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Application for inhibiting activity of trichomonas gallinae in vitro by using bile acid

A technology of bile acid and trichomonas, which is applied in the field of bile acid inhibition of the activity of Trichomonas pigeon, which can solve the problems of body canceration and deformity, damage to the central nervous system, etc.

Active Publication Date: 2021-07-23
INST OF ANIMAL SCI OF CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, with the use of metronidazole and other drugs, there have been reports proving that Trichomonas has obvious drug resistance to several nitroimidazoles such as metronidazole, demetonidazole, ronidazole and carnidazole, and The use of antibiotics can easily induce cancer and deformity of the body, damage the central nervous system, etc.

Method used

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  • Application for inhibiting activity of trichomonas gallinae in vitro by using bile acid
  • Application for inhibiting activity of trichomonas gallinae in vitro by using bile acid
  • Application for inhibiting activity of trichomonas gallinae in vitro by using bile acid

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0027] Embodiment 1: the in vitro inhibitory concentration of bile acid to trichomonas pigeon

[0028] In a pigeon farm in Beijing, the breeding pigeons suffering from trichomoniasis were selected as the collection objects, and the samples of the oral and pharyngeal lesions and the surrounding mucosa of the sick pigeons were collected with cotton swabs soaked in sterile saline, and the cotton swabs were placed in the In a 1.5ml centrifuge tube containing sterile normal saline, centrifuge at 5000r min-1 for 5min, remove the cotton swab and supernatant, resuspend the bottom pellet with sterile PBS, and remove the cells or tissues in the pellet by Percoll density gradient centrifugation to obtain The purified Trichomonas strain liquid is inoculated into TF medium containing 4% fetal bovine serum and 1% penicillin-streptomycin double antibody, and cultured at a constant temperature of 36.5-38°C.

[0029] Identification of worms: collect the purified cultured worms, resuspend the w...

Embodiment 2

[0044] Embodiment 2: the effect of bile acid on Trichomonas pigeon nuclei

[0045] For observing the change of Trichomonas pigeon nuclei under different treatments, it is stained with DAPI fluorescent dye, and operating method is as follows:

[0046] (1) collect the worm bodies after different treatments, and wash 3 times with PBS;

[0047] (2) Add 200 μL DAPI working solution to the washed worms, mix evenly, and incubate at room temperature for 15 minutes;

[0048] (3) Wash away DAPI: after the incubation, wash the worms 3 times with 1mL 1×PBS, and discard the supernatant;

[0049] (4) Resuspend the worm bodies in 200 μL 1×PBS, and observe with a fluorescence microscope.

[0050] Results: DAPI staining observation was performed on Trichomonas treated with metronidazole and bile acid for 24 hours. Compared with the results of DAPI staining of normal Trichomonas, it can be clearly seen that the nuclei in the treated Trichomonas are obvious become larger and rounder, which sh...

Embodiment 3

[0051] Example 3: The Toxic Effect of Bile Acids on Pigeon Embryo Fibroblasts

[0052] Observe and measure metronidazole and bile acid to the toxic effect of pigeon embryo fibroblast, concrete operation is as follows:

[0053] (1) Collect amplified pigeon embryonic fibroblasts, use a counter to count, and adjust the cell density to 4×10 with DMEM complete medium 5 pcs / mL, 37℃, 5%CO 2 The incubator is placed for use;

[0054] (2) Under sterile conditions, add sterilized water around the circumference of the 96-well plate, add 100 μL of cell suspension to each well from the second row, and place at 37°C, 5% CO 2 Incubate in an incubator for 6 hours to make the cells adhere to the wall;

[0055] (3) prepare metronidazole and the bile acid DMED complete culture fluid of identical concentration in the worm body treatment;

[0056] (4) After incubation for 6 hours, the supernatant of the culture solution was discarded, and then 100 μL of metronidazole / bile acid DMEM complete cul...

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Abstract

The invention discloses a method for inhibiting activity of trichomonas gallinae in vitro by using bile acid. The application for inhibiting activity of trichomonas gallinae in vitro by using bile acid comprises the following steps: culturing purified trichomonas gallinae in vitro by using a TF complete culture medium or a DMEM complete culture medium containing 4% of fetal bovine serum (FBS) and 1% of three antibodies (penicillin, streptomycin and amphotericin), and preparing a trichomonas strain solution with a certain density; and culturing 600 microliters of trichomonas strain liquid and 400 microliters of TF complete culture medium of bile acid with the concentration of 0.001%-0.09% at the constant temperature of 36.5-38 DEG C, and observing the state and the death rate of trichomonas at different times. Results show that the bile acid can damage cell nucleuses of trichomonas and further has an inhibition effect on trichomonas, and 100% inhibition effect on trichomonas gallinae can be achieved under treatment of the bile acid with proper concentration. The application for inhibiting activity of trichomonas gallinae in vitro by using the bile acid has the following advantages that the inhibition effect of the bile acid on the trichomonas gallinae cultured in vitro is the same as that of metronidazole on the trichomonas gallinae cultured in vitro, and the purpose of replacing metronidazole can be achieved, so that metronidazole and dimetronidazole are further replaced in production, and antibiotic-free breeding and green development are realized.

Description

technical field [0001] The invention relates to the technical field of inhibition of the activity of trichomonas by drugs, in particular to the application of bile acids for the inhibition of the activity of trichomonas. Background technique [0002] Trichomonas pigeonis is a pear-shaped, oval or spindle-shaped unicellular parasite with a body length of 5 μm to 10 μm and a width of 4 μm to 8 μm. Structures such as columns. Trichomonad pigeonii proliferates through longitudinal binary fission, and its life history only has the trophozoite stage, which is not only the nutrient intake stage, but also its parasitic and transmission stage. The most suitable pH for trichomonas pigeons is 6.8-7.0, the temperature is 36.5-38 ℃, facultative anaerobic is the most suitable growth and reproduction environment. [0003] Pigeon trichomoniasis has widespread and high incidence, and there are reports about the epidemic infection of pigeon trichomoniasis at home and abroad. Domestic repor...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K31/575A61P33/02
CPCA61K31/575A61P33/02
Inventor 陈继兰麻慧边世雄倪爱心葛平壮李云雷孙研研袁经纬
Owner INST OF ANIMAL SCI OF CHINESE ACAD OF AGRI SCI
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