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Journal of Clinical Periodontology Mar 1996Detection of putative pathogens is critical for delineating the etiology and progression of periodontitis. In the present study, we have used a polymerase chain reaction...
Detection of putative pathogens is critical for delineating the etiology and progression of periodontitis. In the present study, we have used a polymerase chain reaction (PCR) assay utilizing primers specific for the lkt A gene of Actinobacillus actinomycetemcomitans, the fimbrial gene of Porphyromonas gingivalis, and tdp A gene of Treponema denticola in order to determine the presence of these pathogens in subgingival plaque samples from periodontitis sites. These gene specific primers were also used to assess the detection of different strains of bacteria in the PCR assays. Primers for P. gingivalis detected P. gingivalis strain 33277, but no product was detected when primers were used with extracts from 4 species of Capnocytophaga, 3 species of Prevotella, 2 species of Porphyromonas other than P. gingivalis, Bacteroides levii, Escherichia coli, 3 strains of A. actinomycetemcomitans and 3 strains of T. denticola. PCR analysis using primers for the lkt A gene of A. actinomycetemcomitans also did not result in a product with any of these bacteria with the exception of a positive result with 3 different strains of A. actinomycetemcomitans. Primers selected from the tdp A gene of T. denticola did not identify any of the bacteria strains tested except T. denticola serovars a, b, and c. Thus, these primers were shown to amplify gene segments that are specific to either P. gingivalis (33277), A. actinomycetemcomitans (33384, 43717 and 43718) or T. denticola (35405, 33521 and 35404). The PCR assay may be used to rapidly detect the presence of periodontal pathogens in the future.
Topics: Adolescent; Adult; Aggregatibacter actinomycetemcomitans; Bacterial Proteins; Bacteroides; Capnocytophaga; Child; Child, Preschool; DNA Primers; Dental Plaque; Disease Progression; Escherichia coli; Fimbriae, Bacterial; Genes, Bacterial; Humans; Middle Aged; Periodontitis; Polymerase Chain Reaction; Porphyromonas; Porphyromonas gingivalis; Prevotella; Serotyping; Species Specificity; Treponema
PubMed: 8707980
DOI: 10.1111/j.1600-051x.1996.tb02078.x -
Clinical Infectious Diseases : An... Jun 1995Pigmented anaerobic gram-negative rods are currently categorized as 17 species distributed in three genera: Prevotella, Porphyromonas, and Bacteroides. These organisms... (Review)
Review
Pigmented anaerobic gram-negative rods are currently categorized as 17 species distributed in three genera: Prevotella, Porphyromonas, and Bacteroides. These organisms are often encountered in clinical specimens but are also found as part of the indigenous flora on various mucosal surfaces. Several studies are presently assessing the association of individual species with health and disease. For example, Porphyromonas gingivalis and Porphyromonas endodontalis are key putative pathogens in adult periodontitis and root canal infections, respectively. Porphyromonas asaccharolytica is prevalent in extraoral infections. The Porphyromonas species of animal origin have been isolated from infected bite wounds in humans. Isolates closely resembling Bacteroides levii have been recovered from various types of human infections. According to preliminary reports, Prevotella intermedia tends to be associated more often with periodontal disease than with a healthy oral cavity. In the laboratory, enzyme profiling facilitates the identification of these pigmented rods. Beta-Lactamase production is more common among prevotella species (30%-50%) than among Porphyromonas species (< 10%).
Topics: Animals; Gram-Negative Anaerobic Bacteria; Humans; Pigmentation; Porphyromonas; Prevotella
PubMed: 7548548
DOI: 10.1093/clinids/20.supplement_2.s187 -
Oral Diseases Mar 1995Eight oligonucleotides based upon regions of the small subunit 16S ribosomal RNA gene sequences were analysed against a background of their position within the molecule...
Oligonucleotide probes to the 16S ribosomal RNA: implications of sequence homology and secondary structure with particular reference to the oral species Prevotella intermedia and Prevotella nigrescens.
