Dataset Information

Analysis, characterization, and loci of the tuf genes in lactobacillus and bifidobacterium species and their direct application for species identification.

ABSTRACT: We analyzed the tuf gene, encoding elongation factor Tu, from 33 strains representing 17 Lactobacillus species and 8 Bifidobacterium species. The tuf sequences were aligned and used to infer phylogenesis among species of lactobacilli and bifidobacteria. We demonstrated that the synonymous substitution affecting this gene renders elongation factor Tu a reliable molecular clock for investigating evolutionary distances of lactobacilli and bifidobacteria. In fact, the phylogeny generated by these tuf sequences is consistent with that derived from 16S rRNA analysis. The investigation of a multiple alignment of tuf sequences revealed regions conserved among strains belonging to the same species but distinct from those of other species. PCR primers complementary to these regions allowed species-specific identification of closely related species, such as Lactobacillus casei group members. These tuf gene-based assays developed in this study provide an alternative to present methods for the identification for lactic acid bacterial species. Since a variable number of tuf genes have been described for bacteria, the presence of multiple genes was examined. Southern analysis revealed one tuf gene in the genomes of lactobacilli and bifidobacteria, but the tuf gene was arranged differently in the genomes of these two taxa. Our results revealed that the tuf gene in bifidobacteria is flanked by the same gene constellation as the str operon, as originally reported for Escherichia coli. In contrast, bioinformatic and transcriptional analyses of the DNA region flanking the tuf gene in four Lactobacillus species indicated the same four-gene unit and suggested a novel tuf operon specific for the genus Lactobacillus.

SUBMITTER: Ventura M

PROVIDER: S-EPMC262312 | biostudies-literature | 2003 Nov

REPOSITORIES: biostudies-literature

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Analysis, characterization, and loci of the tuf genes in lactobacillus and bifidobacterium species and their direct application for species identification.

Ventura Marco M Canchaya Carlos C Meylan Valèrie V Klaenhammer Todd R TR Zink Ralf R

Applied and environmental microbiology 20031101 11

We analyzed the tuf gene, encoding elongation factor Tu, from 33 strains representing 17 Lactobacillus species and 8 Bifidobacterium species. The tuf sequences were aligned and used to infer phylogenesis among species of lactobacilli and bifidobacteria. We demonstrated that the synonymous substitution affecting this gene renders elongation factor Tu a reliable molecular clock for investigating evolutionary distances of lactobacilli and bifidobacteria. In fact, the phylogeny generated by these tu ...[more]

PMID: 14602655

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Project description:IntroductionCoagulase-negative staphylococci (CoNS) are normal inhabitants of human skin and mucous membranes. However, CoNS represent one of the major nosocomial pathogens, especially in immunocompromised patients. The increasing incidence of CoNS and mainly methicillin-resistant strains underlines the need for an accurate identification of Staphylococcus isolates at the species level. Analysis of the tuf gene proved to be an accurate tool for the species identification of CoNS. The aims of this study were to identify the CoNS species by tuf gene-based polymerase chain reaction method and sequencing, and to determine the frequency of CoNS clinical isolates resistant to methicillin (MRCoNS) and other antibiotics.MethodsA total of 200 staphylococci isolates were collected from various clinical samples. Phenotyping methods were used for initial identification followed by polymerase chain reaction amplification of tuf gene with subsequent sequencing. The phylogenetic relationships among species were analyzed using the neighbor-joining method based on the partial gene sequence of tuf. Microbroth dilution test was used for screening methicillin resistance, and disk diffusion susceptibility testing was performed for evaluation of antibiotic resistance among the isolates.ResultsIn the present study, 125 isolates were identified as CoNS; among them, Staphylococcus epidermidis 54(43.2%) and Staphylococcus haemolyticus 50 (40.0%) were demonstrated as the most prevalent species. Resistance to methicillin was detected in 54.4% of the CoNS based on microbroth dilution method. In disk diffusion susceptibility testing, the greatest resistance of CoNS was demonstrated for cefoxitin (65.4%), cotrimethoxazole (54.4%), and clindamycin (49.6%), while daptomycin (87.2%) and linezolid (83.2%) showed the greatest effectiveness for CoNS isolates.ConclusionOur results confirmed the predominance of S. epidermidis and S. haemolyticus among CoNS isolates. The high prevalence of MRCoNS strains is a serious concern and strongly suggests the need for control program measures in our hospitals in order to reduce MRCoNS infections, especially in immunocompromised patients.

Project description:Members of the bacterial genus Bifidobacterium generally dominate the fecal microbiota of infants. The species Bifidobacterium longum is prevalent, but the B. longum subsp. longum and B. longum subsp. infantis strains that are known to colonize the infant bowel are not usually differentiated in microbiota investigations. These subspecies differ in their capacities to metabolize human milk oligosaccharides (HMO) and may have different ecological and symbiotic roles in humans. Quantitative PCR provides a quick analytical method by which to accurately ascertain the abundances of target species in microbiotas and microcosms. However, amplification targets in DNA extracted from samples need to be dependably differential. We evaluated the tuf gene sequence as a molecular target for quantitative PCR measurements of the abundances of B. longum subsp. infantis and B. longum subsp. longum in fecal microbiotas. This approach resulted in the detection of a tuf gene variant (operational taxonomic unit 49 [OTU49]) in Chinese infants that has sequence similarities to both B. longum subsp. infantis and B. longum subsp. longum We compared the genome sequence and growth and transcriptional characteristics of an OTU49 isolate cultured in HMO medium to those of other B. longum subsp. infantis cultures. We concluded from these studies that OTU49 belongs to B. longum subsp. infantis, that dependable quantitative PCR (qPCR) differentiation between the B. longum subspecies cannot be achieved by targeting tuf gene sequences, and that functional genes involved in carbohydrate metabolism might be better targets because they delineate ecological functions.IMPORTANCE High-throughput DNA sequencing methods and advanced bioinformatics analysis have revealed the composition and biochemical capacities of microbial communities (microbiota and microbiome), including those that inhabit the gut of human infants. However, the microbiology and function of natural ecosystems have received little attention in recent decades, so an appreciation of the dynamics of gut microbiota interactions is lacking. With respect to infants, rapid methodologies, such as quantitative PCR, are needed to determine the prevalences and proportions of different bifidobacterial species in observational and microcosm studies in order to obtain a better understanding of the dynamics of bifidobacterial nutrition and syntrophy, knowledge that might be used to manipulate the microbiota and perhaps ensure the better health of infants.

Dataset Information

Analysis, characterization, and loci of the tuf genes in lactobacillus and bifidobacterium species and their direct application for species identification.

Publications

Analysis, characterization, and loci of the tuf genes in lactobacillus and bifidobacterium species and their direct application for species identification.

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