TGEV and PEDV combined live vaccine and preparation method thereof

A technology of live vaccines and viruses, which is applied in the direction of medical formulas, medical preparations with non-active ingredients, medical preparations containing active ingredients, etc. It can solve the problems of poor protection against swine diarrhea, unsatisfactory effects, and economic losses.

Active Publication Date: 2015-07-22
兆丰华生物科技(南京)有限公司 +3
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Pigs over 5 weeks old have a low mortality rate, but due to the impact of diarrhea, slow growth and low feed remuneration, resulting in serious economic losses
[0005] Immunization is an important means to prevent and control the disease. There are reports on the use of vaccines at home and abroad, but the effect is not satisfactory
Especially in recent years, the protect

Method used

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  • TGEV and PEDV combined live vaccine and preparation method thereof
  • TGEV and PEDV combined live vaccine and preparation method thereof
  • TGEV and PEDV combined live vaccine and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0032] Example 1 Attenuated strain source

[0033] 1. Attenuated HB08 strain of porcine transmissible gastroenteritis virus

[0034] Take transmissible gastroenteritis antigen positive disease material from a pig farm in Hebei, add PBS according to the ratio of 1:5 (weight: volume), freeze and thaw repeatedly 3 times, centrifuge to get supernatant, filter with 0.22μm filter membrane, add to the filtrate Trypsin with a final concentration of 20 μg / ml was treated at 37° C. for 1.5 hours.

[0035] A monolayer of ST cells was inoculated according to conventional methods, and blank cells were set as controls. Significant and stable CPE changes appeared at the 17th generation ( figure 1 ). Design the detection primer pair according to the S gene: S-F: 5'-TATTTGTGGTYTTGGTYGTAATGC-3'; S-R: 5'-GGCTGTTTGGTAACTAATTTRCCA-3', perform 32 cycles at 95°C 45S, 50°C 45S, 72°C for 1min, then extend at 72°C for 5min . The amplified S gene fragment is 886bp, see the results figure 2 .

[0...

Embodiment 2

[0055] Embodiment 2 attenuated strain characteristic

[0056] 1. Attenuated HB08 strain of porcine transmissible gastroenteritis virus

[0057] The 125th and 165th passage viruses were diluted to 10 with DMEM respectively. 3.0 TCID 50 / ml, mixed with an equal amount of TGEV-specific serum, neutralized at 37°C for 1 hour, inoculated into 6 wells of a 96-well cell plate covered with a single layer of ST cells, and set 6 wells of non-neutralizing virus positive control cells and only Inoculate 6 wells of the negative control cells in the cell maintenance solution, culture in a 5% CO2 incubator at 37°C for 72-120 hours, and observe the cytopathic changes. Results The cells in the test group had no pathological changes, while the virus control group showed typical CPE changes.

[0058] The 145th generation seed virus of the attenuated porcine transmissible gastroenteritis virus HB08 strain was inoculated with 10 7.0 TCID 50 Inoculate 3 healthy piglets at the age of 3 at a dos...

Embodiment 3

[0067] Example 3 Preparation of dual vaccine

[0068] 1. Preparation of virus liquid for seedling production

[0069] Inoculate the well-growing monolayer cells ST and Vero E6 with the 155th generation of the attenuated porcine transmissible gastroenteritis virus strain HB08 and the 135th generation of the attenuated porcine epidemic diarrhea virus strain ZJ08, discard the cell growth medium, and follow the 2% After inoculation at 37°C for 1 hour, replenish the cell growth maintenance solution and continue culturing. When the CPE reached 80%, the virus was harvested, frozen and thawed 3 times at -20°C / room temperature, and the same batch of viruses were mixed together. Two batches of vaccine virus liquids were prepared by this method, and samples were taken for sterility test, mycoplasma test, exogenous virus test and determination of virus content at the same time. Choose virus content ≥ 10 6.0 TCID 50 / ml of virus liquid is used for seedling production.

[0070] 2 Mixin...

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Abstract

The invention discloses a combined live vaccine of transmissible gastroenteritis virus of swine (TGEV) and porcine epidemic diarrhea virus (PEDV) and a preparation method thereof. An attenuated swine transmissible gastroenteritis virus HB08 and an attenuated porcine epidemic diarrhea virus ZJ08 which are self-separated, attenuated and stored respectively undergo viral multiplication on ST cells and VeroE6 cells, and seedling and freeze drying are then carried out by adding a freeze-drying protective additive into a virus solution which is qualified after inspected. The two diseases, transmissible gastroenteritis of swine and porcine epidemic diarrhea virus, which are epidemic in clinic at present, can be effectively prevented by the use of the combined live vaccine.

Description

technical field [0001] The invention belongs to the field of veterinary biological products, and in particular relates to a porcine transmissible gastroenteritis virus and porcine epidemic diarrhea virus dual live vaccine and a preparation method thereof. Background technique [0002] Porcine transmissible gastroenteritis (TGE) is an important contact viral intestinal infectious disease that causes pig diarrhea. The mortality rate of pigs over 5 weeks old is low, but due to the impact of diarrhea, the growth is slow and the feed remuneration is low, causing serious economic losses. The pathogen was identified by Doyle et al. in 1946. Since the late 1960s, the disease has been prevalent in many provinces and cities in my country. [0003] Porcine epidemic diarrhea (porcine epidemic diarrhea, PED) is a highly acute, contagious viral intestinal infectious disease that causes diarrhea in pigs, characterized by watery diarrhea, vomiting, dehydration and decreased appetite. The...

Claims

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Application Information

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IPC IPC(8): A61K39/225A61K47/42A61K47/26A61P31/14A61K39/215
Inventor 王贵华侯艳红赵亚荣于萍萍刘明明满坤陈翠云卢会英
Owner 兆丰华生物科技(南京)有限公司
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