A novel urease-negative Helicobacter species associated with colitis and typhlitis in IL-10-deficient mice.
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A spiral-shaped bacterium with bipolar, single-sheathed flagella was isolated from the intestines of IL-10 (interleukin-10)-deficient (IL-10(-/-)) mice with inflammatory bowel disease. The organism was microaerobic, grew at 37 and 42 degrees C, and was oxidase and catalase positive but urease negative. On the basis of 16S rRNA gene sequence analysis and biochemical and phenotypic criteria, the organism is classified as a novel helicobacter. Cesarean section-rederived IL-10(-/-) mice without helicobacter infection did not have histological evidence of intestinal inflammation. However, helicobacter-free IL-10(-/-), SCID/NCr, and A/JNCr mice experimentally inoculated with the novel urease-negative Helicobacter sp. developed variable degrees of inflammation in the lower intestine, and in immunocompetent mice, the experimental infection was accompanied by a corresponding elevated immunoglobulin G antibody response to the novel Helicobacter sp. antigen. These data support other recent studies which demonstrate that multiple Helicobacter spp. in both naturally and experimentally infected mice can induce inflammatory bowel disease. The mouse model of helicobacter-associated intestinal inflammation should prove valuable in understanding how specific microbial antigens influence a complex disease process. (+info)
Nucleotide changes in mitochondrial 16S rRNA gene from different mammalian cell lines.
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The partial nucleotide sequences of mitochondrial 16S rRNA gene were analyzed in five rodent cell lines, prior to the analysis of mutation spectrum in the gene. Total DNA was isolated from V79 and CHO-K1 cell lines from Chinese hamster and murine cell lines, Balb Y SV and PCC4 AG Cap, and the 3' terminal regions including the peptidyl transferase domain which is the target for chloramphenicol, a selective inhibitor of mitochondrial protein synthesis, were amplified by polymerase chain reaction (PCR) using two sets of primers and directly sequenced. In Chinese hamster cells, C to T transition at one site was observed in CHO-K1, and either A was deleted at the sequence of AA in all three cell lines, relative to the V79-cell sequence registered in GenBank. One G to A transition mutation in heteroplasmic state was observed in mouse PCC4 AG Cap cells which have chloramphenicol resistant phenotype, whereas there was no change in the Balb Y SV cell line, relative to the L-cell sequence. These mutation sites were located outside the peptidyl transferase domain. (+info)
A cytotoxic ribonuclease targeting specific transfer RNA anticodons.
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The carboxyl-terminal domain of colicin E5 was shown to inhibit protein synthesis of Escherichia coli. Its target, as revealed through in vivo and in vitro experiments, was not ribosomes as in the case of E3, but the transfer RNAs (tRNAs) for Tyr, His, Asn, and Asp, which contain a modified base, queuine, at the wobble position of each anticodon. The E5 carboxyl-terminal domain hydrolyzed these tRNAs just on the 3' side of this nucleotide. Tight correlation was observed between the toxicity of E5 and the cleavage of intracellular tRNAs of this group, implying that these tRNAs are the primary targets of colicin E5. (+info)
Phylogenetic analysis of Piscirickettsia salmonis by 16S, internal transcribed spacer (ITS) and 23S ribosomal DNA sequencing.
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Piscirickettsia salmonis, the etiologic agent of piscirickettsiosis, is a systemic disease of salmonid fish. Variations in virulence and mortality have been observed during epizootics at different geographical regions and in laboratory experiments with isolates from these different locations. This raises the possibility that biogeographical patterns of genetic variation might be a significant factor with this disease. To assess the genetic variability the 16S ribosomal DNA, the internal transcribed spacer (ITS) and the 23S ribosomal DNA of isolates from 3 different hosts and 3 geographic origins were amplified using the polymerase chain reaction (PCR). Results of this analysis confirm that P. salmonis is a member of the gamma subgroup of the Proteobacteria and show that the isolates form a tight monophyletic cluster with 16S rDNA similarities ranging from 99.7 to 98.5%. The ITS regions were 309 base pairs (bp), did not contain tRNA genes, and varied between isolates (95.2 to 99.7% similarity). Two-thirds of the 23S rRNA gene was sequenced from 5 of the isolates, yielding similarities ranging from 97.9 to 99.8%. Phylogenetic trees were constructed based on the 16S rDNA, ITS and 23S rDNA sequence data and compared. The trees were topologically similar, suggesting that the 3 types of molecules provided similar phylogenetic information. Five of the isolates are closely related (> 99.4% 16S rDNA similarity, 99.1% to 99.7% ITS and 99.3 to 99.8% 23S rDNA similarities). The sequence of one Chilean isolate, EM-90, was unique, with 16S rDNA similarities to the other isolates ranging from 98.5 to 98.9%, the ITS from 95.2 to 96.9% and the 23S rDNA from 97.6 to 98.5%. (+info)
Degradation of 1,2-dibromoethane by Mycobacterium sp. strain GP1.
