Amplified 16S Ribosomal DNA Research Articles

Phylogenetic Position of Taylorella equigenitalis Determined by Analysis of Amplified 16S Ribosomal DNA Sequences

The 16S ribosomal DNA sequence of Taylorella equigenitalis (formerly Haemophilus equigenitalis), the causative organism of contagious equine metritis, was determined. A phylogenetic analysis of this sequence revealed a phylogenetic position of T. equigenitalis in the beta subclass of the class Proteobacteria apart from the position of Haemophilus influenzae, which belongs to the gamma subclass of Proteobacteria. A close phylogenetic relationship among T. equigenitalis, Alcaligenes xylosoxidans, and Bordetella bronchiseptica was detected; Spirillum volutans and Chromobacterium fluviatile (Iodobacter fluviatile) were in the same group but slightly removed. This relationship is surprising in view of the considerable differences in the G + C contents of the genomes of these bacteria.

Restriction fragment length polymorphism analysis of 16S ribosomal DNA of Streptococcus and Enterococcus species of bovine origin.

Twelve bacterial species including Streptococcus uberis, S. parauberis, S. agalactiae, S. dysgalactiae, S. bovis, S. mitis, S. salivarius, S. saccharolyticus, Enterococcus faecium, E. faecalis, E. avium, and Aerococcus viridans were examined for their 16S ribosomal DNA fingerprint patterns. Oligonucleotide primers complementary to 16S rRNA genes were used to amplify by the polymerase chain reaction 16S ribosomal gene fragments from genomic DNAs. The molecular sizes of the amplified 16S ribosomal DNA (rDNA) fragments from the 12 species examined ranged from 1,400 to 1,500 bp. Restriction fragment length polymorphism analysis of 16S rDNA was performed with 11 different restriction endonucleases. All 12 species examined could be differentiated on the basis of characteristic 16S rDNA fingerprint patterns by using the restriction endonucleases HhaI, RsaI, and MspI. A scheme for the differentiation of the 12 species is presented. Eleven isolates representing 11 species were obtained from cows with intramammary infections and were examined by 16S rDNA fingerprinting. All 11 species isolated from cows were differentiated by using HhaI, RsaI, and MspI restriction endonucleases. The results of this study demonstrate the potential application of 16S rDNA fingerprinting for the identification and differentiation of bacterial species.

Detection of Leptospiraceae by amplification of 16S ribosomal DNA

The polymerase chain reaction (PCR) was developed to detect Leptospiraceae. Primers were used to amplify a 631 base-pair (bp) 5′-region of 16S rDNA. Representative strains from the species, Leptospira interrogans sensu stricto, L. borgpetersenii, L. noguchii, L. santarosai, L. weilii, L. inadai, L. meyeri and the single member strain of Leptonema were amplified. In contrast, strains representing the saprophytic species, L. biflexa, L. wolbachii and L. parva were not amplified. There was no PCR product from 23 phylogenetically unrelated species of bacteria. As little as 10-1 pg of purified DNA and as few as 10-1 leptospires could be detected using this PCR analysis. Isolates of leptospires from clinical sources gave a positive PCR band, but those from surface waters did not.

Detection of Leptospiraceae by amplification of 16S ribosomal DNA

Detection of Leptospiraceae by amplification of 16S ribosomal DNA