Enterococcus faecalis for treating colitis

A technology for Enterococcus faecalis and colitis, applied in the field of Enterococcus faecalis

Pending Publication Date: 2022-03-01
SUZHOU PRECISION BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, this strain is derived from chickens and is used to improve poultry enteritis caused

Method used

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  • Enterococcus faecalis for treating colitis
  • Enterococcus faecalis for treating colitis
  • Enterococcus faecalis for treating colitis

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0099] Example 1 Isolation and identification of Enterococcus faecalis

[0100] (1) Strain isolation and sequencing

[0101] Under sterile conditions, the prepared YK-20210225 donor fecal bacteria liquid was diluted 10 times, and three concentration gradients were selected, streaked and inoculated on NA and MRS medium, and three replicate plates were incubated at a constant temperature of 37°C. Anaerobic culture in the box for 72 hours, then pick different forms of colonies for Gram staining and microscopic examination, and carry out 2 times of streaking and purification in MRS medium, then staining and microscopic examination, observe the bacterial morphology and whether it is purified, and then carry out 16S For rRNA sequencing, universal primers were used, wherein the forward primer was 27F, whose sequence was shown in SEQ ID NO.2; the reverse primer was 1429R, whose sequence was shown in SEQ ID NO.3.

[0102] The PCR reaction system is as follows:

[0103] Prem...

Embodiment 2

[0137]Example 2 Detection of Resistance to Artificial Gastrointestinal Fluid and Detection of Bile Salt Resistance

[0138] (1) MTT solution preparation: Weigh 0.5g, add 100mL PBS to dissolve (final concentration is 5mg / mL), filter and sterilize, aliquot into 15mL centrifuge tubes, freeze at -20°C for later use, valid for half a year.

[0139] (2) Simulated Gastric Fluid (SGF) experiment

[0140] 1. Take 2.0g NaCl and 3.2g pepsin (Soleibao pepsin, 1:3000, marked as 800-2500 activity units per mg), add 7.0mL 37% dilute hydrochloric acid and water to dissolve to 1000mL , namely, the pH value of the solution is 1.2;

[0141] 2. Adjust the pH to pH 1.2 (simulating the pH of the fasting intestinal fluid of a human), pH 2.0, pH 3.0 (simulating the pH of the intestinal fluid of a human after meals and the pH of the fasting intestinal fluid of a mouse) and pH 4.0, and filter to sterilize;

[0142] 3. Bacteria collection: Bacterial strains were cultured at 37°C for 8 hours, and reach...

Embodiment 3

[0166] Embodiment 3 in vitro drug effect test

[0167] LPS mother liquor configuration: dissolve in DMEM, prepare 1 mg / mL mother liquor, and store in -20°C refrigerator.

[0168] Experimental operation: RAW264.7 cell culture (DMEM high glucose medium + 10% FBS, 37°C + 5% CO 2 For culture), the cells in a better culture state are blown off the adherent cells with the culture medium (the ones that cannot be blown off can be discarded), blown evenly, and the cells are counted, and diluted to obtain a certain concentration of cell culture medium (DMEM +10% FBS), seeded in 24-well cell culture plate, the number of cells per well is about 2×10 6 per well, the inoculum volume is 250 μL. After incubation for 16 hours, set blank control group, LPS (1 μg / mL), LPS (1 μg / mL) + 10% culture supernatant of PRS-217-05 bacteria (that is, the strains were cultured in MRS liquid medium for 24 hours, and then processed by high-speed centrifugation Obtain the bacterial culture supernatant, the ...

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Abstract

The invention provides enterococcus faecalis which is preserved in the China Center for Type Culture Collection on September 10, 2021, the preservation number of the enterococcus faecalis is CCTCC NO: M 20211156, and the preservation date of the enterococcus faecalis is September 10, 2021. The enterococcus faecalis CCTCC NO: M 20211156 provided by the invention has better tolerance to gastrointestinal fluid, and particularly has better tolerance to artificial intestinal fluid with pH of 6.8. The strain can normally grow in ox bile powder culture media with different concentrations of 1%-4%, has a certain tolerance degree to ox bile salt, and can normally grow and breed. The expression quantity of cell inflammatory factors can be reduced.

Description

technical field [0001] The invention belongs to the field of biomedicine, and in particular relates to an enterococcus faecalis for treating colitis. Background technique [0002] Inflammatory bowel disease (Inflammatory bowel disease, IBD) is a chronic and recurrent gastrointestinal inflammatory disease of unknown cause, and its incidence has gradually increased worldwide in recent years. The etiology and pathogenesis of IBD are not very clear at present, and most studies believe that the pathogenesis involves genetic factors, infection factors, and abnormal intestinal immune function. Since the pathogenesis of the disease is in the colon, rectum, ileum and other parts of the intestinal tract that are most exposed to bacteria, many scholars speculate that the bacteria in the intestinal tract may be involved in the pathogenesis of the disease. It has been reported that the intestinal flora of the patient is out of balance. In the intestinal flora, the number of bacteria suc...

Claims

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Application Information

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IPC IPC(8): C12N1/20C12N1/04A61K35/74A61P1/00A61P29/00A61P37/02A23K10/18A23L29/00A23L33/135C12R1/01
CPCC12N1/20C12N1/04A61K35/74A61P1/00A61P29/00A61P37/02A23K10/18A23L33/135A23L29/065A23V2002/00A23V2200/3204
Inventor 朱永亮刘丹穆晓静张叶静朱蒙蒙马梦楠马泽为黄琦
Owner SUZHOU PRECISION BIOTECH CO LTD
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