Detection of Actinomyces species using nonradioactive riboprobes coupled with polymerase chain reaction. 1996

M Kiyama, and K Hiratsuka, and S Saito, and T Shiroza, and H Takiguchi, and Y Abiko
Department of Biochemistry, Nihon University School of Dentistry at Matsudo, Sakaecho-Nishi, Matsudo, Chiba 271, 2-870-1, Japan.

We have been focusing our attention on the detection and identification of oral bacteria which are frequently associated with periodontal disease. In previous studies, Actinomyces species-specific riboprobes were generated and used to identify this microorganism. However, problems lie in the low sensitivity of this method. We have developed a novel system for the detection of Actinomyces species using nonradioactive riboprobes coupled with polymerase chain reaction (PCR) in this study. This system employs two procedures; initially, DNA fragments specific for the target microorganism are amplified by PCR, and these specific fragments are further hybridized with nonradioactive riboprobes. PCR analysis using chromosomal DNA isolated from Actinomyces species including laboratory strains, clinical isolates, and Actinomyces naeslundii (ATCC 12104) indicated the presence of the predicted common 756-bp fragment, a portion of the sialidase gene. These amplified DNA fragments were effectively visualized by hybridization with the digoxigenin-labeled riboprobes corresponding to the internal region of the amplified sialidase gene. With this system, approximately three orders of magnitude less chromosomal DNA was sufficient for the detection of specific microorganisms compared to the conventional riboprobe systems.

UI MeSH Term Description Entries
D009439 Neuraminidase An enzyme that catalyzes the hydrolysis of alpha-2,3, alpha-2,6-, and alpha-2,8-glycosidic linkages (at a decreasing rate, respectively) of terminal sialic residues in oligosaccharides, glycoproteins, glycolipids, colominic acid, and synthetic substrate. (From Enzyme Nomenclature, 1992) Sialidase,Exo-alpha-Sialidase,N-Acylneuraminate Glycohydrolases,Oligosaccharide Sialidase,Exo alpha Sialidase,Glycohydrolases, N-Acylneuraminate,N Acylneuraminate Glycohydrolases,Sialidase, Oligosaccharide
D004269 DNA, Bacterial Deoxyribonucleic acid that makes up the genetic material of bacteria. Bacterial DNA
D006801 Humans Members of the species Homo sapiens. Homo sapiens,Man (Taxonomy),Human,Man, Modern,Modern Man
D000190 Actinomyces A genus of gram-positive, rod-shaped bacteria whose organisms are nonmotile. Filaments that may be present in certain species are either straight or wavy and may have swollen or clubbed heads.
D012313 RNA A polynucleotide consisting essentially of chains with a repeating backbone of phosphate and ribose units to which nitrogenous bases are attached. RNA is unique among biological macromolecules in that it can encode genetic information, serve as an abundant structural component of cells, and also possesses catalytic activity. (Rieger et al., Glossary of Genetics: Classical and Molecular, 5th ed) RNA, Non-Polyadenylated,Ribonucleic Acid,Gene Products, RNA,Non-Polyadenylated RNA,Acid, Ribonucleic,Non Polyadenylated RNA,RNA Gene Products,RNA, Non Polyadenylated
D013045 Species Specificity The restriction of a characteristic behavior, anatomical structure or physical system, such as immune response; metabolic response, or gene or gene variant to the members of one species. It refers to that property which differentiates one species from another but it is also used for phylogenetic levels higher or lower than the species. Species Specificities,Specificities, Species,Specificity, Species
D015342 DNA Probes Species- or subspecies-specific DNA (including COMPLEMENTARY DNA; conserved genes, whole chromosomes, or whole genomes) used in hybridization studies in order to identify microorganisms, to measure DNA-DNA homologies, to group subspecies, etc. The DNA probe hybridizes with a specific mRNA, if present. Conventional techniques used for testing for the hybridization product include dot blot assays, Southern blot assays, and DNA:RNA hybrid-specific antibody tests. Conventional labels for the DNA probe include the radioisotope labels 32P and 125I and the chemical label biotin. The use of DNA probes provides a specific, sensitive, rapid, and inexpensive replacement for cell culture techniques for diagnosing infections. Chromosomal Probes,DNA Hybridization Probe,DNA Probe,Gene Probes, DNA,Conserved Gene Probes,DNA Hybridization Probes,Whole Chromosomal Probes,Whole Genomic DNA Probes,Chromosomal Probes, Whole,DNA Gene Probes,Gene Probes, Conserved,Hybridization Probe, DNA,Hybridization Probes, DNA,Probe, DNA,Probe, DNA Hybridization,Probes, Chromosomal,Probes, Conserved Gene,Probes, DNA,Probes, DNA Gene,Probes, DNA Hybridization,Probes, Whole Chromosomal
D016133 Polymerase Chain Reaction In vitro method for producing large amounts of specific DNA or RNA fragments of defined length and sequence from small amounts of short oligonucleotide flanking sequences (primers). The essential steps include thermal denaturation of the double-stranded target molecules, annealing of the primers to their complementary sequences, and extension of the annealed primers by enzymatic synthesis with DNA polymerase. The reaction is efficient, specific, and extremely sensitive. Uses for the reaction include disease diagnosis, detection of difficult-to-isolate pathogens, mutation analysis, genetic testing, DNA sequencing, and analyzing evolutionary relationships. Anchored PCR,Inverse PCR,Nested PCR,PCR,Anchored Polymerase Chain Reaction,Inverse Polymerase Chain Reaction,Nested Polymerase Chain Reaction,PCR, Anchored,PCR, Inverse,PCR, Nested,Polymerase Chain Reactions,Reaction, Polymerase Chain,Reactions, Polymerase Chain
D016924 Actinomyces viscosus A species of ACTINOMYCES found in the oral cavity of man and hamsters. It has been isolated from actinomycotic lesions in swine, cats, and dogs and has been identified as a causative agent of animal diseases.
D017403 In Situ Hybridization A technique that localizes specific nucleic acid sequences within intact chromosomes, eukaryotic cells, or bacterial cells through the use of specific nucleic acid-labeled probes. Hybridization in Situ,Hybridization, In Situ,Hybridizations, In Situ,In Situ Hybridizations

