| Koumiss is a complicated ecosystem which bears many strains of Lactic acid bacteria (LAB)with excellent industrial properties. It is extremely necessary to isolate and identify these strains tofurther abound our industrial microbe resources and to protect this traditional product as well as toavoid the loss of LAB resources. Several trials were conducted to isolate and identify LAB strainsfrom traditional home-made Koumiss collected from Inner Mongolia-Lanqi region. Thebiodiversity and biological characteristics of these LABs were studied. Using streptococcus mutansas the indicator microorganism, specific strain of these LABs was identified to inhibit the growth ofstreptococcus mutans. Then the inhibiting substance was separated and its physico-chemicalproperties were further studied. All these work together established the base for the exploration offunctional LAB usage.29 separated cultures were isolated from 4 different koumiss examples, the complete 16SrRNA gene of these strains were sequenced, and phylogenetic trees were constructed. The isolateswere identified to the most related strain which had the highest affinity. The identified strains weresix L. lactis, five L. plantarum, five E. faecium, four E. faecalis, two L. casei, two L. helveticus, twoS. thermophilus, one L. delbrueckii subsp, bulgaricus, one Leu. mesenteroides, one Leu. garlicum.Therefore, the result showed that the koumiss mainly bears Lactobacillus, Enterococcus andLactococcus, additionally, there were sparing genuses of Leuconostoc and Streptococcus et al. Onthe species level, more than 2/3 of the total LAB species were predominant strains composed of L.plantarum (17.2% of isolates), E. faecium (17.2%), E. faecalis (13.8%) and L. lactis (20.7%). Inaddition, there were also some species of L. helveticus (6.9%), L. casei (6.9%), S. thermophilus(6.9%), L. delbrueckii subsp, bulgaricus (3.4%), Leu. mesenteroides (3.4%) and Leu.garlicum(3.4%).One strain designated KLDS1.0432 (L. helveticus) of the isolated LABs was found to inhibitthe growth of the indicator streptococcus mutans. Its fermentation broth still had the anti-bacterialactivity after neutralized to pH 5.0, but lost this activity after treated by trypsin and pepsin, whichindicated that the anti-bacterial substance was protein or peptide.The grow curve corresponding to the production of bacteriocin of KLDS 1.0432 was monitored,The result showed that bacteriocin was produced when the strain entered the logarithmic growthphase (6h). The highest production of bacteriocin was reached when the strain entered its stationaryphase (26h), and the yield would not increase any more. To get considerable amount of bacteriocin,the fermentation time should be more than 26h.The bacteriocin was purified by using the 'Hiload 16/60 Superdex 75 prep-grade' chromatographic column and the (?)KTA purifier 100 system. The result showed that theanti-bacterial activity increased to 26 times of the unpurified fermentation broth. Finally, theTricine-SDS-PAGE technology was used to estimate the molecular size of the bacteriocin and itssize was found to be between 7823 Da and 14400Da.LAB biodiversity in Chinese traditional koumiss was primarily studied in this study. It wasestablished to use 16S rRNA technology as feasible routine to identify LAB and to study thebiodiversity of microorganisms in traditional dairy products. Bacteriocin-producing strain wasisolated which provided the possibility to develop industrial probiotic strain with good property andindependent intellectual property right.
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