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Applied and Environmental Microbiology, November 2003, p. 6908-6922, Vol. 69, No. 11
0099-2240/03/$08.00+0     DOI: 10.1128/AEM.69.11.6908-6922.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Analysis, Characterization, and Loci of the tuf Genes in Lactobacillus and Bifidobacterium Species and Their Direct Application for Species Identification

Marco Ventura,1* Carlos Canchaya,1 Valèrie Meylan,1 Todd R. Klaenhammer,2 and Ralf Zink1,{dagger}

Nestlé Research Center, 1000 Lausanne 26, Switzerland,1 Department of Food Science, College of Agriculture and Life Sciences, North Carolina State University, Raleigh, North Carolina 27695-76242

Received 16 June 2003/ Accepted 19 August 2003

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.

* Corresponding author. Mailing address: Department of Microbiology, National University of Ireland, Cork, Western Road, Cork, Ireland. Phone: 353 21 4901365. Fax: 353 21 4903101. E-mail: m.ventura{at}ucc.ie.

{dagger} Present address: Cognis, Dusseldorf, Germany.

Applied and Environmental Microbiology, November 2003, p. 6908-6922, Vol. 69, No. 11
0099-2240/03/$08.00+0     DOI: 10.1128/AEM.69.11.6908-6922.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



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