Evaluation of the Ribosomal Protein S1 Gene (rpsA) as a Novel Biomarker forMycobacteriumSpecies Identification

Objectives. To evaluate the resolution and reliability of therpsAgene, encoding ribosomal protein S1, as a novel biomarker for mycobacteria species identification.Methods. A segment of therpsAgene (565 bp) was amplified by PCR from 42 mycobacterial reference strains, 172 nontuberculosis mycobacteria clinical isolates, and 16M. tuberculosiscomplex clinical isolates. The PCR products were sequenced and aligned by using the multiple alignment algorithm in the MegAlign package (DNASTAR) and the MEGA program. A phylogenetic tree was constructed by the neighbor-joining method.Results. Comparative sequence analysis of therpsAgene provided the basis for species differentiation within the genusMycobacterium. Slow- and rapid-growing groups of mycobacteria were clearly separated, and each mycobacterial species was differentiated as a distinct entity in the phylogenetic tree. The sequences discrepancy was obvious betweenM. kansasiiandM. gastri, M. chelonaeandM. abscessus, M. aviumandM. intracellulare, andM. szulgaiandM. malmoense, which cannot be achieved by 16S ribosomal DNA (rDNA) homologue genes comparison. 183 of the 188 (97.3%) clinical isolates, consisting of 8 mycobacterial species, were identified correctly byrpsAgene blast.Conclusions. Our study indicates thatrpsAsequencing can be used effectively for mycobacteria species identification as a supplement to 16S rDNA sequence analysis.

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Identification of Mycobacterial Species by Comparative Sequence Analysis of the RNA Polymerase Gene (rpoB)

Journal of Clinical Microbiology ◽

10.1128/jcm.37.6.1714-1720.1999 ◽

1999 ◽

Vol 37 (6) ◽

pp. 1714-1720 ◽

Cited By ~ 244

Author(s):

Bum-Joon Kim ◽

Seung-Hyun Lee ◽

Mi-Ae Lyu ◽

Seo-Jeong Kim ◽

Gill-Han Bai ◽

...

Keyword(s):

Sequence Analysis ◽

Rna Polymerase ◽

Phylogenetic Tree ◽

Comparative Sequence Analysis ◽

Clinical Isolates ◽

Species Differentiation ◽

Rrna Gene ◽

Alignment Algorithm ◽

Mycobacterial Species ◽

Comparative Sequence

For the differentiation and identification of mycobacterial species, the rpoB gene, encoding the β subunit of RNA polymerase, was investigated. rpoB DNAs (342 bp) were amplified from 44 reference strains of mycobacteria and clinical isolates (107 strains) by PCR. The nucleotide sequences were directly determined (306 bp) and aligned by using the multiple alignment algorithm in the MegAlign package (DNASTAR) and the MEGA program. A phylogenetic tree was constructed by the neighbor-joining method. Comparative sequence analysis of rpoB DNAs provided the basis for species differentiation within the genusMycobacterium. Slowly and rapidly growing groups of mycobacteria were clearly separated, and each mycobacterial species was differentiated as a distinct entity in the phylogenetic tree. Pathogenic Mycobacterium kansasii was easily differentiated from nonpathogenic M. gastri; this differentiation cannot be achieved by using 16S rRNA gene (rDNA) sequences. By being grouped into species-specific clusters with low-level sequence divergence among strains of the same species, all of the clinical isolates could be easily identified. These results suggest that comparative sequence analysis of amplified rpoB DNAs can be used efficiently to identify clinical isolates of mycobacteria in parallel with traditional culture methods and as a supplement to 16S rDNA gene analysis. Furthermore, in the case of M. tuberculosis, rifampin resistance can be simultaneously determined.

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The Evolution and Sequence Diversity of FhuA inSalmonellaandEscherichia

10.1101/320069 ◽

2018 ◽

Author(s):

Yejun Wang ◽

Xiongbin Chen ◽

Guoqiang Zhu ◽

Aaron P. White ◽

Wolfgang Köster

Keyword(s):

