Live bacterial vaccine for controlling intestinal infection caused by clostridium difficile, preparation method thereof and application thereof

A technology for Clostridium difficile and intestinal infection, which is applied to a live bacteria vaccine for preventing and treating intestinal infection caused by Clostridium difficile and its preparation and application fields, and achieves the effects of being convenient for storage, transportation and taking.

Inactive Publication Date: 2012-10-24
THE FIRST AFFILIATED HOSPITAL OF GUANGZHOU MEDICAL UNIV (GUANGZHOU RESPIRATORY CENT) +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there is no study of live bacterial vaccines using Streptococcus lactis as an expression vector for intestinal infection caused by Clostridium difficile toxin A positive

Method used

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  • Live bacterial vaccine for controlling intestinal infection caused by clostridium difficile, preparation method thereof and application thereof
  • Live bacterial vaccine for controlling intestinal infection caused by clostridium difficile, preparation method thereof and application thereof
  • Live bacterial vaccine for controlling intestinal infection caused by clostridium difficile, preparation method thereof and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0056] 1. Vectors for expressing antigens in vaccines (expression bacteria)

[0057] Lactococcus lactis (Streptococcus lactis; Lactococcus lactis) is used as the carrier to express the 14CDTA protein molecule (antigen) of Clostridium difficile. A kind of bacteria that is harmless to human body and relatively safe (generally regarded as safe, GRAS) is also a kind of food industry bacteria, which is widely used in the dairy industry, such as the lactic acid streptococcus (CICC) preserved in China Industrial Microorganism Culture Collection Center No.: 6023), etc.

[0058] Culture Streptococcus lactis in M17 medium (Difco, Detroit, Mich) or similar medium in an anaerobic or aerobic environment at 30°C. For ThyA gene-deficient Streptococcus lactis, thymidine should be added to the medium (thymidine), the concentration of which is about 20 μM, M17 medium is a medium produced by Difco, and it can be used only after adding distilled water under high pressure.

[0059] 2. Using the ...

Embodiment 2

[0108] Live Recombinant Lactobacillus Vaccine Colonizes Intestines of Golden Hamsters

[0109] In order to observe the recombinant Streptococcus nisin oral vaccine expressing 14CDTA, two recombinants that secrete and express 14CDTA with TETC as a biological adjuvant and membrane-anchored 14CDTA with TETC as a biological adjuvant constructed by the method in Example 1 Streptococcus lactis,) on the prevention and treatment of Clostridium difficile infection, the shuttle plasmid pTRKH2 was used as a carrier to express 14CDTA fusion protein in Streptococcus lactis, and the promoter 59 of Lactococcus lactis subsp.cremoris was used in the experiment (promoter 59, p59) (GeneBank accession No.M24806,) was used as the promoter, and the RBS and signal peptide (75-181, GeneBank accessionNo. M60178) sequence was used as RBS and signal peptide; the terminator of Usp45 gene was selected as the terminator (1514-1553, GeneBank accession No.M60178); 14CDTA expression unit was inserted after th...

Embodiment 3

[0118] Effect of Recombinant Lactobacillus Vaccine on Prevention and Treatment of Clostridium difficile Enteritis

[0119] Lactobacillus LL-PNBCL1003 secreting and expressing 14CDTA with TETC as a biological adjuvant, membrane-anchored Streptococcus lactis LL-PNBCL2003 expressing 14CDTA with TETC as a biological adjuvant, and non-recombinant Lactobacillus (containing a non-recombinant plasmid The construction process of Streptococcus lactis LL-pTRKH2 of pTRKH2 is the same as that in Example 2.

[0120]1. Experimental grouping and immunization methods Take 32 male Syrian golden hamsters (hereinafter referred to as golden hamsters) with a body weight of 100-130 grams, and randomly divide them into 4 experimental groups with 8 animals in each group. The first group (8 animals) is given Normal saline gavage was used as a blank control group, and the second group (8 rats) was given non-recombinant plasmid Streptococcus lactis (LL-pTRKH2) as a positive control ( figure 2 A), the t...

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Abstract

The invention discloses a live bacterial vaccine for controlling intestinal infection caused by positive clostridium difficile of clostridium difficile toxin A, wherein the vaccine contains thymidylate synthetase ThyA gene defective recombined lactic streptococci; and tests of the vaccine verify that the vaccine is safe and effective. The invention also discloses a preparation method of the live bacterial vaccine, and a use of the live bacterial vaccine in the preparation of medicines for controlling the intestinal infection caused by the positive clostridium difficile of the clostridium difficile toxin A.

Description

technical field [0001] The invention relates to a live bacterial vaccine for preventing and treating intestinal infection caused by Clostridium difficile and its preparation method and application, in particular to a live bacterial vaccine for preventing and treating intestinal infection caused by Clostridium difficile toxin A-positive Clostridium difficile . Background technique [0002] Clostridium difficile (CD) is a Gram-positive anaerobic bacillus and is one of the most important pathogens of hospital infection. Studies have shown that clinically about 10-20% of antibiotic-associated diarrhea and nearly 100% of pseudomembranous colitis are related to this bacterium. Clostridium difficile-associated diarrhea (Clostridium difficile-associated diarrhea, CDAD) mainly occurs in people who take antibiotics for a long time, and can also occur in other situations that affect the normal intestinal flora and reduce the resistance to Clostridium difficile colonization, such as th...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K39/07A61P31/04C12N15/74C12N1/21C12R1/145C12R1/46
Inventor 陈学清杨晓强
Owner THE FIRST AFFILIATED HOSPITAL OF GUANGZHOU MEDICAL UNIV (GUANGZHOU RESPIRATORY CENT)
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