Isolation and identification of microorganisms in pit mud and primary analysis of caproic acid bacteria metabolites

The strains were isolated from strong-flavor (Nongxiangxing) Baijiu pit mud by a variety of medium enrichment methods and anaerobic isolation techniques, their basic taxonomic information was determined by morphological observation, 16S rRNA gene sequencing, and phylogenetic analysis, and the metabolites of caproic acid bacteria of Oscillospiraceae were analyzed by GC-MS and HPLC. The results showed that a total of 63 strains of bacteria were isolated. Based on the 16S rRNA gene sequences, all strains could be identified to genus and belonged to 2 class, 3 order, 8 families and 15 genus, including Clostridium, Lactiplantibacillus, Staphylococcus, Sporolactobacillus, Haloimpatiens, Paraclostridium, Rummeliibacillus, Caproiciproducens, etc. Among them, there were 9 strains showed low sequence similarity (96.17%-97.70%) to the proximate bacteria, which might be potential new species. Both of 3 caproic acid bacteria of Oscillospiraceae was capable of producing acetic acid, butyric acid, caproic acid by fermentation using RCM medium, and the caproic acid production of Caproicibacterium sp. 20-5D, Caproiciproducens sp. CPB-2 and Caproiciproducens sp. YR-4 was 3.84 g/L, 1.03 g/L and 0.35 g/L, respectively, with strong ability of producing mixed acids. The functional bacteria isolated from the pit mud of strong-flavor Baijiu had the great significance for enriching the numbers of culturable microbes in pit mud and the application and development of functional microorganisms.