Study On Diversity And Biofilm Formation Property Of Microbial On Kefir Biofilm

Kefir is a kind of natural fermentation agent. The particle formation mechanism has been one of the hotspots in the field of scientific research, from the perspective of biological membrane of analysis of the particles formation mechanism is a new research direction. However there is little report about biofilm of Kefir currently, this paper focus on the formation, microbial diversity and growth of Kefir biofilm, in order to provide ideas for research the mechanism of Kefir grains formation and establish method of forming artificial Kefir.This paper compared the macro and micro structure, growth rate, fermentation characteristics, yield of Kefir polysaccharide and the formation of biofilm about two kinds of Kefir which originate from Qinghai and Northeast. Compared with the Kefir from Northeast, the Qinghai Kefir macroscopic and microscopic structure is denser, with the passage times increased, the growth of Qinghai Kefir is significantly higher than that of the Northeast folk Kefir(P<0.05), during the process of fermentation, the change of Oxidation-Reduction Potential in Qinghai Kefir grain was advantageous to aerobic bacteria,facultative anaerobe and anaerobic bacteria, the p H was significantly higher than that of the Northeast folk Kefir(P<0.05). After fermentation, the yield of polysaccharide in Qinghai Kefir is significantly higher than that of Northeast. Kefir biofilm formation at the best time for 3 days determined through the biofilm formation of Specific Biofilm Formation and viable count method, there can be seen obviously three-dimensional membrane structure, therefore, the Qinghai Kefir was chosen as the research object for the next experiment. The formation of Kefir biofilm was observed by laser confocal microscopy, polysaccharide and protein content were observed in the biofilm forming process, the protein content changed little, polysaccharide content increased significantly, and the biological membrane on total number of bacteria was increased with the incubation time added. The number of bacteria was biggest when 3 days.The microbial diversity of Kefir biofilm is detected by high throughput method, and compared with the natural Qinghai Kefir grain. Bacterial phylum level analysis shows that Kefir biofilm bacteria was mainly have 2 kinds and raw grains was mainly 5. Fungal phylum level analysis shows that Kefir biofilm bacteria was mainly have 1 kinds and raw grains are mainly 4. Bacterial genus level analysis showed that 6 genera of bacteria were got in biofilm, 12 genera of bacteria were got in natural grain. 2 genera of fungi were got in biofilm, 7 genera of fungi were found in natural grain. Besides the above identified species, Kefir biofilm and the natural grain are also contains uncultured and unclassified species. 6 species were isolated and identified in Kefir biofilm via traditional scribe methods: Kluyveromyces marxianus、Lactococcus lactis、Leuconostoc mesenteroides 、 Lactobacillus kefiri 、 Lactobacillus sunkii 、 Acetobacter orientalis. Studies of the hydrophobicity, zeta potential, auto-aggregation, co-aggregation and biofilm formation on the 6 kinds of microbial showed that, when the hydrophobicity was more bigger, the zeta was more lower, the Specific Biofilm Formation was more bigger for Lactic acid bacterial, but for yeast and acetobacter the change was not obvious. Compared with cultured alone and lactic acid bacteria co-cultured with yeast than acetic acid bacteria; the co-aggregation and biofilm formation ability were strongest when 6 species were co-cultured, but remain weaker than the Kefir biofilm, the results suggests that the strains were not isolated in Kefir biofilm also played an important role on the formation of Kefir biofilm.The effect of added exogenous substanceswas on Kefir biofilm growth was studied. The results showed that the best condition of promoting the growth of Kefir biofilm under concentration was 1.61% Ca Cl2, 2.26% casein, 5.55% lactose. Compared to the control group the Specific Biofilm Formation was 2.55±0.06, and improved by 66.24%, the 97.70% accuracy of model was verified and the corresponding number of viable cells was(11.6±0.19)×108CFU/m L and improved by 27.47%. The growth of Kefir biofilm can promote through optimized the culture conditions, thus accelerate the formation of the grain structure.