Sequence Analysis ◽

Phylogenetic Tree ◽

Gene Mutations ◽

Comparative Sequence Analysis ◽

Sequence Diversity ◽

Bacterial Cells ◽

Bacterial Survival ◽

Bacterial Strains ◽

Evolutionary Patterns ◽

Potential Ligand

ABSTRACTThefhuACDBoperon, present in a number ofEnterobacteriaceae, encodes components essential for the uptake of ferric hydroxamate type siderophores. FhuA acts not only as transporter for physiologically important chelated ferric iron, but also as receptor for various bacteriophages, toxins and antibiotics, which are pathogenic to bacterial cells. In this research, thefhuAgene distribution and sequence diversity were investigated inEnterobacteriaceae, especiallySalmonellaandEscherichia. Comparative sequence analysis resulted in afhuAphylogenetic tree that did not match the expected phylogeny based on housekeeping sequence analysis or trees offhuCDBgenes. ThefhuAsequences showed a unique mosaic-clustering pattern. On the other hand, the gene sequences showed high conservation for strains from the same serovar or serotype. In total, six clusters were identified from FhuA proteins inSalmonellaandEscherichia, among which typical peptide fragment variations could be defined. Six fragmental insertions / deletions and two substitution fragments were discovered, which could well classify the different clusters. Structure modeling demonstrated that all the six featured insertions/deletions and one substitution fragment are located at the apexes of the long loops of FhuA external pocket. These frequently mutated regions are likely under high selection pressure, and bacterial strains could have escaped from phage infection or toxin / antibiotics attack viafhuAgene mutations while maintaining the siderophore uptake activity essential for bacterial survival. The unusualfhuAclustering suggests that high frequency exchange offhuAgenes has occurred between enterobacterial strains after distinctive species were established.IMPORTANCEThe enterobacterialfhuACDBoperon encodes proteins which mediate the uptake of siderophores to supply the cells with iron essential for bacterial survival. Here we show different evolutionary patterns for thefhugenes within the same operon. ThefhuAhas a phylogenetic tree that does not match the species phylogeny, whereas the rest of thefhugenes do. ThefhuAgenes showed inter-species sequence convergence and conservation within specific serovars and serotypes. Nearly all of the significant sequence differences among FhuA clusters are located in potential ligand-binding sites on the extracellular surface of fhuA-encoding receptors. The unusualfhuAclustering suggests the frequent recombination and exchange offhuAgenes between enterobacterial strains in the evolutionary state after distinctive species were established.Our findings suggested either a new evolutionary mechanism or local gene recombination infhuAthat is in contrast to previous evolutionary hypotheses that have formed under the assumption of no recombination.

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E. coli translation initiation factor IF2 - an extremely conserved protein. Comparative sequence analysis of the infB gene in clinical isolates of E. coli

FEBS Letters ◽

10.1016/s0014-5793(97)01472-5 ◽

1997 ◽

Vol 419 (2-3) ◽

pp. 281-284 ◽

Cited By ~ 14

Author(s):

Søren A de A. Steffensen ◽

Ane B Poulsen ◽

Kim K Mortensen ◽

Hans U Sperling-Petersen

Keyword(s):

Sequence Analysis ◽

Translation Initiation ◽

Comparative Sequence Analysis ◽

Translation Initiation Factor ◽

Clinical Isolates ◽

Initiation Factor ◽

E Coli ◽

Comparative Sequence

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Typing of Candida glabrata in Clinical Isolates by Comparative Sequence Analysis of the Cytochrome c Oxidase Subunit 2 Gene Distinguishes Two Clusters of Strains Associated with Geographical Sequence Polymorphisms

Journal of Clinical Microbiology ◽

10.1128/jcm.38.1.227-235.2000 ◽

2000 ◽

Vol 38 (1) ◽

pp. 227-235

Author(s):

Gerdine F. O. Sanson ◽

Marcelo R. S. Briones

Keyword(s):

United States ◽

Sequence Analysis ◽

Phylogenetic Relationships ◽

Candida Glabrata ◽

The United States ◽

Comparative Sequence Analysis ◽

Clinical Isolates ◽

Coding Region ◽

Comparative Sequence ◽

Oxidase Subunit

ABSTRACT We tested whether comparative sequence analysis of the mitochondrion-encoded cytochrome c oxidase subunit 2 gene ( COX2 ) could be used to distinguish intraspecific variants of Candida glabrata . Mitochondrial genes are suitable for investigation of close phylogenetic relationships because they evolve much faster than nuclear genes, which in general exhibit very limited intraspecific variation. For this survey we used 11 clinical isolates of C. glabrata from three different geographical locations in Brazil, 10 isolates from one location in the United States, 1 American Type Culture Collection strain as an internal control, and the published sequence of strain CBS 138. The complete coding region of COX2 was amplified from total cellular DNA, and both strands were sequenced twice for each strain. These sequences were aligned with published sequences from other fungi, and the numbers of substitutions and phylogenetic relationships were determined. Typing of these strains was done by using 17 substitutions, with 8 being nonsynonymous and 9 being synonymous. Also, cDNAs made from purified mitochondrial polyadenylated RNA were sequenced to confirm that our sequences correspond to the expressed copies and not nuclear pseudogenes and that a frameshift mutation exists in the 3′ end of the coding region (position 673) relative to the Saccharomyces cerevisiae sequence and the previously published C. glabrata sequence. We estimated the average evolutionary rate of COX2 to be 11.4% sequence divergence/10 8 years and that phylogenetic relationships of yeasts based on these sequences are consistent with rRNA sequence data. Our analysis of COX2 sequences enables typing of C. glabrata strains based on 13 haplotypes and suggests that positions 51 and 519 indicate a geographical polymorphism that discriminates strains isolated in the United States and strains isolated in Brazil. This provides for the first time a means of typing of Candida strains that cause infections by use of direct sequence comparisons and the associated divergence estimates.

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