Eight oligonucleotides based upon regions of the small subunit 16S ribosomal RNA gene sequences were analysed against a background of their position within the molecule and their two-dimensional structure to rationalise their use in recognising Prevotella intermedia and Prevotella nigrescens. The 41 clinical isolates from both oral and respiratory sites and two reference strains were subjected to DNA-DNA hybridisation and multilocus enzyme electrophoresis to confirm their identity. Alignment of oligonucleotide probes designated I Bi-2 to I Bi-6 (for P. intermedia) and 2Bi-2 (for P. nigrescens) with the 16S rRNA suggested that these probes lacked specificity or were constructed from hypervariable regions. A 52-mer oligonucleotide (designated Bi) reliably detected both species. Because of the high degree of concordance between the 16S rRNAs of both species, it was necessary to vary the stringency of hybridisation conditions for detection of both species. Thus probe I Bi-I recognised P. intermedia while I Bi-I detected both P. intermedia and P. nigrescens at low stringency. However, under conditions of high stringency only P. nigrescens was recognised by probe 2Bi-I. These probes were highly specific and did not hybridise with DNA from the closely related P. corporis, nor other periodontal pathogens such as Fusobacterium nucleatum, Actinobacillus actinomycetemcomitans, Treponema denticola and several pigmented species such as Prevotella melaninogenica, P. denticola, P. loescheii, Porphyromonas asaccharolytica, Py. endodontalis, Py. gingivalis, Py. levii, and Py. macacae.
Topics: Bacterial Typing Techniques; Base Sequence; DNA, Bacterial; Humans; Molecular Sequence Data; Nucleic Acid Conformation; Nucleic Acid Hybridization; Oligonucleotide Probes; Prevotella; Prevotella intermedia; RNA, Bacterial; RNA, Ribosomal, 16S; Sequence Alignment; Sequence Analysis, RNA; Sequence Homology, Nucleic Acid
PubMed: 7553378
DOI: 10.1111/j.1601-0825.1995.tb00154.x -
Acta Microbiologica Et Immunologica... 1995Resistance to beta-lactam antibiotics of 183 clinical isolates belonging to Bacteroides, Porphyromonas and Prevotella was tested by a micro-broth dilution MIC method....
Resistance to beta-lactam antibiotics of 183 clinical isolates belonging to Bacteroides, Porphyromonas and Prevotella was tested by a micro-broth dilution MIC method. Beta-lactamase production was screened by using nitrocefin sticks. Prevalence of resistance to different beta-lactam antibiotics was higher among the Bacteroides strains other than B. fragilis parallel with a beta-lactamase production (88% and 96%, respectively). Resistance was observed less frequently among Porphyromonas and Prevotella strains. Cefoxitin resistance was 11.5% and amoxicillin/clavulanic acid 3.5% among Bacteroides isolates, whereas no resistance was found to these antibiotics among the Porphyromonas and Prevotella strains. All strains tested were susceptible for imipenem. Beta-lactamase production of selected isolates was tested quantitatively. Beta-lactamase of B. fragilis 1 and B. levii differed in their isoelectric points, substrate profiles and inhibition by clavulanic acid, sulbactam and tazobactam.
