Novel multivalent carrier vaccine for shrimp and application thereof

A carrier vaccine and prawn technology, applied in the field of genetic bioengineering, can solve the problems of unsatisfactory, inconvenient administration, and difficulty in the protection period of vaccine immunity beyond 28 days, etc., and achieve the effect of long protection period and long multivalent cross protection

Active Publication Date: 2013-01-30
马悦 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] However, there are still many shortcomings in the existing research work: 1) The administration is inconvenient, mainly by injection, and it is difficult to carry out productive breeding applications; 2) The immune protection period of the existing vaccines is difficult to exceed 28 days , cannot meet the actual needs of the immune protection period (more than 40 days) in shrimp production and breeding, so there is no production application value and development prospect
Before the present invention was made, such a vector vaccine had not been seen in any similar reports or invention patents so far.

Method used

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  • Novel multivalent carrier vaccine for shrimp and application thereof
  • Novel multivalent carrier vaccine for shrimp and application thereof
  • Novel multivalent carrier vaccine for shrimp and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0027] Example 1: Preparation of Multivalent Vector Vaccine Against Shrimp White Spot Syndrome and Vibriosis

[0028] 1. Culture medium preparation

[0029] (1) DSM liquid medium: bacterial nutrient broth (Difco) 8g, 10% (w / v) KCl 10ml, 1.2% (w / v) MgSO 4 ·7H 2 O 10ml, 1M NaOH 1.5ml, adjust pH to 7.6, ddH 2 O was adjusted to 1000ml. After autoclaving, cool to 50°C and add sterilized 1M Ca(NO 3 ) 2 , 0.01MMnCl 2 , 1mM FeSO 4 1ml each.

[0030] (2) PBS buffer: Na 2 HPO 4 2H 2 O 2.74g / L, NaH 2 PO 4 ·H 2 O 0.63g / L, the ingredients were dissolved in 1000ml of distilled water, and sterilized at 121°C for 20 minutes.

[0031] (3) LB medium: tryptone (Tryptone) 10g / L, yeast extract (Yeast extract) 5g / L, sodium chloride (NaCl) 10g / L, adjust the pH to 7.0.

[0032] 2. Recovery of the plate medium: take the seeds of the Bacillus subtilis recombinant strain with the preservation number CCTCCNo: M 2012395 stored at -80°C and inoculate them on the LB solid plate by streaking...

Embodiment 2

[0038] Example 2: Evaluation of immune protection against white spot syndrome by oral immunization of Penaeus vannamei

[0039] Breeding of Penaeus vannamei: The purchased experimental shrimp (Penaeus vannamei) was temporarily raised in a 3000L aquaculture tank after being tested for no WSSV carrying, with filtered fresh seawater (seawater salinity was 25g / L), and the aquaculture water temperature was maintained at 26 ±2°C, after 1 week of holding, remove unhealthy individuals. After continuing to breed for one week (average body weight 10±1.2g / tail), they were randomly divided into groups with 30 tails in each group, and each group paralleled 3 water tanks (100L / tank).

[0040] Preparation of WSSV virus suspension: Dissect the gills and pancreas of diseased shrimp, weigh 1 g, cut them into pieces and grind them under liquid nitrogen, add TN buffer (50 mmol / L Tris- HCl, pH 7.6, 0.4 mol / L NaCl), use a glass homogenizer to homogenize in an ice bath, centrifuge the homogenate...

Embodiment 3

[0047] Example 3: Evaluation of Immune Protection of Chinese Shrimp against White Spot Syndrome by Oral Immunization

[0048] Feeding of Penaeus chinensis: The experimental shrimp (Penaeus chinensis) purchased from Tianjin was temporarily raised in a 3000L aquaculture tank after being tested for no WSSV. ±2°C, after 1 week of holding, remove unhealthy individuals. After continuing to breed for one week (average body weight 10g±0.9g / tail), they were randomly divided into 100L water tanks, with 30 tails in each group, and 3 parallel water tanks in each group (100L / tank).

[0049] Preparation of WSSV virus suspension: Dissect the gills and pancreas of diseased shrimp, weigh 1 g, cut them into pieces and grind them under liquid nitrogen, add TN buffer (50 mmol / L Tris- HCl, pH 7.6, 0.4 mol / L NaCl), use a glass homogenizer to homogenize in an ice bath, centrifuge the homogenate at 4°C, 7000 g for 20 min, discard the precipitate; filter the supernatant through a 0.45 μm microporo...

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Abstract

The invention relates to a novel multivalent carrier vaccine for a shrimp. The vaccine is characterized by comprising a bacillus subtilis recombination strain obtained by a genetic engineering technique, the strain belongs to the category of bacillus subtilis HT5303 and is preserved in China Center for Type Culture Collection, and the preservation number is CCTCC NO.: M 2012395. When spores are formed by the bacillus subtilis vaccine strain, white spot syndrome virus VP281 protein can be demonstrated on the surfaces of the spores, and fusion protein of cell-penetrating peptides, white spot syndrome virus VP19 and vibrio harveyi 3-glyceraldehyde-phosphate dehydrogenase (GAPDH) can be secreted and expressed when vegetative cells are formed. Compared with other vaccines, the multivalent carrier vaccine has the advantages of cross protection and long protection period. Moreover, the vaccine can be added into feeds and has the advantages of resistance against high-temperature setting of feeds and broad prospects for commercial development.

Description

technical field [0001] The invention belongs to the technical field of genetic bioengineering, and relates to a novel polyvalent vector vaccine for prawn white spot syndrome virus (WSSV) and vibriosis and its application, in particular to a polyvalent vaccine for preventing and treating prawn white spot syndrome and vibriosis. Valence carrier vaccines and their applications. Background technique [0002] Shrimp is a high-value aquatic product mainly produced in Asia and Latin America for export purposes. At present, the main cultured species in my country are Penaeus vannamei, Penaeus monodon and Penaeus sinensis. However, in the past 10 years, there have been quite serious problems in shrimp farming, among which white spot syndrome virus (White spot syndrome virus, WSSV), taura virus (Taura Syndrome Virus, TSV) and Vibrio are the main culprits of the disease . Shrimp white spot syndrome (WSS) is an fulminant infectious disease caused by shrimp white spot syndrome virus (W...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K48/00A61K39/385A61K39/295A61P31/20A61P31/04C12N1/21C12N15/34C12R1/125
Inventor 不公告发明人
Owner 马悦
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