Intestines

Example 3

16S rRNA sequencing of ileal biopsies showed that the mucosally-associated bacteria from pediatric CD patients had reduced alpha diversity (Faith's phylogenetic diversity) compared to non-IBD patients (FIG. 2A and FIG. 2B). In addition, by examining the beta diversity, we found that microbial communities were more dissimilar among CD patients than a separately recruited, slightly younger non-IBD patient cohort (FIG. 2C and FIG. 2D). In the pediatric population, healthy volunteers are not available. The non-IBD controls were culled from pediatric patients with abdominal pain, who had no evidence of intestinal inflammation by histology, and were primarily comprised of individuals with functional abdominal pain and irritable bowel syndrome. Analysis of the bacterial composition of CD and non-IBD patients revealed distinguishing taxa between non-IBD and CD patients (FIG. 9A, FIG. 9B and FIG. 9C). These results suggested that the non-IBD controls in this study, while slightly younger on average and exhibiting GI complaints without evidence of inflammation were nevertheless more homogeneous and maintained an overall diverse microbial community.

Example 20

Colitis in mouse was induced by adding 3% DSS (dextran sulfate sodium) in the drinking water for 12 consecutive days. Besides DSS placement, mice were daily treated with GLP-2 analogs (40 μg/kg/day) [GLP-2G is the GLP2 sequence with G2S mutation and is a known drug called teduglutide]. Cyclosporine A (20 mg/kg/day) was used for the positive control group and PBS for the negative control group. During the experiment period, body weight was measured every day.

Intestinal Weight Body Weight Measurement

Animals were sacrificed after 12 days of treatment. Small intestine was immediately excited and flushed with PBS. After PBS was gently squished out, intestinal weight was weighed using an analytical balance. Intestinal vs. body weight ratio was then calculated and analyzed.

Body weight and intestine weight versus body weight in DSS-induced colitis mice after daily administration of mTA68 for 12 days are shown in FIG. 23.

Example 5

The Lactobacillus ingested through the oral cavity passes through the stomach with the lower acidity and the intestines with high digestive enzymes and are exposed to low pH of gastric acid, pepsin, intestinal bile salts and digestive enzymes. Therefore, in order to utilize microorganisms as probiotics, gastric juice resistance is essential to survive in low pH and enzymes, and bile juice resistance is essential to survive in extreme intestinal environment. In accordance with the present disclosure, experiments were conducted to identify resistance to artificial gastric juice and bile juice of the above two strains with superior inhibitory effects against Gardnerella vaginalis and Candida albicans. The pH of the gastric juice in the body is maintained at about 3.0, and the food passes through the stomach for about 3 hours. In general, when maintaining viable cell count for 3 hours or more at pH 3, the cells has the high resistance to acidity. In order to identify the intestinal viability of Lactobacillus, survival experiments for artificial gastric juice and artificial bile juice were conducted with reference to Maragkoudakis' method. MG4272 and MG4288 strains were streaked on MRS plate medium and incubated at 37° C. for 24 hours, and the resulting colonies were inoculated in MRS liquid medium and incubated (37° C., 24 hours). Then, 2% passage was incubated for 24 hours in fresh MRS medium. The culture medium was then centrifuged (4,000×g, 4° C., 5 minutes) and washed twice with phosphate-buffer saline (PBS, pH 7.4). The washed cells were adjusted to OD₆₀₀ 1.0 (10⁸ to 10⁹ CFU/mL) and used for resistance experiments to the artificial gastric juice and artificial bile solution, respectively. As a control, 900 μL of pH 7 PBS was added to 100 μL of diluted Lactobacillus and the mixture was shaken and the number of viable cells was measured immediately. In order to identify the resistance to gastric juice, pepsin (Sigma-Aldrich, Saint Louise, USA) was dissolved in 3 g/L of pH 3 to pH 4 PBS to prepare an artificial gastric juice. 100 μL of lactobacillus diluent was added to 900 μL of artificial gastric juice, shaken, and cultured at 37° C. In 3 hours, the viable cell count was measured. To identify resistance to the artificial bile juice, pancreatin (Sigma-Aldrich, Saint Louise, USA) was dissolved in 1 g/L at pH 7 to pH 8 to prepare artificial bile juice. 100 μL of lactobacillus diluent was added to 900 μL of artificial bile juice, shaken and incubated at 37° C. In 4 hours, the viable cell count was measured. The measured results are shown in Table 1 in terms of log CFU/ml.

As shown in Table 1 both strains of MG4272 and MG4288 were identified to maintain the viable cell count of 10⁸ CFU/mL or more after 3 hours at pH 3, thereby identifying excellent acid resistance. In the artificial bile resistance test, both strains of MG4272 and MG4288 were identified to maintain the viable cell count of 10⁸ CFU/mL or more, thereby identifying excellent bile resistance.