Study On Identification Of Bifidobacterium In Prescription Milk Powder Of Probiotics By Genotyping Technique

Bifidobacterium is the most important culture in human or animal digestive tract that the mallerais scholars Tissier DE of French Pasteur institute found it from breastfeeding faeces separation in 1899. Along with the development of bifidobacteria, it can produce special substances and inhibit the invasion of pathogenic bacteria, which have effect of regulating the body’s immune ability. In addition, Bifidobacterium can maintain the micro ecological balance,co-treating polysaccharide in the body, so that the species gradually be applied to all kinds of infant food, medicine, and so on.In order to monitor the safety of dairy products, strains that are used in dairy products is strictly regulated, such as " the "list of strains can be used for babies and infants food"(health ministry announcement no. 25, 2011), the "GB 10765-2010 national food safety standards Infant formula". Those rules stipulate the amount and types of bacteria strainsin in the infant dairy products.In addition, the national standard "GB 2012-4789.34 national food safety standards food microbiology detection Bifidobacterium identification" rules the conventional detection method of Bifidobacterium in food. The conventional detection method that it aimed at the number of species or physiological and biochemical experiments of strains is not good and poorly in repeatability and identification of strain level, which can not meet the national standards’ regulatory requirements.In recent years, molecular classification technology, that is based on gene sequence classification, is a kind of new biological technology,containing PFGE, RFLP,PCR, multiple PCR, PCR- RFLP, 16 S rRNA sequencing technology, MLST technology, etc.16 S rRNA gene sequences is deemed to the "living fossil" of species evolution, is internationally recognized as a wide range of species identification method.16S-23 S rRNA genes that is section sequence located in the 16 S r RNA genes and 23 S r RNA gene. Compared with the high conservative relative to the 16 S rRNA genes, the16S- 23 S rRNA gene have more variation band,so that it can be used to distinguish species that genes is very close. MLST classification technology is a new type of molecular classification method, that is formed through Maiden scholars improved the multiple locus enzyme electrophoresis. The technology based on seven housekeeping genes, so that it is not only have certain relatively conservative, but also has certain mutant, and can show genetic relationship of strains accuratly.In this paper, 16 S rRNA gene sequences, 16S-23 S rRNA gene sequence, MLSTbased classification technology and system evolution analysis are uesd to identify 12 standard strains of Bifidobacterium and 30 strains isolated from 20 probiotics powder samples.The main results were as follows:(1), The proportion of probiotics infant powder brand in domestic product is36.4%, through the research and the brand’s official website, among 75% of probiotics infants cow powdered, 25% of probiotics baby sheep powder. In survey, the powder label that is agree with government standard, including 56%of percent of pass in domestic brand, and the percent of pass in import brands less than 50%.(2), In the testing of the number of living Bacterium in samples,we found that 16 of 20 samples reaches the national standard, and the percent of pass is 90%. The number of information in the label and the number of living bacterium according with national standard is 13 and the percent is 65%.(3), 16 S r RNA gene sequences technology can identify 12 strains standard and30 sample strains. In the evolutionary tree, bootstrap values of most branches are more than 80 and has good resolution. On accout of high convervation of 16 Sr RNA gene sequences, this technology can only identifident to species level.The analysis of the samples can be identifidented accurately.(4), Similarity to 16 S rRNA gene sequences classification technology, 16S-23 S r RNA classification technology can accurately differentiate Bifidobacterium to species level. The degree of differentiation is above 90. The identification of sample strains has high resolution compared to 16 S rRNA classification technique.(5), MLST classification technology, can accurately distinguish between bifidobacterium five bifidobacterium species. The degree of differentiation is 100. In addition, the technology differentiate B.animalis. ssp. lactis ATCC27673 and other B.animalis. ssp. lactis, appraisal to intraspecific level. The technology owes high distinguishability.