Culture-dependent and -independent analyses of bacterial compositions and its contributions to formation of γ-aminobutyric acid and poly-γ-glutamic acid in Cheonggukjang

Abstract

This study was performed to unveil bacterial compositions and their contributions to the formation of γ-aminobutyric acid (GABA) and poly-γ-glutamic acid (γ-PGA) in Cheonggukjang. To predict possible key factors contributing to the content of the bioactive compounds in Cheonggukjang, commercial products were analyzed for various parameters. The content of GABA and γ-PGA showed a negative (R² = 0.61 - 0.73) and positive correlation (R² = 0.53 - 0.96) with antioxidative activity. Consistently, GABA content showed a moderate negative correlation with γ-PGA content (R² = 0.58). Among the physicochemical and microbial parameters, only salinity showed a moderate negative correlation with γ-PGA content (R² = 0.75), which might be due to the inhibition of bacterial growth. It was also suggested that multiple factors (including bacterial species) were involved in the formation of GABA and γ-PGA in Cheonggukjang. To reveal dominant bacterial species and further presume their contributions to the bioactive compound formation in Cheonggukjang, both culture-independent (metagenomic) and -dependent (culturomic) methods were used. Culture-independent method showed that Bacillus piscis was dominant (23.37 - 94.89 %), followed by B. hisashii (0.00 - 62.45 %) and B. coagulans (0.00 - 13.82 %). Considering the quantitative speciation data on the bioactive compound content in Cheonggukjang (and bacterial production capability) together, it was further elucidated that B. piscis contributed primarily to the bioactive compound formation. Unlike this, culture-dependent analysis revealed that B. licheniformis and B. subtilis were dominant (30.0 - 47.6 and 17.5 - 39.5 %, respectively). Based on the quantitative speciation data on the bacterial production capability of GABA and γ-PGA, B. subtilis was the primarily contributing bacterial species to the bioactive compound formation. Consequently, it was observed that the bacterial compositions and their contributions to the bioactive compound formation determined by the two methods differed considerably, i.e., B. piscis and B. subtilis were identified to be prominent bacterial contributors, respectively, depending on the method used.

Keywords: Bacillus piscis; Bacillus subtilis; Bioactive compounds; Cheonggukjang; Culture-dependent method; Culture-independent method; Metagenomic analysis; Poly-γ-glutamic acid; Traditional microbiological analysis; γ-aminobutyric acid.