriae require the expression of additional fimbria-associated genes.

The nucleotide sequence of the chromosomal DNA flanking the Actinomyces naeslundii (formerly A. viscosus) T14V type 1 fimbrial structural subunit gene (fimP) was determined. Six open reading frames (ORFs), in the order 5' ORF3, ORF2, ORF1,fimP, ORF4, ORF5, ORF6 3', were identified. ORF1 encoded a protein of 408 amino acid residues (Mr = 39,270) and had significant sequence homology with the A. naeslundii T14V type 1 and A. naeslundii WVU45 type 2 fimbrial structural subunits. An in-frame fusion of ORF1 to the malE gene of the expression vector, pMAL-c2, yielded a protein that was immunostained with antibodies raised against the maltose binding protein and A. naeslundii T14V whole bacteria. Digestion of the fusion protein with factor Xa released a protein (apparent molecular mass of 34 kDa) that was immunostained only with the antibody directed against A. naeslundii T14V whole bacterial cells. Integration plasmids carrying a kanamycin resistance gene (kan) that was used to substitute for ORF1 or for DNA fragments internal to the coding region of the other five ORFs were used to transform A. naeslundii T14V. Neither type 1 fimbriae nor the 65-kDa fimbrial structural subunit was detected in mutants obtained by allelic replacement of ORF1 or ORF2. Mutants obtained by allelic replacement of ORF3 or ORF4 expressed only the 65-kDa fimbrial structural subunit. These mutants did not bind, in vitro, to proline-rich proteins that serve as the receptors for Actinomyces type 1 fimbriae. In contrast, a mutant in which the integration plasmid DNA had been inserted at a site close to the carboxyl terminus of ORF6 expressed type 1 fimbriae and had adherence properties similar to those observed in the wild-type strain. These results demonstrate the existence of additional genes near fimP that are likely to be involved in the synthesis and function of cell surface fimbriae of A. naeslundii T14V.

Actinomyces viscosus↗
PubMed2001

Roles of fructosyltransferase and levanase-sucrase of Actinomyces naeslundii in fructan and sucrose metabolism.

The ability of Actinomyces naeslundii to convert sucrose to extracellular homopolymers of fructose and to catabolize these types of polymers is suspected to be a virulence trait that contributes to the initiation and progression of dental caries and periodontal diseases. Previously, we reported on the isolation and characterization of the gene, ftf, encoding the fructosyltransferase (FTF) of A. naeslundii WVU45. Allelic exchange mutagenesis was used to inactivate ftf, revealing that FTF-deficient stains were completely devoid of the capacity to produce levan-type (beta2,6-linked) polysaccharides. A polyclonal antibody was raised to a histidine-tagged, purified A. naeslundii FTF, and the antibody was used to localize the enzyme in the supernatant fluid. A sensitive technique was developed to detect levan formation by proteins that had been separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and the method was used to confirm that the levan-synthesizing activity of A. naeslundii existed predominantly in a cell-free form, that a small amount of the activity was cell associated, and that the ftf mutant was unable to produce levans. By using the nucleotide sequence of the levanase gene of a genospecies 2 A. naeslundii, formerly Actinomyces viscosus, a portion of a homologue of this gene (levJ) was amplified by PCR and inserted into a suicide vector, and the resulting construct was used to inactivate the levJ gene in the genospecies 1 strain WVU45. A variety of physiologic and biochemical studies were performed on the wild-type and LevJ-deficient strains to demonstrate that (i) this enzyme was the dominant levanase and sucrase of A. naeslundii; (ii) that LevJ was inducible by growth in sucrose; (iii) that the LevJ activity was found predominantly (>90%) in a cell-associated form; and (iv) that there was a second, fructose-inducible fructan hydrolase activity produced by these strains. The data provide the first detailed molecular analysis of fructan production and catabolism in this abundant and important oral bacterium.

Actinomyces↗
PubMed1995

Phosphorylating enzymes involved in glucose fermentation of Actinomyces naeslundii.

Enzymatic activities involved in glucose fermentation of Actinomyces naeslundii were studied with glucose-grown cells from batch cultures. Glucose could be phosphorylated to glucose 6-phosphate by a glucokinase that utilized polyphosphate and GTP instead of ATP as a phosphoryl donor. Glucose 6-phosphate was further metabolized to the end products lactate, formate, acetate, and succinate through the Embden-Meyerhof-Parnas pathway. The phosphoryl donor for phosphofructokinase was only PPi. Phosphoglycerate kinase, pyruvate kinase, and acetate kinase coupled GDP as well as ADP, but P(i) compounds were not their phosphoryl acceptor. Cell extracts showed GDP-dependent activity of phosphoenolpyruvate carboxykinase, which assimilates bicarbonate and phosphoenolpyruvate into oxaloacetate, a precursor of succinate. Considerable amounts of GTP, polyphosphate, and PPi were found in glucose-fermenting cells, indicating that these compounds may serve as phosphoryl donors or acceptors in Actinomyces cells. PPi could be generated from UTP and glucose 1-phosphate through catalysis of UDP-glucose synthase, which provides UDP-glucose, a precursor of glycogen.

