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Haemonchus contortus nano-material subunit vaccine and application thereof

A technology for Haemonchus contortus and subunit vaccines, applied in vaccines, applications, veterinary vaccines, etc., can solve problems such as difficult vaccine research and development, achieve strong immune protection ability, high application value, prolong or enhance specificity effect of immune response

Active Publication Date: 2020-08-21
NANJING AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In addition, the complex reaction mechanism between the parasite and the host also makes the development of vaccines difficult.

Method used

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  • Haemonchus contortus nano-material subunit vaccine and application thereof
  • Haemonchus contortus nano-material subunit vaccine and application thereof
  • Haemonchus contortus nano-material subunit vaccine and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0045] Embodiment 1. Preparation of Haemonchus contortus nanomaterial subunit vaccine

[0046] 1. Extraction of total RNA from Haemonchus contortus

[0047] Total RNA was extracted according to the instructions of Inviotrgen TRIzol, and the specific steps were as follows:

[0048] (1) Add 1mL TRIzol reagent to the centrifuge tube containing adult worms of Haemonchus contortus, transfer it to a homogenizer soaked in 0.1% DEPC, grind it fully on ice, transfer it to a new centrifuge tube and let it stand at room temperature 5min;

[0049] (2) Add 200 μL of chloroform to the centrifuge tube, shake the tube vigorously by hand for 15 seconds, and let it stand at room temperature for 3 minutes;

[0050] (3) Centrifuge at 12,000 rpm for 15 minutes at 4°C, and carefully transfer the upper aqueous phase to a new centrifuge tube;

[0051] (4) Add 500 μL of isopropanol to the centrifuge tube, mix it upside down, and let stand at room temperature for 10 minutes;

[0052] (5) centrifuge...

Embodiment 2

[0128] Example 2. Detection of immune protection of a nanomaterial subunit vaccine of Haemonchus contortus

[0129] 1. Experimental Design

[0130] Mice were randomly assigned to 3 groups of 8 and vaccinated only once on day 0. Mice were sacrificed on day 14. The three groups were PBS group, rHCA59 group and rHCA59-PLGA group. Multiple subcutaneous (SC) injections were performed using 1 ml of vaccine (content 20 μg rHCA59 protein). .

[0131] 2. Observation of immune protection effect

[0132] 2.1 Detection of antibodies in serum

[0133] (1) The mouse antiserum was collected on the 14th day and sacrificed. Antibody testing by ELISA. The levels of IgG1, IgM and IgG2a in serum samples were determined with a mouse ELISA kit according to the manufacturer's instructions (HengYuan, Shanghai, China).

[0134] (2) Briefly, the wells of a 96-well microtiter plate were coated with purified antibodies (anti-mouse IgG1, IgG2a or IgM), and mouse serum samples diluted in PBS were a...

Embodiment 3

[0148] Example 3. Detection of immune protection of a nanomaterial subunit vaccine of Haemonchus contortus

[0149] 1. Experimental Design

[0150] Fifteen goats (3-6 months old) were divided into 3 groups, with 5 goats in each group, which were the rHCA59-PLGA group (both immunized and challenged) with the nanomaterial subunit vaccine of Haemonchus contortus, and the positive control group (only The worms were challenged and not immunized), and the negative control group (the worms were not challenged and not immunized).

[0151] The experimental group was first immunized and boosted with 500 μg of Haemonchus contortus nanomaterial subunit vaccine dissolved in 1 mL of PBS on the 0th day and the 14th day, respectively. On day 28, goats in the experimental group and the positive control group were infected with 8000 third-stage larvae of Haemonchus contortus. Goats were slaughtered on day 63. After attacking the worms, closely observe the sheep's feeding situation, coat and me...

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Abstract

The invention discloses a Haemonchus contortus nano-material subunit vaccine and applications thereof. The Haemonchus contortus nano-material subunit vaccine is a subunit vaccine prepared by wrappinghaemonchus contortus recombinant protein HCA59 with nano-material PLGA, the recombinant HCA59 protein has an amino acid sequence as shown in SEQ ID NO. 1. The subunit vaccine can be used for preventing haemonchus contortus infection of sheep. The particle size of the nano subunit vaccine is 75 to 402nm; the Haemonchus contortus nano-material subunit vaccine has good biocompatibility and unique physicochemical properties, has the advantages of targeting property, slow release property, safety, high efficiency and the like, and can be decomposed and metabolized in an animal body; the vaccine hasan irritant antigen effect of stimulating dendritic cells, can promote the capability of converting mononuclear cells into the dendritic cells, has relatively strong immunoprotection capability, andcan remarkably reduce the ovum discharge rate and adult reduction rate of goats infected with haemonchus contortus clinically.

Description

technical field [0001] The invention relates to the technical field of biological veterinary medicine, and relates to a nano-subunit vaccine of Haemonchus contortus. Background technique [0002] Haemonchus contortus mainly parasitizes the abomasum of ruminants such as camels, cattle, sheep, and deer, and feeds on the blood of the host. The survey found that the annual cost of controlling Haemonchus contortus has reached several million dollars in some European countries. In the current market, chemical drugs are mainly used to prevent and control the infection of Haemonchus contortus, but due to people's excessive dependence on chemical drugs, the emergence of drug-resistant strains of Haemonchus contortus makes the effect of drugs no longer ideal. In addition, due to widespread concerns about drug residues and some biosafety factors, it is difficult for some anti-nematode drugs to be widely used in the prevention and control of Haemonchus contortus. [0003] Therefore, ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K39/00A61K47/34A61P33/12C12N15/70C12N15/12
CPCA61K39/002A61K47/34A61P33/12C12N15/70C07K14/4354A61K2039/552Y02A50/30
Inventor 李祥瑞徐立新宋小凯严若峰王强强
Owner NANJING AGRICULTURAL UNIVERSITY
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