Related Publications

M Kiyama, and K Hiratsuka, and S Saito, and T Shiroza, and H Takiguchi, and Y Abiko
January 1994, Journal of immunological methods,
M Kiyama, and K Hiratsuka, and S Saito, and T Shiroza, and H Takiguchi, and Y Abiko
June 1992, Rinsho byori. The Japanese journal of clinical pathology,
M Kiyama, and K Hiratsuka, and S Saito, and T Shiroza, and H Takiguchi, and Y Abiko
August 1995, Clinical chemistry,
M Kiyama, and K Hiratsuka, and S Saito, and T Shiroza, and H Takiguchi, and Y Abiko
December 1995, Rinsho byori. The Japanese journal of clinical pathology,
M Kiyama, and K Hiratsuka, and S Saito, and T Shiroza, and H Takiguchi, and Y Abiko
January 1993, Methods in molecular biology (Clifton, N.J.),
M Kiyama, and K Hiratsuka, and S Saito, and T Shiroza, and H Takiguchi, and Y Abiko
May 2002, Journal of virological methods,
M Kiyama, and K Hiratsuka, and S Saito, and T Shiroza, and H Takiguchi, and Y Abiko
October 1994, BioTechniques,
M Kiyama, and K Hiratsuka, and S Saito, and T Shiroza, and H Takiguchi, and Y Abiko
October 1997, Plant disease,
M Kiyama, and K Hiratsuka, and S Saito, and T Shiroza, and H Takiguchi, and Y Abiko
October 1992, Journal of clinical microbiology,
M Kiyama, and K Hiratsuka, and S Saito, and T Shiroza, and H Takiguchi, and Y Abiko
August 1990, Analytical biochemistry,
Copied contents to your clipboard!