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The newly isolated bacterial strain GP1 can utilize 1, 2-dibromoethane as the sole carbon and energy source. On the basis of 16S rRNA gene sequence analysis, the organism was identified as a member of the subgroup which contains the fast-growing mycobacteria. The first step in 1,2-dibromoethane metabolism is catalyzed by a hydrolytic haloalkane dehalogenase. The resulting 2-bromoethanol is rapidly converted to ethylene oxide by a haloalcohol dehalogenase, in this way preventing the accumulation of 2-bromoethanol and 2-bromoacetaldehyde as toxic intermediates. Ethylene oxide can serve as a growth substrate for strain GP1, but the pathway(s) by which it is further metabolized is still unclear. Strain GP1 can also utilize 1-chloropropane, 1-bromopropane, 2-bromoethanol, and 2-chloroethanol as growth substrates. 2-Chloroethanol and 2-bromoethanol are metabolized via ethylene oxide, which for both haloalcohols is a novel way to remove the halide without going through the corresponding acetaldehyde intermediate. The haloalkane dehalogenase gene was cloned and sequenced. The dehalogenase (DhaAf) encoded by this gene is identical to the haloalkane dehalogenase (DhaA) of Rhodococcus rhodochrous NCIMB 13064, except for three amino acid substitutions and a 14-amino-acid extension at the C terminus. Alignments of the complete dehalogenase gene region of strain GP1 with DNA sequences in different databases showed that a large part of a dhaA gene region, which is also present in R. rhodochrous NCIMB 13064, was fused to a fragment of a haloalcohol dehalogenase gene that was identical to the last 42 nucleotides of the hheB gene found in Corynebacterium sp. strain N-1074. (+info)
Rapid and easy detection of Helicobacter pylori by in situ hybridization.
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Various in situ hybridization (ISH) methods have been used to identify Helicobacter pylori, a causative organism responsible for chronic gastritis and peptic ulcer disease, but they were hard to perform and time consuming. To detect H. pylori in a rapid and easily reproducible way, we developed synthetic biotinylated oligonucleotide probes which complement rRNA of H. pylori. Formalin-fixed and paraffin-embedded tissues from 50 gastric biopsy specimens were examined. Using a serologic test and histochemical stain (Warthin-Starry silver stain and/or Giemsa stain) as a standard, 40 of them were confirmed to be H. pylori-positive. Our ISH was quickly carried out within one hr and results were compared with those obtained from immunohistochemical stain. The ISH produced a positive reaction in 38 of 40 cases (95%). All H. pylori-negative cases failed to demonstrate a positive signal. The ISH has a sensitivity comparable to those of conventional histochemical and immunohistochemical stain, and has high specificity. In conclusion, ISH with a biotinylated oligonucleotide probe provides a useful diagnostic method for detecting H. pylori effectively in routinely processed tissue sections. (+info)
Formation of hydride-Meisenheimer complexes of picric acid (2,4, 6-trinitrophenol) and 2,4-dinitrophenol during mineralization of picric acid by Nocardioides sp. strain CB 22-2.
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There are only a few examples of microbial conversion of picric acid (2,4,6-trinitrophenol). None of the organisms that have been described previously is able to use this compound as a sole source of carbon, nitrogen, and energy at high rates. In this study we isolated and characterized a strain, strain CB 22-2, that was able to use picric acid as a sole source of carbon and energy at concentrations up to 40 mM and at rates of 1.6 mmol. h(-1). g (dry weight) of cells(-1) in continuous cultures and 920 micromol. h(-1). g (dry weight) of cells(-1) in flasks. In addition, this strain was able to use picric acid as a sole source of nitrogen at comparable rates in a nitrogen-free medium. Biochemical characterization and 16S ribosomal DNA analysis revealed that strain CB 22-2 is a Nocardioides sp. strain. High-pressure liquid chromatography and UV-visible light data, the low residual chemical oxygen demand, and the stoichiometric release of 2.9 +/- 0.1 mol of nitrite per mol of picric acid provided strong evidence that complete mineralization of picric acid occurred. During transformation, the metabolites detected in the culture supernatant were the [H-]-Meisenheimer complexes of picric acid and 2,4-dinitrophenol (H--DNP), as well as 2,4-dinitrophenol. Experiments performed with crude extracts revealed that H--DNP formation indeed is a physiologically relevant step in picric acid metabolism. (+info)
Geographic distribution and genetic diversity of Ceanothus-infective Frankia strains.
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Little is known about Ceanothus-infective Frankia strains because no Frankia strains that can reinfect the host plants have been isolated from Ceonothus spp. Therefore, we studied the diversity of the Ceonothus-infective Frankia strains by using molecular techniques. Frankia strains inhabiting root nodules of nine Ceanothus species were characterized. The Ceanothus species used represent the taxonomic diversity and geographic range of the genus; therefore, the breadth of the diversity of Frankia strains that infect Ceanothus spp. was studied. DNA was amplified directly from nodular material by using the PCR. The amplified region included the 3' end of the 16S rRNA gene, the intergenic spacer, and a large portion of the 23S rRNA gene. A series of restriction enzyme digestions of the PCR product allowed us to identify PCR-restriction fragment length polymorphism (RFLP) groups among the Ceanothus-infective Frankia strains tested. Twelve different enzymes were used, which resulted in four different PCR-RFLP groups. The groups did not follow the taxonomic lines of the Ceanothus host species. Instead, the Frankia strains present were related to the sample collection locales. (+info)