Acetate Kinase↗
PubMed1961

Antigenic structure and serological relationships of Mycobacterium, Actinomyces, Streptococcus, and Diplococcus.

Kwapinski, Jerzy B. (South African Institute for Medical Research, Johannesburg), and Marshall L. Snyder. Antigenic structure and serological relationships of Mycobacterium, Actinomyces, Streptococcus, and Diplococcus. J. Bacteriol. 82:632-639. 1961.-Chemical fractions, 72 in all, from four strains of Mycobacterium, four strains of Actinomyces, and four strains of Streptococceae, were tested in homologous and heterologous antisera prepared against intact cells and chemical fractions of Mycobacterium tuberculosis. Antigens isolated from the cytoplasm of these bacteria proved to be serologically related to each other. In contrast, most of the cell-wall antigens, except those extracted with either 30% sodium hydroxide or formamide at 100 to 130 C, were species specific. Sera prepared against the individual fractions of Mycobacterium reacted only with the strain of Mycobacterium from which they were isolated.

Actinomyces↗
PubMed1969

Determination and analysis of Actinomyces israelii serotypes by fluorescent-antibody procedures.

This study was undertaken to determine the antigenic relationships between serotypes of Actinomyces israelii with fluorescent-antibody (FA) procedures. In addition, the antigenic relationships between A. israelii and other members of the genus Actinomyces were studied by the same methods. Seven FA conjugates were used to determine the serological characteristics of 28 isolates believed to represent A. israelii, serotypes 1 and 2. The results showed that the lower dilutions of serotype 1 conjugates stained serotype 2 antigens; however, serotype 2 conjugates did not stain serotype 1 antigens. Serotype 1 conjugate could be made specific by adsorption. A. israelii serotype 1 conjugate cross-reacted also with A. naeslundii, but this cross-reaction could be eliminated by adsorption or dilution. Serotype 2 conjugate appeared to be specific for A. israelii serotype 2. Adsorption studies revealed antigenic variants among the various A. israelii serotype 1 and 2 isolates. However, all isolates could be identified by direct FA staining with appropriate conjugates. One isolate previously identified as A. israelii was shown, on the basis of FA studies, to be an A. naeslundii. A polyvalent diagnostic reagent was prepared which was specific for A. israelii serotypes 1 and 2. This reagent should find application in diagnostic and reference laboratories.

Actinomyces↗
PubMed1969

Morphological, biochemical, and serological studies on 64 strains of Actinomyces israelii.

A comparative study of 64 strains of Actinomyces israelii was done with the use of techniques standardized by the Subgroup on Taxonomy of the Microaerophilic Actinomycetes. Emphasis was placed on the range of variation to assist recognition of clinical isolates and aid in differentiation from Actinomyces-like organisms. None of the strains was positive for catalase or indole, or in the Voges-Proskauer test; 90% were methyl red-positive and 62% were nitrate-positive. Acid was produced from: glucose (100%), xylose (100%), salicin (98%), raffinose (95%), lactose (89%), cellobiose (83%), mannose (78%), arabinose (76%), inositol (58%), mannitol (48%), starch (31%), glycogen (0%), glycerol (0%), and rhamnose (0%). A. israelii can be identified by the fluorescent-antibody method, but there is no single morphological or biochemical characteristic which can be used for its identification. By both fluorescent-antibody and gel-diffusion techniques, the serological classification of A. israelii group D with serotypes 1 and 2 was verified. Eleven serotype 2 strains were compared morphologically, biochemically, and serologically with 53 serotype 1 strains. All but two of the serotype 2 strains produced viscous growth in broth and none fermented arabinose.

Actinomyces↗
PubMed1981

Demonstration of Actinomyces and Arachnia species in cervicovaginal smears by direct staining with species-specific fluorescent-antibody conjugate.

For direct observation of microaerophilic actinomycetes by fluorescent antibody, a procedure was developed in which pepsin treatment and rhodamine conjugate of normal serum were used to reduce nonspecific staining in cervicovaginal smears. Actinomyces israelii, Actinomyces naeslundii, and Arachnia propionica were observed in cervicovaginal smears from women who did use and who did not use an intrauterine contraceptive device. A. israelii was found more commonly in women with an intrauterine contraceptive device, but no evidence was obtained that the use of an intrauterine contraceptive device influenced the presence of either A. propionica or A. naeslundii.

