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KMID : 0545120110210121280
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Journal of Microbiology and Biotechnology
2011 Volume.21 No. 12 p.1280 ~ p.1286
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Development of a Quantitative PCR for Detection of Lactobacillus plantarum Starters During Wine Malolactic Fermentation
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Cho Gyu-Sung
Sabrina Krauss
Melanie Huch
Maret Du Toit
Charles M.A.P. Franz
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Abstract
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A quantitative, real-time PCR method was developed to enumerate Lactobacillus plantarum IWBT B 188 during the malolactic fermentation (MLF) in Grauburgunder wine. The qRT-PCR was strain-specific, as it was based on primers targeting a plasmid DNA sequence, or it was L. plantarum-specific, as it targeted a chromosomally located plantaricin gene sequence. Two 50 l wine fermentations were prepared. One was inoculated with 15 g/hl Saccharomyces cerevisiae, followed by L. plantarum IWBT B 188 at 3.6 × 106 CFU/ml, whereas the other was not inoculated (control). Viable cell counts were performed for up to 25 days on MRS agar, and the same cells were enumerated by qRT-PCR with both the plasmid or chromosomally encoded gene primers. The L. plantarum strain survived under the harsh conditions in the wine fermentation at levels above 105/ml for approx. 10 days, after which cell numbers decreased to levels of 103 CFU/ml at day 25, and to below the detection limit after day 25. In the control, no lactic acid bacteria could be detected throughout the fermentation, with the exception of two sampling points where ca. 1 × 102 CFU/ml was detected. The minimum detection level for quantitative PCR in this study was 1 × 102 to 1 × 103 CFU/ml. The qRT-PCR results determined generally overestimated the plate count results by about 1 log unit, probably as a result of the presence of DNA from dead cells. Overall, qRT-PCR appeared to be well suited for specifically enumerating Lactobacillus plantarum starter cultures in the MLF in wine.
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KEYWORD
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Lactobacillus plantarum, malolactic fermentation, LAB, wine, starter cultures, qRT-PCR
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