Isolation And Identification Of Predominant Oenococcus Oeni From Chinese Wine

In order to screen Oenococcus oeni with good performance for the MLF, The excellent Oenococcus oeni for Chinese wine were isolated and identification from Xinjiang wine, Heibei wine, Gansu wine after Alcohol fermentation with ATB isolation medium. The main results of the experiment as follows:1. Forty-nine strains of malalactic bacteria were isolated and purified from Xinjiang wine, Heibei wine, Gansu wine after Alcohol fermentation with ATB isolation medium. According Cultivating characteristics, tomorphological, malic acid decomposition, catalase experiment and physiological biochemical characteristics, they were identified to be O. oeni.2. The 49 isolates were tested for their capacity of resistance to the individual stress condition of pH, ethanol%(V/V) and SO₂ content.According to the resuits,the forteen selected strains'growth under the combination of external stress physical conditions of ethanol (V/V),pH and SO₂ (mg/L) were also tested,the results showed that, in selective strains, the resistance of XJ-2a is stronger than SD-2a, and XJ-2b, HB-1A, XJ-3a, HB-1B have higher applicability in wine producing.3. The analysis of species-specific PCR was used to make molecule identify. The result as follows: The forteen better resistant strains of single factor all have only one clear strips with 1025bp, the results showed that these forteen strains were O. oeni.4. The analysis of 16S rRNA PCR was used to make molecule identify. The results as follows: The results showed that the 16S rRNA gene sequence similarity of forteen strains and model strain were all above 99 %, comparing with the type strains of O. oeni from GeneBank database. It further proved that the forteen strains were O. oeni, and the homologous phylogenetic tree had been established based on 16S rRNA sequence of O. oeni. Besides, The results of 16S rRNA gene sequences similarity matrix of model strains of 7 Genera in streptococcaceae showed that the similarity of the gene sequences of forteen strains reaches 100%, and the relationship of Leuconostoc mesenteroides and O. oeni is the closest, the similarity reached 83.5%.5. The determination of O.oeni-XJ-2a's growth curve. The results showed that the Optical Density (OD₆₀₀) of O.oeni-XJ-2a's biggest bacterium levitation liquid is more than O.oeni-SD-2a's only 0.267, in other words, O.oeni-XJ-2a have better growth characteristics.In conclusion, according to tomorphological and physiological biochemical characteristics, which showed that these strains belong to O. oeni; Finally, the species-specific PCR and homology analysis of 16S rRNA sequences also support the results.