High-throughput Sequencing Analysis Of Microbial Community Succession In Naturally Fermented Bean Dregs And Development Of Complex Bacterial Liquid Fermented Bean Dregs

Okara is a by-product of the production of soy products and is often used as feed and compost,but the overall utilization rate is low.At present,some places also use it as raw materials to produce okara products,such as okara cake,moldy bean dregs,bean dregs paste,etc.Most okara products mainly rely on natural fermentation during the production process,and the raw materials are susceptible to contamination.In addition,the product production process is not standardized,resulting in the unstable flora of the okara itself.Therefore,fermented bean dregs face the disadvantages of uneven quality in the production process.In this paper,the composition and changes of bacteria in traditional okara were explored by combining high-throughput sequencing with traditional strain isolation,and the selection of pure fermented cultures was carried out on this basis.After determining the pre-selected strains,the combination of sensory and physical and chemical indicators was used as the basis for the preparation of composite bacterial liquid,and then single-factor experiments and response surface optimization experiments were carried out to determine the best fermentation process,and the microorganisms in the products obtained by the two fermentation methods were analyzed,and the physical and chemical indicators before and after bean dregs were compared to illustrate the fermentation effect.The main methods and research results of the paper are summarized as follows:(1)Isolation and identification of traditional strains in okara and pre-selection of fermentation strains.High-throughput gene sequencing technology was used to sample and analyze a total of 5 samples in 3 stages of traditional bean dregs fermentation.At the bacterial phylum level,proteomycetes,firmicutes,actinomycetes and cyanobacteria are the mainstay,and the presence of proteomycetes and firmicutes runs through the whole fermentation process.The categories at the bacterial species level were approximately the same in the 5 samples.Among them,Lactobacillus,Acinetobacter jowlii,Delfortella,Red String of Erythros,and Lactobacillus swiss are the main bacterial species of the entire fermentation system.At the fungal phylum level,the ascomycetes,mucormycetes,basidiomycetes and zygomycetes were the main phylums.The phylum Mucormyces exists throughout the fermentation process,while the ascomycetes phylum gradually disappears with fermentation.The categories and quantities of fungal species in the five samples varied.Mucorrhage,Curly Mucorrhage,Multibranched Mucorus,Aspergillus broom,Epiphylococcus nigra and Mucorus totalis are the main fungal species of the entire fermentation system.By traditional strain isolation method,10 strains were identified,4 strains were Bacillus species,1 strain was Panmycetes,3 strains were Aspergillus strains,1 strain was yeast genus,and 1 strain was Leuconostomyces.This coincides with high-throughput sequencing results and demonstrates the accuracy of the assay.(2)Fermentation experiment and response surface optimization of complex bacterial liquid.Aspergillus oryzae,Yeast gum,Bacillus subtilis,and Lactobacoccus lactis were selected as fermentation strains,and after the strain combination and ratio experiments,it was concluded that when Aspergillus oryzae: Bacillus subtilis:Lactococcus lactis = 1:1:2,the highest score was scored according to the scoring system,and a compound fermentation liquid was made accordingly.From the single-factor experiment,when the moisture content is 70%,the fermentation time is6 d,the bacterial inoculation amount is 6%,and the fermentation temperature is 25℃,the compound fermentation effect of bean dregs is the best,and the response surface optimization experiment is carried out on this basis,when the process conditions are:moisture content 70.85%,fermentation time 5.91 d,bacterial inoculation amount6.17%,the theoretical score value is the highest,9.49 points.Combined with the actual operation,it was modified to 71% water content,6.0% inoculation amount and fermentation time of 6 days.Under this process,three parallel experiments were carried out,and the actual sensory score was 9.54 ± 0.12 points.(3)Detection of compound fermented okara results.Sequencing of the finished product of compound fermentation showed that after compound fermentation,the OTU types of bacteria increased compared with traditional fermentation,and the total number decreased.Compared with traditional fermentation,the number of bacterial species contained in the finished product has changed significantly,such as the number of enterococcus,Acinetobacter and Leucontomycetes,while the number of Bacillus has increased significantly.Compared with traditional fermentation,the total number of OTU species of fungi decreased,and the number of fungal species contained in the finished product changed significantly.For example,the genus Mucormyces decreased from 98.63% to 4.50%,and Aspergillus increased from0.07% to 86.21%.Moreover,the limit of pathogenic bacteria in the finished bean dregs meets the national standards,which ensures the safety and edibility of the finished bean dregs.The physical and chemical indexes before and after fermentation of bean dregs were detected,and it was found that in addition to the decrease in protein and insoluble dietary fiber content,the content of amino acid nitrogen,reducing sugar,total acid and soluble dietary fiber increased.The structural observation of fermented bean dregs by scanning electron microscopy showed that the particle structure of the bean dregs was broken and dispersed into small pieces after fermentation,and the function changed significantly.