Actinomyces↗
PubMed1989

Detection of sialidase (neuraminidase) activity in Actinomyces species by using 2'-(4-methylumbelliferyl)alpha-D-N-acetylneuraminic acid in a filter paper spot test.

A rapid method for the detection of acetylneuraminyl hydrolase, EC 3.2.1.18 (sialidase or neuraminidase), was developed by using 2'-(4-methylumbelliferyl)alpha-D-N-acetylneuraminic acid as substrate in a filter paper spot test. The method was compared to conventional assays that use 2'-(4-methylumbelliferyl)alpha-D-N-acetylneuraminic acid and bovine submaxillary mucin and was found to be in excellent agreement. Organisms with greater than 10 U of enzyme activity (in nanomoles per minute per milligram of cell protein) gave positive reactions, while those with 2.7 to 9.0 U gave only weak reactions. Isolates with less than 2.7 U of activity were detected upon prolonged incubation. Sialidase activity was detected in 79% of 71 clinical isolates representing five species of Actinomyces. The percentage of sialidase-producing isolates of each species varied considerably: Actinomyces israelii, 63%; A. meyeri, 73%; A. naeslundii, 85%; A. odontolyticus, 73%; and A. viscosus, 100%.

Actinomyces↗
PubMed1999

Clinical spectrum of infections due to the newly described Actinomyces species A. turicensis, A. radingae, and A. europaeus.

Over a 7-year period, we isolated 294 Actinomyces-like organisms (ALOs) which were not clearly identifiable. Using well-defined probes coding for sequences specific for recently described Actinomyces species (A. turicensis, A. radingae, and A. europaeus), we were able to identify 128 strains. The majority belonged to the A. turicensis species. A. radingae was found only in patients with skin-related pathologies. A. europaeus was also detected in patients with urinary tract infections. The main sources of A. turicensis were genital infections, followed by skin-related and urinary tract infections. Additional clinical pictures were appendicitis, cholecystitis, ear, nose, and throat infections, and bacteremia. In a small number of patients these ALOs were found as the only pathogen. Strains of the three species were tested by two widely used biochemical identification methods. A. turicensis was easily identifiable by both these methods. We conclude that these ALOs are not infrequent pathogens and are found in a wide range of human infections. At least A. turicensis is easily identifiable by clinical diagnostic laboratories.

Actinomyces↗
PubMed2002

Late prosthetic hip joint infection with Actinomyces israelii in an intravenous drug user: case report and literature review.

Late infections with Actinomyces israelii have been described for prosthetic hip joints but not in association with intravenous drug use. We present a case of a 43-year-old intravenous drug user who developed A. israelii infection in connection with a hip prosthesis 11 years after implantation, and we review four previously reported cases of Actinomyces prosthetic joint infections.

Actinomyces↗
PubMed1980

Actinomyces-like organisms in cervical smears from women using intrauterine contraceptive devices.

Cervical smears from 293 users of intrauterine contraceptive devices attending family planning clinics in East Fife, Dundee, and Angus were stained by Papanicolaou and Gram's methods and examined for actinomyces-like organisms. Of the 128 women using plastic devices, 40 gave smears positive for these organisms. In contrast only two positive smears were obtained from the 165 women using devices containing copper and none from a control group of 300 women taking oral contraceptives. Colonisation was more common in women whose plastic devices had been in situ for over two years. Correlations between the presence of these organisms and recorded incidences of pain and both clinical and cytological evidence of inflammation of the lower genital tract were highly significant (p = 0.00001, p < 0.00001, and p < 0.00001 respectively). The results suggest that plastic intrauterine contraceptive devices predispose to colonisation by actinomyces-like organisms, particularly after long-term use. Hence if the apparently bacteriostatic action of copper devices is confirmed these should probably be more widely used.

Actinomyces↗
PubMed1983

Actinomyces in the female genital tract. A preliminary report.

Actinomyces spp were isolated by culture of endocervical specimens from two groups of women attending the department of genitourinary medicine of the Royal Hallamshire Hospital, Sheffield. The first group consisted of 78 users of intrauterine contraceptive devices (IUCD) of whom 20 (25.6%) were culture positive. The second group contained 63 women using various forms of contraception 12 (19%) of whom were culture positive. None of these 12 women had an IUCD or foreign body in situ. The results suggest that Actinomyces spp may be part of the commensal flora of the genital tract in some women.

Actinomyces↗