A specific oligonucleotide primer for the rapid detection of Lactobacillus lindneri by polymerase chain reaction

Abstract

A polymerase chain reaction (PCR) method was developed for the rapid detection of the beer-spoilage heterofermentative lactic acid bacterium Lactobacillus lindneri. Three strains, the Chinese brewery isolate DA1, the Japanese commercial beer isolate BG2, and the Japanese brewery isolate SE3, which were serologically classified as belonging to L. lindneri, were used in this study. After sequencing the 16S rDNA of the isolates DA1 and BG2 and the typical beer-spoilage heterofermentative Lactobacillus brevis L63, these sequences were compared with published data. A L. lindneri specific PCR primer, DA-40, was then constructed based on the V1 variable region of 16S rDNA. The specificity of PCR using the L. lindneri specific primer DA-40 and the universal primer 907r was examined using five L. lidneri strains: the three isolates described above and two strains from culture collection, DSM 20690 and DSM 20692. A variety of beer-spoilage lactic acid bacteria, including 71 Lactobacillus strains and 13 Pediococcus strains, were also included in this examination. No PCR product was obtained from any DNA with the exception of the five L. lindneri strains, indicating that the L. lindneri specific primer DA-40 was highly specific. The detection limit for L. lindneri in beer was 63 CFU/100 mL of beer.

Application of molecular identification tools for Lactobacillus, with a focus on discrimination between closely related species: A review

“…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;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). However, probes based on other genes such as pyrDFE have also been successfully applied to resolve the differences among some closely related species (Briengel, Curk, & Hubert, 1996).…”

Isolation of a Hop-Sensitive Variant ofLactobacillus lindneriand Identification of Genetic Markers for Beer Spoilage Ability of Lactic Acid Bacteria

“…Among the beer spoilers, several species of lactic acid bacteria (LAB) are reported to be responsible for approximately 70% of spoilage incidents caused by microorganisms (2, 3). For this reason, species-specific identification methods based on PCR have been widely evaluated for potential applications to microbiological quality control (6,19,41,42,44). Although species-specific PCR tests are rapid and reasonably accurate, there are two problems for applying this approach to the quality control of breweries.…”

Random amplified polymorphic DNA-PCR based cloning of markers to identify the beer-spoilage strains of Lactobacillus brevis, Pediococcus damnosus, Lactobacillus collinoides and Lactobacillus coryniformis

“…The fact that the cloned ORF1 and ORF3 encode a putative dolicol phosphate mannose synthetase and a teichoic acid glycosylation protein, respectively, suggested to us that these genes could be responsible for production of a cell wall sugar molecule. Yasui et al. (1997); Tsuchiya et al.…”