Topics: Anti-Bacterial Agents; Bacteroides; Drug Resistance, Microbial; Porphyromonas; Prevotella; beta-Lactamases; beta-Lactams
PubMed: 8548202
DOI: No ID Found -
Journal of Bacteriology Feb 1994The phylogenetic structure of the bacteroides subgroup of the cytophaga-flavobacter-bacteroides (CFB) phylum was examined by 16S rRNA sequence comparative analysis.... (Comparative Study)
Comparative Study
The phylogenetic structure of the bacteroides subgroup of the cytophaga-flavobacter-bacteroides (CFB) phylum was examined by 16S rRNA sequence comparative analysis. Approximately 95% of the 16S rRNA sequence was determined for 36 representative strains of species of Prevotella, Bacteroides, and Porphyromonas and related species by a modified Sanger sequencing method. A phylogenetic tree was constructed from a corrected distance matrix by the neighbor-joining method, and the reliability of tree branching was established by bootstrap analysis. The bacteroides subgroup was divided primarily into three major phylogenetic clusters which contained most of the species examined. The first cluster, termed the prevotella cluster, was composed of 16 species of Prevotella, including P. melaninogenica, P. intermedia, P. nigrescens, and the ruminal species P. ruminicola. Two oral species, P. zoogleoformans and P. heparinolytica, which had been recently placed in the genus Prevotella, did not fall within the prevotella cluster. These two species and six species of Bacteroides, including the type species B. fragilis, formed the second cluster, termed the bacteroides cluster. The third cluster, termed the porphyromonas cluster, was divided into two subclusters. The first contained Porphyromonas gingivalis, P. endodontalis, P. asaccharolytica, P. circumdentaria, P. salivosa, [Bacteroides] levii (the brackets around genus are used to indicate that the species does not belong to the genus by the sensu stricto definition), and [Bacteroides] macacae, and the second subcluster contained [Bacteroides] forsythus and [Bacteroides] distasonis. [Bacteroides] splanchnicus fell just outside the three major clusters but still belonged within the bacteroides subgroup. With few exceptions, the 16 S rRNA data were in overall agreement with previously proposed reclassifications of species of Bacteroides, Prevotella, and Porphyromonas. Suggestions are made to accommodate those species which do not fit previous reclassification schemes.
Topics: Bacteroides; Base Sequence; DNA Primers; Molecular Sequence Data; Phylogeny; Porphyromonas; RNA, Ribosomal, 16S; Sequence Alignment; Sequence Homology, Nucleic Acid
PubMed: 8300528
DOI: 10.1128/jb.176.3.725-732.1994 -
FEMS Immunology and Medical Microbiology Mar 1993Black-pigmented Gram-negative anaerobes are causative agents of pyogenic infections and are closely linked to various forms of periodontal diseases. Whereas many studies... (Comparative Study)
Comparative Study
Black-pigmented Gram-negative anaerobes are causative agents of pyogenic infections and are closely linked to various forms of periodontal diseases. Whereas many studies have shown a high incidence of plasmids in intestinal Bacteroides spp., there have been only a few reports of plasmid analyses in pigmented Gram-negative anaerobes. According to previous reports and confirmed in this study, plasmids can be present in Porphyromonas asaccharolytica, Prevotella intermedia, Pr. melaninogenica, and B. levii but have not been detected in P. gingivalis or other black-pigmented species. There were no correlations between plasmids and phenotypes such as resistance to antibiotics or bacteriocinogenicity. The highest carriage rate was found in isolates from cases of chronic otitis media, but the relationship between this site of infection and a high incidence of plasmids could be incidental. The size of plasmids ranged from 1.5 to 29 MDa. Plasmids with molecular weight > 10 MDa were described for the first time in these organisms. Repeated plasmid analyses showed that the plasmid patterns were generally stable.
Topics: Bacteroidaceae; Bacteroidaceae Infections; Humans; Phenotype; Plasmids
PubMed: 8518750
DOI: 10.1111/j.1574-695X.1993.tb00311.x -
Journal of Clinical Microbiology Sep 1991A rapid method for presumptive identification of black-pigmented gram-negative anaerobic rods was developed. Using filter paper spot tests for indole production,...
A rapid method for presumptive identification of black-pigmented gram-negative anaerobic rods was developed. Using filter paper spot tests for indole production, sialidase, alpha-glucosidase, beta-glucosidase, alpha-fucosidase, and trypsinlike enzyme activities, 100% of Porphyromonas gingivalis, Prevotella intermedia, and Bacteroides levii and 89% of Prevotella corporis isolates were correctly identified to the species level. Porphyromonas asaccharolytica and Porphyromonas endodontalis could not be differentiated from each other but could be distinguished from all other species tested. Similarly, Prevotella denticola, Prevotella loescheii, and Prevotella melaninogenica could not be differentiated from each other. The methods described are based on 4-methylumbelliferone derivatives of the various substrates and are simple to perform, rapid (less than 15 min), and applicable to difficult-to-cultivate anaerobic rods.
