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Aims: Tempeh is a soy-based traditional food fermented by Rhizopus oligosporus. Although this mold is the main microorganism responsible for tempeh fermentation, various unknown bacteria presence in tempeh could enhance tempeh’s nutritional value. This study is aimed to examine the identity of bacteria in tempeh bacterial community by combining metagenomics analysis and culturable technique. Methodology and results: Samples were obtained from a tempeh producer which consists of raw soybeans, fresh water used to soak the beans, soaking water after the beans were soaked for 18 h, dehulled-soybean before inoculation, starter culture, and fresh tempeh. All samples were plated onto Enterobacteriaceae and Lactic Acid Bacteria agar media, and the total DNA was extracted for metagenomics analysis based on 16S rRNA gene cloning and High-Throughput Sequencing (HTS). Metagenomic analysis indicated that Firmicutes and Proteobacteria were the predominant and subdominant bacteria, respectively, while the culturable technique showed Proteobacteria were the predominant bacteria. Firmicutes species detected in tempeh were similar to the ones in the soaking water, which were populated by Lactobacillus. However, another predominant bacteria from tempeh, Enterococcus, was similar to minor population of Enterococcus detected in dehulled-soybean before inoculation. Based on the cloned 16S rRNA genes, we observed L. agilis, L. fermentum, and E. cecorum as the predominant bacteria in tempeh. The starter culture, which was dominated by Clostridium, did not alter bacterial community in tempeh, since its proportion was only 2.7% in tempeh clean reads. Conclusion, significance and impact of study: The dominant bacteria in tempeh was Lactobacillus from Firmicutes. The bacterial community in tempeh was not affected by the starter culture used, but mainly because of the soybean soaking process.

Metagenome analysis of tempeh production: Where did the bacterial community in tempeh come from?