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Applied and Environmental Microbiology, August 2001, p. 3450-3454, Vol. 67, No. 8
0099-2240/01/$04.00+0   DOI: 10.1128/AEM.67.8.3450-3454.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Differentiation of Lactobacillus plantarum, L. pentosus, and L. paraplantarum by recA Gene Sequence Analysis and Multiplex PCR Assay with recA Gene-Derived Primers

Sandra Torriani,1,2 Giovanna E. Felis,1 and Franco Dellaglio1,*

Dipartimento Scientifico e Tecnologico, Facoltà di Scienze MM.FF.NN., Università degli Studi di Verona, 37134 Verona,1 and DOFATA, Università degli Studi di Catania, 95124 Catania,2 Italy

Received 12 February 2001/Accepted 27 April 2001

In this study, we succeeded in differentiating Lactobacillus plantarum, Lactobacillus pentosus, and Lactobacillus paraplantarum by means of recA gene sequence comparison. Short homologous regions of about 360 bp were amplified by PCR with degenerate consensus primers, sequenced, and analyzed, and 322 bp were considered for the inference of phylogenetic trees. Phylograms, obtained by parsimony, maximum likelihood, and analysis of data matrices with the neighbor-joining model, were coherent and clearly separated the three species. The validity of the recA gene and RecA protein as phylogenetic markers is discussed. Based on the same sequences, species-specific primers were designed, and a multiplex PCR protocol for the simultaneous distinction of these bacteria was optimized. The sizes of the amplicons were 318 bp for L. plantarum, 218 bp for L. pentosus, and 107 bp for L. paraplantarum. This strategy permitted the unambiguous identification of strains belonging to L. plantarum, L. pentosus, and L. paraplantarum in a single reaction, indicating its applicability to the speciation of isolates of the L. plantarum group.

* Corresponding author. Mailing address: Dipartimento Scientifico e Tecnologico, Facoltà di Scienze MM.FF.NN., Università degli Studi di Verona, Strada Le Grazie, 15, Ca' Vignal 2-37134 Verona, Italy. Phone: 39 045 8027917. Fax: 39 045 8027928. E-mail: dellaglio{at}sci.univr.it.

Applied and Environmental Microbiology, August 2001, p. 3450-3454, Vol. 67, No. 8
0099-2240/01/$04.00+0   DOI: 10.1128/AEM.67.8.3450-3454.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


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