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Mycoplasma hyopneumoniae culture medium and preparation method thereof

A technology of Mycoplasma hyopneumoniae and culture medium, which is applied in the field of Mycoplasma hyopneumoniae culture medium and its preparation, can solve the problems of difficult to exceed titer and high serum dosage, and achieve the effects of reducing production costs, reducing pollution risks, and reducing allergic stress reactions.

Active Publication Date: 2017-06-30
JIANGSU NANNONG HI TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Since the isolation and identification of Mycoplasma hyopneumoniae, there have been different formulations of culture media, from the earliest boiled pig lung tissue inactivated cell culture medium, pig lung embedding block cell-free medium, to the later Friis medium, Jiangsu Academy of Agricultural Sciences Mycoplasma hyopneumoniae can be cultured in the KM2 culture medium and the Mycoplasma hyopneumoniae medium in the veterinary biological product manufacturing and inspection regulations, but the culture titer is difficult to exceed 10 9 CCU, the amount of serum is also relatively high

Method used

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  • Mycoplasma hyopneumoniae culture medium and preparation method thereof
  • Mycoplasma hyopneumoniae culture medium and preparation method thereof

Examples

Experimental program
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Embodiment 1

[0043] ——Preparation of culture medium of the present invention

[0044] 1. Preparation of 20× Hank's solution:

[0045] 1) 20×Hank's1: Weigh 16g of NaCl, MgSO 4 ·7H 2 O 0.5g, KCl 1.0g, MgCl · ·6H 2 O 0.5g, each component was dissolved in 90ml of deionized water one by one, weighed CaCl 2 Dissolve 0.5g in 10ml deionized water and mix the two;

[0046] 2) 20×Hank's2: Weigh Na 2 HPO 4 12H 2 O 0.4g, KH 2 PO 4 0.35 g and 5 g of glucose were dissolved in 98 ml of deionized water one by one, and 2 ml of 1% phenol red solution was added.

[0047] 2. Preparation of basal medium:

[0048] Measure 20×Hank’s 1 35ml and 20×Hank’s 2 35ml in 722ml deionized water, weigh 2g of yeast extract, 0.5g of milk protein hydrolyzate, and 6g of beef heart extract, stir and dissolve one by one, adjust to 10M NaOH pH to 7.4, autoclave at 115°C for 20min.

[0049] 3. Preparation of auxiliary medium:

[0050] 1) 5% arginine solution: Weigh 5 g of L-arginine and dissolve it in 100 ml of dei...

Embodiment 2

[0056] ——Preparation of culture medium of the present invention

[0057] (1) Preparation of 20× Hank's solution:

[0058] 1) 20×Hank's1: Weigh NaCl 12g, MgSO 4 ·7H 2 O 0.1g, KCl 0.4g, MgCl 2 ·6H 2 O 0.1g, each component was dissolved in 90ml deionized water one by one, weighed CaCl 2 Dissolve 0.2g in 10ml deionized water and mix the two;

[0059] 2) 20×Hank's2: Weigh Na 2 HPO 4 12H 2 O 0.1g, KH 2 PO 4 0.10 g and 5 g of glucose were dissolved in 96 ml of deionized water one by one, and 4 ml of 1% phenol red solution was added.

[0060] (2) Preparation of basal medium:

[0061] Measure 20×Hank’s 1 25ml and 20×Hank’s 2 25ml in 825ml deionized water, weigh 5g of yeast extract, 3g of milk protein hydrolyzate, and 12g of beef heart extract, stir and dissolve one by one, adjust the pH with 10M NaOH To 7.2, autoclave at 115°C for 20 minutes.

[0062] (3) Preparation of auxiliary medium:

[0063] 1) 5% arginine solution: Weigh 5 g of L-arginine and dissolve it in 100 ml...

Embodiment 3

[0069] ——Preparation of culture medium of the present invention

[0070] (1) Preparation of 20× Hank's solution:

[0071] 1) 20×Hank's1: Weigh 15g of NaCl, MgSO 4 ·7H 2 O 0.2g, KCl 0.6g, MgCl 2 ·6H 2 O 0.3g, each component was dissolved in 90ml deionized water one by one, weighed CaCl 2 Dissolve 0.3g in 10ml deionized water, and mix the two;

[0072] 2) 20×Hank's2: Weigh Na 2 HPO 4 12H 2 O 0.2g, KH 2 PO 4 0.20 g and 2 g of glucose were dissolved in 97 ml of deionized water one by one, and 3 ml of 1% phenol red solution was added.

[0073] (2) Preparation of basal medium:

[0074] Measure 20×Hank’s 1 30ml and 20×Hank’s 2 30ml in 780ml deionized water, weigh 3g of yeast extract, 2g of milk protein hydrolyzate, and 8g of beef heart extract, stir and dissolve one by one, adjust the pH with 10M NaOH To 7.3, autoclave at 115°C for 20min.

[0075] (3) Preparation of auxiliary medium:

[0076] 1) 5% arginine solution: Weigh 5 g of L-arginine and dissolve it in 100 ml o...

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Abstract

The invention relates to a mycoplasma hyopneumoniae culture medium and a preparation method thereof. The mycoplasma hyopneumoniae culture medium consists of a basic culture medium and an auxiliary culture medium, wherein the basic culture medium has major components of Hank's liquid, lactoprotein hydrolysate, a yeast extract and an ox heart extract, sterilization can be performed through autoclaving, and a pollution risk caused by filtration sterilization is greatly reduced; and the auxiliary culture medium comprises pig serum, argenine, cysteine, phenol red solution, penicillium and the like, the pig serum is sterilized through cobalt radiation, and the rest components are mixed with inactivated pig serum after filtration sterilization. The titer of a lapinized attenuated strain of the mycoplasma hyopneumoniae cultured with the culture medium provided by the invention is 109-1010 CCU, the minimum culture time may be 40 h, the generation of old bacteria and aged bacteria is greatly reduced, the usage amount of the pig serum can be as low as 8%, the allergic stress reaction caused by the pig serum is reduced, and the production cost of an enterprise is lowered.

Description

technical field [0001] The invention belongs to the technical field of veterinary biology, and relates to a culture medium for mycoplasma hyopneumoniae and a preparation method thereof. [0002] technical background [0003] Mycoplasma swine pneumonia, also known as swine panting disease, is a chronic respiratory infectious disease caused by Mycoplasma hyopneumoniae. Clinically, it is characterized by cough, wheezing and typical "shrimp meat"-like lesions in the lungs, with high morbidity and low mortality. Mycoplasma hyopneumoniae can cause upper respiratory commensal bacteria such as Pasteurella multocida to proliferate in the lungs by inhibiting innate and acquired lung immunity and promote the occurrence of diseases. the occurrence of disease. Although M. hyopneumoniae is not the only pathogen that causes lung disease, it is known as an enhancer of pathogens that can cause lung disease and is therefore a major cause of economic loss. [0004] The effective prevention a...

Claims

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Application Information

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IPC IPC(8): C12N1/20C12R1/35
CPCC12N1/20
Inventor 车巧林董彦鹏缪芬芳耿晓眉靳蒙蒙胡静雅沈晓红
Owner JIANGSU NANNONG HI TECH
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