Porcine circovirus culture method and application

A technology of porcine circovirus and a culture method, applied in the field of veterinary biological products, can solve the problems of vaccination failure, instability, weakened respiratory tract immunity, etc.

Active Publication Date: 2014-12-31
PU LIKE BIO ENG
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The Ross study at the University of Iowa found that after Mhp infection, the ability of lymphocytes to produce antibodies decreased, cellular immunity decreased, and the ability of alveolar macrophages to phagocytize and clear pathogens also decreased, and the activity of suppressive T cells increased, resulting in respiratory immunity. Weakened disease resistance, which makes it easier for other pathogens to invade, and can cause secondary infection of porcine reproductive and respiratory syndrome, porcine circular ring, etc., resulting in the failure of various vaccinations
[0006] The biological characteristics of PCV2 are quite special. Although it can exist in the body

Method used

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  • Porcine circovirus culture method and application
  • Porcine circovirus culture method and application
  • Porcine circovirus culture method and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0073] The cultivation of different components of embodiment 1 and the mensuration of content

[0074] 1.1 Cultivation of different components

[0075] 1.1.1 PCV2 culture alone

[0076] To cultivate PK15 cells, drain the cell culture medium from the monolayer PK15 cells that have grown to 80%-100%.

[0077] Inoculate the basic seed virus of PCV2 SH strain on PK15 cells at 0.01 MOI, absorb at 37°C for 30 minutes, add the cells prepared by MEM medium containing 4% (V / V) calf serum and 2mmol / l D-glucosamine hydrochloride to maintain cultured at 37°C for 5-7 days, freeze-thawed at -40°C for 2-3 times, and the virus was harvested as culture 1.

[0078] 1.1.2 Co-culture of PCV2 and Mhp

[0079] To cultivate PK15 cells, drain the cell culture medium from the monolayer PK15 cells that have grown to 80%-100%.

[0080] Inoculate PCV2 SH strain and Mycoplasma hyopneumoniae HN0613 strain basic seed virus / bacteria into PK15 cells at 0.01 MOI of PCV2SH strain and 1% (V / V) of Mycoplasma ...

Embodiment 2

[0094] Co-culture of embodiment 2 PCV2 and Mhp

[0095] 2.1 Co-culture of PCV2 and Mhp

[0096] Prepare the co-culture of PCV2 and Mhp according to Example 1.1.2, wherein the PCV2 strains are respectively SD strain and WH strain, that is, respectively cultivate the co-culture of PCV2 SD strain and Mycoplasma hyopneumoniae HN0613 strain, and PCV2 WH strain and pig The co-culture of Mycoplasma pneumoniae HN0613 strain was named as culture 5 and culture 6 in turn.

[0097] 2.2 Determination of PCV2 and Mhp content in culture 5 and culture 6

[0098] According to the method in Example 1.2, the contents of PCV2 and Mhp in culture 5 and culture 6 were measured respectively, and the measurement results are shown in Table 2.

[0099] Table 2 Determination results of different components after cultivation

[0100]

Embodiment 3

[0101] Example 3 Preparation of PCV2 Single Vaccine and PCV2-Mhp Vaccine Composition

[0102] 3.1 Inactivation and inspection

[0103] Culture 1, culture 2, culture 5 and culture 6 prepared in Example 1-2 were inactivated by directly adding 10% formaldehyde solution at 0.1% (V / V), inactivated at 37°C for 24 hours, each Stir every 4 hours. The inactivated culture 1 and culture 2 were sampled for sterility and inactivation test respectively, and the culture 1, culture 2, culture 5 and culture 6 that passed the inspection were respectively used as antigen solutions and stored at 4°C.

[0104] 3.2 Preparation of Mycoplasma hyopneumoniae single vaccine

[0105] Prepare according to the method of Mycoplasma hyopneumoniae described in Chinese patent CN103127497A (Zhang Xuke, Sun Jinzhong, Bai Chaoyong. Porcine circovirus type 2, Mycoplasma pneumoniae dual inactivated vaccine and preparation method thereof. Chinese patent CN103127497A), and obtain Mycoplasma hyopneumoniae concentrat...

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Abstract

The invention provides a porcine circovirus culture method and application and a mycoplasma hyopneumoniae culture method. The porcine circovirus culture method can simultaneously culture porcine circovirus and mycoplasma hyopneumoniae, simultaneously proliferate the titer of the porcine circovirus and the mycoplasma hyopneumoniae, and obtain high titer porcine circovirus. A vaccine combination prepared by adding mycoplasma hyopneumoniae antigens into the porcine circovirus which is prepared by the porcine circovirus culture method can be used for the prevention and/or treatment of diseases caused by the porcine circovirus and the mycoplasma hyopneumoniae.

Description

technical field [0001] The invention belongs to the technical field of veterinary biological products, and in particular relates to a method for cultivating porcine circovirus and its application, and a method for cultivating mycoplasma hyopneumoniae. Background technique [0002] In 1974, German scholar Tischer et al. detected a cell pollutant from the continuously passaged pig kidney cell line PK15 cells, which was a virus-like particle that could not cause cytopathic changes. In 1982, the scholar further research confirmed that the genome of the virus is composed of single-stranded circular DNA, thus naming it Porcine Circovirus (PCV). [0003] Porcine circovirus belongs to the genus Circovirus of the family Circoviridae and is the smallest animal virus ever discovered, with an average diameter of 17 nm. According to the difference of pathogenicity, antigenicity and nucleotide sequence of porcine circovirus, PCV is divided into two genotypes of non-pathogenic PCV1 and pa...

Claims

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Application Information

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IPC IPC(8): C12N7/00C12N1/20A61K39/12A61P31/20C12R1/35C12R1/93
Inventor 张许科孙进忠王延辉王莹田克恭
Owner PU LIKE BIO ENG
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