Development of Culture-Independent Detection Method for Beer Spoilage Lactic Acid Bacteria

Abstract

In microbiological quality control of final beer products, 100–400 ml beer samples are traditionally filtered on a membrane filter, followed by a lengthy culturing process in/on a laboratory medium that usually requires 3–14 days. This study aims to detect beer spoilage lactic acid bacteria (LAB) in a culture-independent fashion, using a direct polymerase chain reaction (PCR) approach. In order to detect a trace level of spoilage LAB in beer products, pressure cycling technology was evaluated to determine if this technology could improve the direct recovery of DNA from membrane filters for PCR detection. In this study, a mixed cellulose ester filter was adopted to allow a larger volume of beer to be filtrated. As a result of the optimization of the DNA extraction process that minimizes the loss of DNA and the effect of PCR inhibitors, 100 (1–10) cells/300 ml beer were successfully detected for beer spoilage LAB. Furthermore, a challenge test using 3,000 ml beer showed that 100 cells of Levilactobacillus (Lactobacillus) brevis were detectable without the negative effects of beer-originated PCR inhibitors, indicating that a trace level of contamination, corresponding to 1 cell in 300 ml of beer products, is detectable. It was also shown that the test was completed in approximately 8 hours. Therefore, this newly developed method is considered to be useful for the culture-independent direct detection of an extremely low level of beer spoilage LAB without a traditional culturing process.

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Key Words:

• Beer spoilage
• culture-independent detection
• lactic acid bacteria
• pressure cycling technology

Acknowledgments

We are grateful to Megumi Ito of Adecco Ltd. for providing technical assistance and useful discussion for this study.

Disclosure statement

No potential conflict of interest was reported by the authors.