Phylogenetic Analysis of Genes Coding for 16S rRNA in Mammalian Ureaplasmas

Abstract

Phylogenetic relationships among species of the genus Ureaplasma were elucidated by analyzing 16s rRNA sequence information. The 16s rRNA genes of six strains of the mammalian Ureaplasma species were amplified by PCR and were sequenced directly by a primer walking method. The phylogenetic tree based on the nucleotide sequence of the 16s rRNA genes corresponded to the evolutionary history of the host animal species.The genus Ureaplasma in the family Mycoplasmataceae is composed of organisms that hydrolyze urea. The following six species have been recognized in the genus Ureaplasma: Ureaplasma urealyticum, isolated from humans; Ureaplasma diversum, isolated from bovines; Ureaplasma gallorale, isolated from birds; Ureaplasma felinum and Ureaplasma cati, isolated from cats; and UreapZasma canigenitalium, isolated from dogs. The accumulated data suggest that the human species U. urealyticum consists of two different clusters, the T960 and parvo biovars, which can be distinguished on the basis of the results of polyacrylamide gel electrophoresis of cell proteins (12,20), DNA-DNA homology data (1), restriction endonuclease cleavage patterns (13), restriction fragment length polymorphism data (4), manganese susceptibility data (14), and DNA modification system data (2). The genome sizes of U. urealyticum strains have been estimated to be 760 to 1,170 kbp by pulsedfield gel electrophoresis (9). Two loci for rRNA operons have been determined on the genome of U. urealyticum (3). In the present study, we sequenced and compared the 16s ribosomal DNAs (rDNAs) of the mammalian Ureaplasma species, including the two biovars of U. urealyticum, because phylogenetic analysis of the mycoplasmas has been greatly improved by analyzing 16s rRNA sequences (21).U. diversum A417T (T = type strain) was obtained from C. J.Howard and R. N. Gourlay, Institute for Research on Animal Diseases, Compton, England (8). U. felinum FT2-BT, U. cati F2T, and U. canigenitalium D6P-CT were all obtained from our laboratory stock cultures (5, 6). All of the strains were grown aerobically in liquid medium at 37°C as described previouslyThe almost complete 16s rDNA sequences of U. diversum, U. felinum, U. cati, and U. canigenitalium were amplified by PCR by using universal primers F16S (5'-GAGTTTGATCCT GGCTCAGG-3') and R16S (5'-GGGATACCTTGTTAC GACTI'-3'). Both strands of the PCR products were sequenced by the dideoxy termination method (18) by primer walking without molecular cloning.The nucleotide sequences of the 16s rDNAs of the Ureaplasma species were aligned by the method of Higgins et al. (7), using the DNASIS software package (Hitachi Software Engineering Co., Yokohama, Japan). A phylogenetic tree was constructed by the neighbor-joining method (17).The nucleotide sequences of the 16s rDNAs of strains T960T and 27 of the two biovars of U. urealyticum (19) were obtained from nucleotide sequence databases. The nucleotide sequence of U. gallorale was not included in this study since the U. gallorale 16s rRNA gene was not amplified by the PCR primers used,...

Molecular characterization of ureaplasmas isolated from reproductive tract of goats and sheep from Brazil

“…The specific primer set F16S and R16 amplifies nearly the entire 16S rDNA sequence of mollicutes and was used as described by Harasawa and Cassell 14 . PCR products (10 μL) were then electrophoresed in 1% agarose gel with 10 μg/mL of ethidium bromide and visualized with UV light (Vilber Lourmat, Germany).…”

An Epidemiological Survey of Hemoplasma Infection in Iriomote Cats (<i>Prionailurus bengalensis iriomotensis</i>)

“…M. haematoparvum' (Dog, EF416569) [26]; M. haemocanis (Dog, EF416568); M. coccoides (Mouse, AY171918) [14]; M. haemomuris (Mouse, U82963) [19]; 'C. M. haemobos' (Cattle, EF616468) [12]; M. wenyonii (Cattle, DQ641256) [11]; and M. suis (Pig, AY492086 [7]; and U. felinum (domestic cat, D78651) [7].…”

Destructive Polyarthropathy in Aborted Bovine Fetuses: A Possible Association with Ureaplasma diversum Infection?

“…5 was sequenced. High-quality sequence data for 625 base pairs had 99% homology with the published sequence from the type strain of U. diversum (strain A417) 2 and less than 97% homology with other Ureaplasma spp. The sequence amplified from fetus No.…”