| Lactobacillus reuteri possess great propertise and extensively applied in the development of functional product.The immunization regulated function of L.reuteri on the body has been extensively concerned among many probiotic propertise.Traditional microbial research focused on the characteristic evaluation of the strain by simple physiological and biochemical tests.However,researchers can explore the biological functions of microorganism based on the gene structure from perspective in bioinformatics with the maturity of whole genome sequencing technology which promoted the development of lactic acid bacteria(LAB)greatly.The study combined evaluation in vitro with genomics to invistigate the immunomodulatory activity and molecular biological information of L.reuteri J1 due to the strain specificity.The investigation revealed the probiotic mechanism and genetic characteristics of L.reuteri.In this study,L.reuteri J1 was used as the research object,which was isolated from conventional fermented dairy food.At the same time,evaluated the adhesive property,tolerance to the gastrointestinal,antimicrobial ability and immunomodulatory activity.Then,using complete genome sequencing and comparative genomics to explored in-depth.The conclusion will search the immunomodulatory activity and genetic basis of the strain.We obtained the following results through the study:(1)L.reuteri J1 has strong adhesive ability.The number in adhesion was 867±88.0 CFU/100Caco-2 Cells,and the adhesive rate was 87.93%.J1 has strong ability to colonize in the digestive tract compared with control in the experiment.(2)L.reuteri J1 has the potential to colonize in the gastrointestinal tract.Evaluating the tolerance to the gastrointestinal environment by simulating in the simulated artificial gastrointestinal fluid.In the evaluation of tolerance to gastrointestinal,the survival rates of the strain were 86.77% and 97.12% after treatment in the medium of p H 2.0 and p H 3.0 for 3 h,respectively.Treated with the bile salt in concentration of 0.3%,0.5% and 1.0%(v/v)for 4 h,the survival rates of the strain were 105.00%,82.60% and 82.18%,respectively.After being cultured with simulated artificial gastric juice for 3 h and then the cells were transferred to artificial intestinal juice for 8 h,the survival rate was 82.71%.After treated in 7 g/100 m L of saline solution for 24 h,the survival rate was 80.36%.The strain had good tolerance to gastrointestinal tract.(3)The strain has antibacterial ability.Using the cylinder plate method to evaluate the antibacterial ability.The fermentative supernatant of L.reuteri J1 could inhibit the growth of Listeria monocytogenes and Bacillus cereus effectively,but there is no antibacterial ability in the Staphylococcus aureus,Bacillus subtilis,Escherichia coli and Salmonella.The glycerol fermentative supernatant has no effect on the indicator bacteria,and it is judged that there is no ability to convert glycerol into reuterin for J1(4)L.reuteri J1 has immunomodulatory activity.To research the expression levels of genes related to immunization,the RNA were extracted from Caco-2 cells after stimulating of L.reuteri JI,and the expression of immune-related genes in Caco-2 cells were tested by real-time RT-PCR.The conclusion showed that J1 could stimulate the cells evidently.The expression of IL-1?,IL-1? and CCL20 was stimulated and the expression of IL-6 and IL-8 was inhibited.(5)L.reuteri J1 genome contains 1 circular chromosome and 5 circular plasmids.The length of the genome in L.reuteri J1 is 2045363 bp,56896 bp,18542 bp,18059 bp,10362 bp and 8492 bp,respectively,and GC content is 39.3%(chromosome).There are 2189 coding genes,18 genomic islands(GIs),and 88 nc RNA.There are 1928 protein-coding genes annotated into the COG database.The genes interrelated to the synthesis of capsular polysaccharide and lipoteichoic acid were found in the L.reuteri J1 genome,Verifying the immunomodulatory activity of the strain further.(6)Comparative genomic analysis revealed that the relationship of J1 belonged to the genus of L.reuteri.Compared with the other 6 strains of L.reuteri,the average nucleotide identity(ANI)was above 95%. |
PDF Full Text Request