Aspergillus oryzae is an industrially useful species, of which various strains have been identified; however, their genetic relationships remain unclear. A. oryzae was previously thought to be asexual and unable to undergo crossbreeding. However, recent studies revealed the sexual reproduction of Aspergillus flavus, a species closely related to A. oryzae. To investigate potential sexual reproduction in A. oryzae and evolutionary history among A. oryzae and A. flavus strains, we assembled 82 draft genomes of A. oryzae strains used practically. The phylogenetic tree of concatenated genes confirmed that A. oryzae was monophyletic and nested in one of the clades of A. flavus but formed several clades with different genomic structures. Our results suggest that A. oryzae strains have undergone multiple inter-genomic recombination events between A. oryzae ancestors, although sexual recombination among domesticated species did not appear to have occurred during the domestication process, at least in the past few decades. Through inter- and intra-cladal comparative analysis, we found that evolutionary pressure induced by the domestication of A. oryzae appears to selectively cause non-synonymous and gap mutations in genes involved in fermentation characteristics, as well as intra-genomic rearrangements, with the conservation of industrially useful catalytic enzyme-encoding genes.

The microbial community during fermented vegetable production has a large impact on the quality of the final products. Lactic acid bacteria have been well-studied in such processes, but knowledge about the roles of non-lactic acid bacteria is limited. This study aimed to provide useful knowledge about the relationships between the microbiota, including non-lactic acid bacteria, and metabolites in commercial pickle production by investigating Japanese pickles fermented in rice-bran. The samples were provided by six manufacturers, divided into two groups depending on the production conditions. The microbiological content of these samples was investigated by high-throughput sequencing, and metabolites were assessed by liquid chromatography-mass spectrometry and enzymatic assay. The data suggest that Halomonas, halophilic Gram-negative bacteria, can increase glutamic acid content during the pickling process under selective conditions for bacterial growth. In contrast, in less selective conditions, the microbiota consumed glutamic acid. Our results indicate that the glutamic acid content in fermented pickle is influenced by the microbiota, rather than by externally added glutamic acid. Our data suggest that both lactic acid bacteria and non-lactic acid bacteria are positive key factors in the mechanism of commercial vegetable fermentation and affect the quality of pickles.

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Background Salting is a traditional procedure for producing pickled vegetables. Salting can be used as a pretreatment, for safe lactic acid fermentation and for salt stock preparation. This study aimed to provide valuable knowledge to improve pickle production by investigating the dynamics of microbiota and metabolites during the pretreatment and salt stock preparation processes, which have previously been overlooked. The differences in these process conditions would be expected to change the microbiota and consequently influence the content of metabolites in pickles. Methods Samples, collected from eight commercial pickle manufacturers in Japan, consisted of the initial raw materials, pickled vegetables and used brine. The microbiota were analyzed by 16S rRNA sequencing and the metabolites quantified by liquid chromatograph-mass spectrometry. Statistical analyses helped to identify any significant differences between samples from the initial raw materials, pretreatment process and salt stock preparation process groups. Results Under pretreatment conditions, aerobic and facultative anaerobic bacteria were predominant, including Vibrio, a potentially undesirable genus for pickle production. Under salt stock preparation conditions, the presence of halophilic bacteria, Halanaerobium, suggested their involvement in the increase in pyruvate derivatives such as branched-chain amino acids (BCAA). PICRUSt analysis indicated that the enhanced production of BCAA in salt stock was caused not by quantitative but by qualitative differences in the biosynthetic pathway of BCAA in the microbiota. Conclusion The differences in the microbiota between pretreatment and previously studied lactic acid fermentation processes emphasized the importance of anaerobic conditions and low pH under moderate salinity conditions for assuring safe pickle production. The results from the salt stock preparation process suggested that the Halanaerobium present may provide a key enzyme in the BCAA biosynthetic pathway which prefers NADH as a coenzyme. This feature can enhance BCAA production under anaerobic conditions where NADH is in excess. The effects shown in this study will be important for adjusting pickling conditions by changing the abundance of bacteria to improve the quality of pickled vegetables.