Topics: Bacteriological Techniques; Evaluation Studies as Topic; Glucosidases; Gram-Negative Anaerobic Bacteria; Hymecromone; Indoles; Neuraminidase; Pigmentation; alpha-L-Fucosidase
PubMed: 1774320
DOI: 10.1128/jcm.29.9.1955-1958.1991 -
Oral Microbiology and Immunology Aug 1990Eight species of black-pigmented Bacteroides (BPB) were examined for plasmid content. Three classes of plasmids with molecular sizes of 16.0 kilobase (kb), 5.4 kb and...
Eight species of black-pigmented Bacteroides (BPB) were examined for plasmid content. Three classes of plasmids with molecular sizes of 16.0 kilobase (kb), 5.4 kb and 5.0 kb were found in 7 of 125 Bacteroides intermedius strains. One strain each of Porphyromonas (former Bacteroides) asaccharolyticus and of Bacteroides levii were also shown to have plasmids. However, no plasmids were detected in 90 strains of Porphyromonas (Bacteroides) gingivalis. The plasmids were purified and digested with restriction endonucleases for restriction mapping. The maps obtained in this study have given us information on the scheme of constructing recombinants which may be usable as shuttle vectors, by the recombination of BPB and Escherichia coli plasmids.
Topics: Bacteroides; Chromosomes, Bacterial; Cloning, Molecular; DNA, Recombinant; Electrophoresis, Agar Gel; Plasmids; Restriction Mapping
PubMed: 2082244
DOI: 10.1111/j.1399-302x.1990.tb00647.x -
Journal of Clinical Microbiology Mar 1990A filter paper spot test with 2'-(4-methylumbelliferyl)-alpha-D-N-acetylneuraminic acid as a substrate was used to study the prevalence of sialidase activity among...
A filter paper spot test with 2'-(4-methylumbelliferyl)-alpha-D-N-acetylneuraminic acid as a substrate was used to study the prevalence of sialidase activity among gram-negative anaerobic and capnophilic bacteria. A total of 567 isolates representing four genera of obligate anaerobes and four genera of capnophilic organisms was tested. Sialidase activity was detected in 94% of 66 isolates from the Bacteroides fragilis group, 98% of 66 B. bivius isolates, and all isolates of the following species (number of isolates follows species name): B. capillosus, 4; B. levii, 2; B. denticola, 22; B. loescheii, 23; B. melaninogenicus, 32; B. forsythus, 44; and B. buccalis, 2. However, sialidase activity was detected in only 29% of 7 B. buccae isolates, 79% of 14 B. disiens isolates, and 55% of 11 B. oralis isolates. Sialidase activity was not detected among any of 13 isolates of B. gracilis, 12 isolates of B. ureolyticus, 61 isolates of B. intermedius, or 26 isolates of B. corporis. Porphyromonas (Bacteroides) asaccharolytica (20 isolates) and P. endodontalis (8 isolates) did not demonstrate sialidase activity, while 25 isolates of P. gingivalis were sialidase positive. Sialidase activity was found in 10 (100%) of 10 isolates of Capnocytophaga ochracea of C. sputigena but not in any of 4 C. gingivalis isolates. Other gram-negative anaerobic or capnophilic bacteria, including the following, were negative for sialidase activity: Fusobacterium nucleatum, 39 isolates; Wolinella recta, 19 isolates; Eikenella corrodens, 17 isolates; Haemophilus aphrophilus, 10 isolates; and Actinobacillus actinomycetemcomitans, 10 isolates. These data demonstrate sialidase activity in several species of the genera Bacteroides and Porphyromonas and suggest that this characteristic may be useful for identification.
Topics: Bacteria; Bacteroides; Carbon Dioxide; Fusobacterium; Gram-Negative Anaerobic Bacteria; Neuraminidase
PubMed: 2108991
DOI: 10.1128/jcm.28.3.422-425.1990