Suzuki, K.1, Shimokawa, M.2, Asano, S.2, Yako, N.2 and Yamagishi, H.1, (1)Quality Control Center, Asahi Breweries, Ltd., Moriya-shi, Ibaraki, JAPAN, (2)Asahi Breweries, Ltd., Moriya-shi, Ibaraki, JAPAN
Technical Session 8: Microbiology
Wednesday, June 07, 2017
8:15–10:00 a.m.
Harbourview (1st Floor)
Lactobacillus pastorianus was first described by Pasteur in 1876 and isolated by Van Laer in 1892. Therefore, L. pastorianus was recognized as the first beer spoilage Lactobacillus discovered in brewing microbiology. Mysteriously, this honorable species vanished in the brewing history and became almost forgotten. For the progress of brewing microbiology, this historic species should be rediscovered and recharacterized. A century later, one strain (L. paracollinoides) was found from a spoiled beer. This strain, designated as L. paracollinoides JCM 11969T, initially exhibited no culturability on any culture media except for beer. The stepwise adaptation to deMan, Rogosa and Sharpe (MRS) broth, in which degassed beer as a subculture medium was progressively replaced by an increasing proportion of MRS broth in 10 point increments, finally allowed the isolation of L. paracollinoides JCM 11969T. Strikingly, our taxonomic studies indicated L. pastorianus (Van Laer 1892) is identical to L. paracollinoides JCM 11969T at the species level, on the basis of 16S rDNA sequence analysis and DNA-DNA hybridization study. This finding led us to further study of L. paracollinoides, focusing on its culturability on microbiological media. As a consequence, L. paracollinoides JCM 11969T in a hard-to-culture state was found to grow optimally at pH 4.7 but very poorly at pH5.3. No growth was observed at pH 5.6, which is often the approximate pH value adopted by conventional culture media for beer spoilage lactic acid bacteria. Its growth was also inhibited by most of the nutrient sources typically contained in culture media, including sodium acetate, yeast extract, peptone and manganese. These characteristics were also observed with another L. paracollinoides strain JCM 15729 in hard-to-culture state, suggesting these are common features of this species. Furthermore L. paracollinoides JCM 11969T showed a strong sensitivity even to agar, a solidifying agent widely used for culture media in primary isolation. All of these characteristics account for the unculturable nature of L. paracollinoides. Interestingly, the distinguishing pH-dependent growth characters and nutrient sensitivities were no longer observed for the L. paracollinoides strains obtained by the stepwise adaptation to MRS broth, suggesting the long-term habitation in brewing environments is responsible for the hard-to-culture state in L. paracollinoides. In addition, additional environmental surveys revealed the presence of 18 L. paracollinoides strains, all of which exhibited no growth on conventional culture media. Taken together, these findings indicate that the species L. paracollinoides is common and ubiquitous in brewing environments, but extremely difficult to culture upon primary isolation from their natural environments. The indication that L. paracollinoides and L. pastorianus (Van Laer 1892) are synonymous explains why this historic species has been hidden for so many decades despite its ubiquitous distribution in brewing environments. This presentation summarizes 27 years of our studies on their extraordinarily unique culturability and provides useful insights for brewing microbiologists in developing a comprehensive QC culture medium for beer spoilage bacteria.
Koji Suzuki joined Asahi Breweries, Ltd. in 1992 and functions as a manager in the Quality Control Center. He received a Ph.D. degree from Tokyo University in 2004 and two awards from the Brewing Society of Japan in 2007 and 2009 for his work concerning beer spoilage lactic acid bacteria. In 2011, he also received a technology award from the Japanese Society for Bioscience, Biotechnology, and Agrochemistry. At present, he lectures on fermentation food science at Meiji University and serves on an editorial board of the Journal of the Institute of Brewing. He has authored many original and review papers, as well as several book chapters, such as those published by Elsevier Science and Woodhead. He is currently a vice chair of the BCOJ Analysis Committee and also serves as a member of the Industry-Government-Academia Collaboration Committee in the Japanese Society for Food Science and Technology.
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