Phenotypic and genotypic characterization of tetracycline and minocycline resistance in Clostridium perfringens.

The aim of this study was to determine the incidence of tetracycline resistance and the prevalence of tetracycline-resistance genes in strains of Clostridium perfringens isolated from different sources between 1994 and 2005. Susceptibility to tetracycline and minocycline in strains from humans (35 isolates), chickens (15 isolates), food (21 isolates), soil (16 isolates) and veterinary sources (6 isolates) was determined, and tetracycline-resistance genes were detected. Resistance was most common in strains isolated from chickens, followed by those from soils, clinical samples and foods. The most highly resistant strains were found among clinical and food isolates. tetA(P) was the most common resistance gene, and along with tetB(P) was found in all resistant strains and some sensitive strains. One tetracycline-resistant food isolate had an intact tet(M) gene. However, PCR fragments of 0.4 or 0.8 kb with high degrees of identity to parts of the tet(M) sequences of other bacteria were found, mainly in clinical isolates, and often in isolates with tetB(P). No correlation between level of sensitivity to tetracycline or minocycline and the presence of tetA(P), tetB(P) or part of tet(M) was found. The presence of part of tet(M) in some strains of C. perfringens containing tetB(P) may have occurred by recent gene transfer.

Sudan azo dyes and Para Red degradation by prevalent bacteria of the human gastrointestinal tract.

Sudan azo dyes have genotoxic effects and ingestion of food products contaminated with Sudan I, II, III, IV, and Para Red could lead to exposure in the human gastrointestinal tract. In this study, we examined thirty-five prevalent species of human intestinal bacteria to evaluate their capacity to degrade Sudan dyes and Para Red. Among these tested bacterial strains, 23, 13, 33, 30, and 29 out of 35 species tested were able to reduce Sudan I, II, III, IV, and Para Red, respectively, to some extent. Bifidobacterium infantis, Clostridium indolis, Enterococcus faecalis, Lactobacillus rhamnosus, and Ruminococcus obeum were able to reduce completely all four tested Sudan dyes and Para Red. Escherichia coli and Peptostreptococcus magnus were the only two strains that were not able to reduce any of the tested Sudan dyes and Para Red to any significant extent. Metabolites of the reduction of the tested Sudan dyes and Para Red by E. faecalis were isolated and identified by HPLC and LC/ESI-MS analyses and compared with authentic standards. Thus it appears that the ability to reduce Sudan dyes and Para Red except Sudan II is common among bacteria in the human colon.

Mechanism of metronidazole-resistance by isolates of nitroreductase-producing Enterococcus gallinarum and Enterococcus casseliflavus from the human intestinal tract.

Enterococcus casseliflavus and Enterococcus gallinarum strains resistant to metronidazole, nitrofurantoin and nitrofurazone were isolated from fecal samples of a patient with recurrent ulcerative colitis treated with metronidazole. Unlike other metronidazole-resistant bacteria, these strains produced nitroreductase but metabolized metronidazole to compounds that could not be detected by liquid chromatography with UV or mass spectral analysis. Metronidazole-susceptible Clostridium perfringens grew equally well in spent cultures of Enterococcus spp. incubated with or without metronidazole. These data indicate that the nitroreductases produced by these Enterococcus strains did not activate metronidazole to bactericidal metabolites and these bacteria may reduce the effectiveness of metronidazole. We have indirect evidence for an alternative pathway that results in metronidazole resistance. These strains of enterococcus had nitroreductase so resistance should not have occurred.

Microarray method to monitor 40 intestinal bacterial species in the study of azo dye reduction.

Azo dyes are widely used in dye manufacturing, paper printing, textile industries, and as tattoo pigmentation. Since intestinal and skin bacteria can metabolize certain azo dyes to carcinogenic compounds, many researchers have studied the azoreductases of these bacteria. In this study, we used a microarray method to identify the intestinal bacterial species from cultured fecal samples in Brain Heart Infusion (BHI) broth with or without azo dyes that may be involved in azo dye reduction. The microarray was designed to identify 40 bacterial species that are reported in the literature to be predominant in human feces. Results from this study showed 26-30 species are present in the cultured fecal samples. The representative bacteria were then examined for the azo dye reduction activity.

Classification of a polycyclic aromatic hydrocarbon-metabolizing bacterium, Mycobacterium sp. strain PYR-1, as Mycobacterium vanbaalenii sp. nov.

A polycyclic aromatic hydrocarbon (PAH)-utilizing Mycobacterium strain, PYR-1(T), was isolated from petroleum-contaminated estuarine sediments and has been shown by 16S rRNA gene sequencing to be closely related to Mycobacterium aurum ATCC 23366(T) and Mycobacterium vaccae ATCC 15438(T). In this investigation, the 16S rDNA, fatty acid methyl esters, DNA-DNA hybridization, PFGE analysis of restriction-digested total genomic DNA and biochemical tests were used to determine the taxonomic relationship of strain PYR-1(T) to other closely related Mycobacterium species. The sequence of the 16S rRNA gene of strain PYR-1(T) was similar to that of Mycobacterium austroafricanum ATCC 33464(T), except for one gap at position 43. Fatty acid methyl ester analysis also showed similarity to M. austroafricanum ATCC 33464(T); however, the Euclidean distance was greater than 4.0, indicating that these strains were not identical. Dot-blot DNA-DNA hybridization of strain PYR-1(T) with M. austroafricanum indicated less than 40% relatedness. When the total chromosomal DNA of M. aurum ATCC 23366(T), M. austroafricanum ATCC 33464(T) and strain PYR-1(T) was digested with restriction enzyme Xbal and analysed by PFGE, all three organisms gave different restriction patterns. Previous studies from our laboratory have shown that the reverse-phase HPLC elution profiles of mycolic acids of strain PYR-1(T) and M. austroafricanum ATCC 33464(T) have different patterns. Based on phylogenetic analysis using 165 rRNA gene sequences, fatty acid analysis, DNA-DNA hybridization and PFGE analysis and physiological and chemotaxonomic characteristics, it is concluded that strain PYR-1(T) (= DSM 7251(T) = NRRL B-24157(T)) represents a novel species of the genus Mycobacterium, for which the name Mycobacterium vanbaalenii sp. nov. is proposed.