Assignment of the Bacterial Agent of Urinary Calculus in Young Rats by the Comparative Sequence Analysis of the 16S rRNA Genes of Corynebacteria.

Nontuberculous mycobacteria (NTM) have emerged as an important cause of opportunistic nosocomial infections. NTM has frequently been isolated from hospital water distribution systems. The aim of this study was to survey the risk of NTM infections and determine the prevalence of NTM species in the hospital water distribution systems in Tabriz, Iran. One hundred and twenty samples of water from different sources of Tabriz hospitals were collected. The samples were filtered through 0.45-µm pore size membranes and decontaminated with 0.01% cetylpyridinium chloride. The sediment was inoculated onto Lowenstein–Jensen medium and incubated for 8 weeks. For identification to the species level, partial sequence analysis of the hsp65 and 16S rRNA genes were used. NTM were detected in 76 (63.3%) of 120 samples. Potentially pathogenic mycobacteria and saprophytic mycobacteria were isolated. Mycobacterium gordonae was the only single species that was present in all types of water. The prevalence of NTM in Tabriz hospitals' water compared with many investigations on hospital waters was high. This indicates that the immunocompromised patients and transplant recipients are at risk of contamination which necessitates considering decontamination of water sources to prevent such potential hazards.

Download Full-text

Multilocus sequence analysis of Ensifer and related taxa

INTERNATIONAL JOURNAL OF SYSTEMATIC AND EVOLUTIONARY MICROBIOLOGY ◽

10.1099/ijs.0.64344-0 ◽

2007 ◽

Vol 57 (3) ◽

pp. 489-503 ◽

Cited By ~ 180

Author(s):

Miet Martens ◽

Manuel Delaere ◽

Renata Coopman ◽

Paul De Vos ◽

Monique Gillis ◽

...

Keyword(s):

Sequence Analysis ◽

16S Rrna ◽

Housekeeping Genes ◽

16S Rrna Genes ◽

New Combination ◽

Rrna Genes ◽

Multilocus Sequence Analysis ◽

Bootstrap Support ◽

Rrna Gene ◽

Separate Species

Multilocus sequence analysis (MLSA) was performed on representatives of Ensifer (including species previously assigned to the genus Sinorhizobium) and related taxa. Neighbour-joining (NJ), maximum-parsimony (MP) and maximum-likelihood (ML) phylogenies of dnaK, gltA, glnA, recA, thrC and 16S rRNA genes were compared. The data confirm that the potential for discrimination of Ensifer species is greater using MLSA of housekeeping genes than 16S rRNA genes. In incongruence-length difference tests, the 16S rRNA gene was found to be significantly incongruent with the other genes, indicating that this gene should not be used as a single indicator of relatedness in this group. Significant congruence was detected for dnaK, glnA and thrC. Analyses of concatenated sequences of dnaK, glnA and thrC genes yielded very similar NJ, MP and ML trees, with high bootstrap support. In addition, analysis of a concatenation of all six genes essentially produced the same result, levelling out potentially conflicting phylogenetic signals. This new evidence supports the proposal to unite Ensifer and Sinorhizobium in a single genus. Support for an alternative solution preserving the two genera is less strong. In view of the opinions expressed by the Judicial Commission, the name of the genus should be Ensifer, as proposed by Young [Young, J. M. (2003). Int J Syst Evol Microbiol 53, 2107–2110]. Data obtained previously and these new data indicate that Ensifer adhaerens and ‘Sinorhizobium morelense’ are not heterotypic synonyms, but represent separate species. However, transfer to the genus Ensifer is not possible at present because the species name is the subject of a pending Request for an Opinion, which would affect whether a novel species in the genus Ensifer or a new combination based on a basonym would be created.

