Development of Diagnostic Oligonucleotide Probes for Four Lactobacillus Species Occurring in the Intestinal Tract

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Summary

Partial sequences of the 16S rRNA from Lactobacillus acidophilus, Lactobacillus fermentum, Lactobacillus plantarum and Lactobacillus ruminis were used to design diagnostic species-specific DNA oligonucleotide probes. The specificity of the probes was tested against crude rRNA of 30 strains from 12 Lactobacillus species and 18 Gram-positive representatives from various genera. The detection limit of rRNA from homologous strains in mixed cultures (108 cells of a heterologous species) and in 0.6 g minced beef meat corresponded to 3 × 104 cells, and 1 × 105 cells respectively. A rapid lysis technique of cells was developed that facilitates the isolation of nucleic acids directly in 1.5 ml screw-top vials.

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      Citation Excerpt :

      Probes are synthetically prepared oligonucleotides meant to bind to the specific complimentary recognition sequence on the target bacterial genomic DNA. Most often, probes have been designed either against 16S or 23S rRNA genes (Ehrmann, Ludwig, & Schleifer, 1994; Hensiek, Krupp, & Stackebrandt, 1992; Hertel et al., 1991; Hertel, Ludwig, Pot, Kersters, & Schleifer, 1993; Sghir, Antonopoulos, & Mackie, 1998), but 16S rRNA based probes remain more popular due to the smaller size of 16S rRNA (1.5 knt) in comparison to the 2.3 knt of the 23S rRNA gene. Species-specific oligonucleotide probes have been used for the successful identification of a number of important Lactobacillus species (Alander et al., 1999; Ampe, 2000; Chagnaud, Machinis, Coutte, Marecat, & Mercenier, 2001; Drake, Small, Spence, & Swanson, 1996; Lucchini et al., 1998; Nakagawa et al., 1994; Park & Itoh, 2005; Pot et al., 1993; Tilsala-Timisjarvi & Alatossava, 1997; Yasui, Okamoto, & Taguchi, 1997).

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    Professor Dr. Erko Stackebrandt, Dept. of Microbiology, University of Queensland, St. Lucia, Qld. 4072, Australia

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