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Lactobacillus for inhibiting PEDV (Porcine Epidemic Diarrhea Virus) adhesion

A technology of Lactobacillus reuteri and Lactobacillus reuteri, applied in the directions of bacteria, microorganism-based methods, medical preparations containing active ingredients, etc., can solve the problems of difficult prevention and control work and unstable effects.

Active Publication Date: 2018-09-04
NANJING AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, due to the great changes in the epidemic characteristics and incidence of PED in recent years, the effect of the single-use vaccine is obviously unstable, which has brought many difficulties to the prevention and control work.

Method used

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  • Lactobacillus for inhibiting PEDV (Porcine Epidemic Diarrhea Virus) adhesion
  • Lactobacillus for inhibiting PEDV (Porcine Epidemic Diarrhea Virus) adhesion
  • Lactobacillus for inhibiting PEDV (Porcine Epidemic Diarrhea Virus) adhesion

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0032] Example 1: Isolation and Identification of Lactobacillus of Porcine Origin

[0033] 1. Sample collection

[0034] Animal feces were collected in the animal room of an experimental animal center, healthy piglets around 45 days old. Use a sterile cotton swab to collect fresh feces samples from pigs within 30-60 minutes, put them in sealed bags, put them in an ice box and send them back to the laboratory immediately. Randomly sample 5 samples from each pig house, and collect feces from 4 pig houses. There are 20 samples in total. All piglets were free of recent diarrheal disease, antibiotics or feed containing antibiotics, and feed additives containing probiotics. After taking the samples and sending them to the laboratory, store them at 4°C, and complete the isolation of bacteria within 4 hours.

[0035] 2. Medium

[0036] Lactobacillus selective medium (MRS-vancomycin solid medium): peptone 10g, beef extract 10g, yeast extract 5g, glucose 20g, sodium acetate 5g, diam...

Embodiment 2

[0064] Example 2: Lactobacillus reuteri NAU2015807129 acid resistance, bile salt resistance, adhesion ability test

[0065] 1. Acid resistance test

[0066] The test strain was inoculated in the MRS liquid medium according to the inoculum amount of 5% (v / v), and activated for 2 to 3 generations to ensure the high activity of the strain. The activated bacterial solution was inserted into the liquid medium of pH 1.5, pH 2.0 and pH 2.5 and the unacidified medium (as a control) acidified with hydrochloric acid according to 1% (v / v) inoculum, 37 After treatment at ℃ for 2 hours, the samples were diluted in different gradients, and the total number of colonies was measured by the agar plate pouring method, and the changes in the number of viable bacteria before and after treatment were compared. Each experiment was repeated three times.

[0067] Table 4 The growth of NAU2015807129 cultured in MRS with different pH for 2h (mean ± standard deviation)

[0068] Table 4 Survival analy...

Embodiment 3

[0087] Example 3: Lactobacillus reuteri NAU2015807129 inhibits PEDV adhesion test

[0088] In this experiment, vero cells were used to amplify PEDV, using 100TCID 50 The challenge meter infects IPEC-J2 cells.

[0089] IPEC-J2 cells were cultured on a 96-well plate. After the cells were fused to a monolayer, the medium was discarded, washed once with sterilized PBS, and then the following experiments were performed. The experiments were divided into three treatments as follows:

[0090] (1) Group 1 prevention group. Pre-concentrated at 1×10 8 Add 0.1mL of CFU / mL bacterial solution to the cells for 1.5h, wash off thoroughly, and then add TCID with a titer of 100 times 50 / 0.1mL of PEDV virus 0.1mL treated for 1.5h, after thorough washing, 37℃, 5%CO 2 The cells were cultured in an incubator, with 4 replicates for each sample.

[0091] (2) Group 2 repair group. Pre-titer to 100 times TCID 50 / 0.1mL PEDV virus 0.1mL was added to the cells for 1.5h, washed away thoroughly, an...

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Abstract

The invention discloses lactobacillus for inhibiting PEDV (Porcine Epidemic Diarrhea Virus) adhesion and belongs to the field of biological high techniques. The classification name of the strain NAU2015807129 is Lactobacillus reuteri, and is preserved in the China General Microbiological Culture Collection Center in March 27, 2017, and the strain preservation number is CGMCC No.13934. The strain is a feed probiotic which is approved by the Department of Agriculture, can be produced as a probiotic additive of pigs by using fermentation techniques, and can be applied to prevention and treatmenton porcine epizootic diarrhea. The strain NAU2015807129 for inhibiting PEDV adhesion, which is disclosed by the invention, can be prepared into a medicine or a food additive, can be used for preventing and treating porcine epizootic diarrhea, and has great application values.

Description

technical field [0001] The invention relates to a strain of Lactobacillus NAU2015807129 that inhibits PEDV adhesion, belongs to the field of biological high technology, and relates to the isolation, identification and screening methods of Lactobacillus with acid resistance, bile salt resistance, adhesion ability and inhibition of PEDV adhesion, and uses fermentation technology to The bacterium is expanded and cultivated, and the method of biological control is used to prevent and treat porcine epidemic diarrhea, and is suitable for preparing probiotic preparations with the function of preventing and treating porcine epidemic diarrhea. Background technique [0002] Porcine epidemic diarrhea (Porcine epidemic diarrhea, PED) is an acute, contact and highly contagious intestinal infectious disease of pigs caused by porcine epidemic diarrhea virus (Porcine epidemic diarrhea virus, PEDV). The disease occurs only in pigs, and there is no breed difference. The incidence rate of suc...

Claims

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Application Information

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IPC IPC(8): C12N1/20A61K35/747A61P31/14C12R1/225
CPCA61K2035/115A61P31/14C12N1/20C12N1/205C12R2001/225
Inventor 黄克和王添翔倪少闯陈兴祥甘芳
Owner NANJING AGRICULTURAL UNIVERSITY
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