Dataset Information

Analysis of Nucleotide Sequences of the 16S rRNA Gene of Novel Escherichia coli Strains Isolated from Feces of Human and Bali Cattle.

ABSTRACT: Livestock especially cattle are known as a main reservoir of Escherichia coli O157:H7. This bacterium is considered as a pathogenic agent characterized by producing toxins, which are familiarly known as Shiga-like toxin-1 (Stx1) and Stx2. The aim of this work was to analyse the novel sequence of the 16S rRNA gene of strains isolated in this study in order to know the phylogenetic relationships between these sequences and those between the sequences of bacteria available in databanks. The results of this analysis showed that the strains KL-48(2) and SM25(1) that originated from human and cattle feces, respectively, are closely related among them and with respect to E. coli EDL 933, E. coli Sakai, E. coli ATCC 43894, E. coli O111:H-, E. coli O121:H19, E. coli O104:H4, and Shigella sonnei with more than 99% similarity values.

SUBMITTER: Suardana IW

PROVIDER: S-EPMC4172979 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature

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Analysis of Nucleotide Sequences of the 16S rRNA Gene of Novel Escherichia coli Strains Isolated from Feces of Human and Bali Cattle.

Suardana I Wayan IW

Journal of nucleic acids 20140909

Livestock especially cattle are known as a main reservoir of Escherichia coli O157:H7. This bacterium is considered as a pathogenic agent characterized by producing toxins, which are familiarly known as Shiga-like toxin-1 (Stx1) and Stx2. The aim of this work was to analyse the novel sequence of the 16S rRNA gene of strains isolated in this study in order to know the phylogenetic relationships between these sequences and those between the sequences of bacteria available in databanks. The results ...[more]

PMID: 25276419

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Project description:In several developing countries, studies on antimicrobial resistance among bacteria from food animals are rare mostly because of under-resourced laboratories. The objective of this study was to develop and field-test a low cost protocol to estimate the isolate- and sample-level prevalence of resistance to critically important antibiotics among Escherichia coli and Salmonella isolated from dairy cattle feces. Using a predesigned protocol, fecal samples were collected to isolate non-type-specific E. coli and Salmonella using selective media without antibiotic supplements. Besides, samples were screened for E. coli and Salmonella isolates not susceptible to third-generation cephalosporins and quinolones using selective media supplemented with cefotaxime (1.0 μg/mL) and ciprofloxacine (0.5 μg/mL), respectively. All bacterial isolates were further tested for antibiotic susceptibility using disk diffusion. Bacterial isolates not susceptible to third-generation cephalosporins were tested for extended spectrum beta-lactamase (ESBL) phenotype using the combination disk test. Molecular methods were performed on selected bacterial isolates to identify and distinguish genetic determinants associated with the observed phenotypes. Among 85 non-type-specific E. coli isolated from MacConkey agar without antibiotics, the isolate-level prevalence of resistance to tetracycline was the highest (8.2%). Among 37 E. coli recovered from MacConkey agar with cefotaxime, 56.8% were resistant ceftriaxone. Among 22 E. coli isolates recovered from MacConkey agar with ciprofloxacin, 77.3% and 54.5% were resistant to nalidixic acid and ciprofloxacin, respectively. Sixteen Salmonella were isolated and only one demonstrated any resistance (i.e., single resistance to streptomycin). Among E. coli isolates not susceptible to ceftriaxone, an AmpC phenotype was more common than an ESBL phenotype (29 versus 10 isolates, respectively). Whole genome sequencing showed that phenotypic profiles of antibiotic resistance detected were generally substantiated by genotypic profiles. The tested protocol is suited to detecting and estimating prevalence of antimicrobial resistance in bacteria isolated from food animal feces in resource-limited laboratories in the developing world.

Project description:16S rRNA gene sequences are commonly analyzed for taxonomic and phylogenetic studies because they contain variable regions that can help distinguish different genera. However, intra-genus distinction using variable region homology is often impossible due to the high overall sequence identities among closely related species, even though some residues may be conserved within respective species. Using a computational method that included the allelic diversity within individual genomes, we discovered that certain Escherichia and Shigella species can be distinguished by a multi-allelic 16S rRNA variable region single nucleotide polymorphism (SNP). To evaluate the performance of 16S rRNAs with altered variable regions, we developed an in vivo system that measures the acceptance and distribution of variant 16S rRNAs into a large pool of natural versions supporting normal translation and growth. We found that 16S rRNAs containing evolutionarily disparate variable regions were underpopulated both in ribosomes and in active translation pools, even for an SNP. Overall, this study revealed that variable region sequences can substantially influence the performance of 16S rRNAs and that this biological constraint can be leveraged to justify refining taxonomic assignments of variable region sequence data. IMPORTANCE This study reevaluates the notion that 16S rRNA gene variable region sequences are uninformative for intra-genus classification and that single nucleotide variations within them have no consequence to strains that bear them. We demonstrated that the performance of 16S rRNAs in Escherichia coli can be negatively impacted by sequence changes in variable regions, even for single nucleotide changes that are native to closely related Escherichia and Shigella species; thus, biological performance is likely constraining the evolution of variable regions in bacteria. Further, the native nucleotide variations we tested occur in all strains of their respective species and across their multiple 16S rRNA gene copies, suggesting that these species evolved beyond what would be discerned from a consensus sequence comparison. Therefore, this work also reveals that the multiple 16S rRNA gene alleles found in most bacteria can provide more informative phylogenetic and taxonomic detail than a single reference allele.

Dataset Information

Analysis of Nucleotide Sequences of the 16S rRNA Gene of Novel Escherichia coli Strains Isolated from Feces of Human and Bali Cattle.

Publications

Analysis of Nucleotide Sequences of the 16S rRNA Gene of Novel Escherichia coli Strains Isolated from Feces of Human and Bali Cattle.

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