Download Full-text

The Influence of DNA Extraction Procedure and Primer Set on the Bacterial Community Analysis by Pyrosequencing of Barcoded 16S rRNA Gene Amplicons

Molecular Biology International ◽

10.1155/2014/548683 ◽

2014 ◽

Vol 2014 ◽

pp. 1-10 ◽

Cited By ~ 9

Author(s):

Ingo C. Starke ◽

Wilfried Vahjen ◽

Robert Pieper ◽

Jürgen Zentek

Keyword(s):

Sequence Analysis ◽

16S Rrna ◽

Dna Extraction ◽

16S Rrna Genes ◽

Read Length ◽

Rrna Genes ◽

Rrna Gene ◽

Variable Regions ◽

Extraction Procedures ◽

Primer Sets

In this study, the effect of different DNA extraction procedures and primer sets on pyrosequencing results regarding the composition of bacterial communities in the ileum of piglets was investigated. Ileal chyme from piglets fed a diet containing different amounts of zinc oxide was used to evaluate a pyrosequencing study with barcoded 16S rRNA PCR products. Two DNA extraction methods (bead beating versus silica gel columns) and two primer sets targeting variable regions of bacterial 16S rRNA genes (8f-534r versus 968f-1401r) were considered. The SEED viewer software of the MG-RAST server was used for automated sequence analysis. A total of 5.2×105 sequences were used for analysis after processing for read length (150 bp), minimum sequence occurrence (5), and exclusion of eukaryotic and unclassified/uncultured sequences. DNA extraction procedures and primer sets differed significantly in total sequence yield. The distribution of bacterial order and main bacterial genera was influenced significantly by both parameters. However, this study has shown that the results of pyrosequencing studies using barcoded PCR amplicons of bacterial 16S rRNA genes depend on DNA extraction and primer choice, as well as on the manner of downstream sequence analysis.

Download Full-text

Comparative Genetic Analysis of Mycobacterium ulcerans and Mycobacterium marinum Reveals Evidence of Recent Divergence

Journal of Bacteriology ◽

10.1128/jb.182.22.6322-6330.2000 ◽

2000 ◽

Vol 182 (22) ◽

pp. 6322-6330 ◽

Cited By ~ 96

Author(s):

Timothy P. Stinear ◽

Grant A. Jenkin ◽

Paul D. R. Johnson ◽

John K. Davies

Keyword(s):

Sequence Analysis ◽

16S Rrna ◽

Genetic Relationship ◽

Mycobacterium Marinum ◽

Mycobacterium Ulcerans ◽

16S Rrna Genes ◽

Rrna Genes ◽

Mobile Dna ◽

Apparent Paradox ◽

Close Genetic Relationship

ABSTRACT Previous studies of the 16S rRNA genes from Mycobacterium ulcerans and Mycobacterium marinum have suggested a very close genetic relationship between these species (99.6% identity). However, these organisms are phenotypically distinct and cause diseases with very different pathologies. To investigate this apparent paradox, we compared 3,306 nucleotides from the partial sequences of eight housekeeping and structural genes derived from 18M. ulcerans strains and 22 M. marinumstrains. This analysis confirmed the close genetic relationship inferred from the 16S rRNA data, with nucleotide sequence identity ranging from 98.1 to 99.7%. The multilocus sequence analysis also confirmed previous genotype studies of M. ulceransthat have identified distinct genotypes within a geographical region. Single isolates of both M. ulcerans and M. marinum that were shown by the sequence analysis to be the most closely related were then selected for further study. One- and two-dimensional pulsed-field gel electrophoresis was employed to compare the architecture and size of the genome from each species. Genome sizes of approximately 4.4 and 4.6 Mb were obtained forM. ulcerans and M. marinum, respectively. Significant macrorestriction fragment polymorphism was observed between the species. However, hybridization analysis of DNA cleaved with more frequently cutting enzymes identified significant preservation of the flanking sequence at seven of the eight loci sequenced. The exception was the 16S rRNA locus. Two high-copy-number insertion sequences, IS2404 and IS2606, have recently been reported in M. ulcerans, and significantly, these elements are not present in M. marinum. Hybridization of theAseI restriction fragments from M. ulcerans with IS2404 and IS2606 indicated widespread genome distribution for both of these repeated sequences. Taken together, these data strongly suggest that M. ulcerans has recently diverged from M. marinumby the acquisition and concomitant loss of DNA in a manner analogous to the emergence of M. tuberculosis, where species diversity is being driven mainly by the activity of mobile DNA elements.

Download Full-text