Bovine tuberculosis diagnosis marker and application thereof

A diagnostic marker and technology for tuberculosis, applied in disease diagnosis, biological testing, biomaterial analysis, etc., can solve difficulties in popularization and application, high requirements for testing environment, technology and personnel, and inability to distinguish between bacteria-expelling period and non-bacteria-excreting period Tuberculosis cattle and other problems, to achieve the effect of prevention and control, improve the efficiency of diagnosis

Active Publication Date: 2019-06-28
INST OF ANIMAL SCI OF CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Currently established skin test, IFN-γ release test and serological detection methods cannot distinguish between shed and non-sheltered tuberculosis cattle.
Nest PCR detection method can be used to detect Mycobacterium bovis pathogens in nasal secretions, alveolar lavage fluid, and milk. Studies have shown that 23% to 80% of tuberculosis-positive cattle can detect Mycobacterium bovis in nasal swabs pathogens, but this method has relatively high requirements on the detection environment, technology and personnel, and it is difficult to promote and apply at the grassroots level

Method used

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  • Bovine tuberculosis diagnosis marker and application thereof
  • Bovine tuberculosis diagnosis marker and application thereof
  • Bovine tuberculosis diagnosis marker and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0034] Detection and collection of clinical samples involved in the present invention in embodiment 1

[0035] 1. Tuberculin skin test:

[0036] The tuberculin skin test was performed according to the "Diagnostic Criteria for Bovine Tuberculosis" (GB / T 18645-2002). Shave the upper 1 / 3 of the cow's neck, and inject 0.1 mL of purified tuberculin bovis (PPD-B, 250 IU / head) intradermally. The skin thickness at the injection site was measured by the same operator with a vernier caliper before and 72 hours after the injection, and the difference in skin thickness was calculated. When the difference in skin thickness is greater than or equal to 4mm, the cow is positive for tuberculosis; when the difference in skin thickness is less than 2mm, it is judged as negative for tuberculosis; The skin test was performed 60 days after the first test. If the difference in skin thickness in the second test was greater than or equal to 2mm, it was determined to be positive for tuberculosis.

...

Embodiment 2

[0054] Example 2 Screening of Molecular Markers

[0055] 1, sample pretreatment: the tuberculosis bovine PCR positive (bTB) determined in embodiment 1 of random screening PCR-P ), tuberculosis bovine PCR negative (bTB PCR-N ) and 20 tuberculosis-negative cattle (NC) each. in bTB PCR-P As an example, every 10 plasma samples were mixed in equal volumes to form 2 biologically replicated plasma pooled samples (bTB PCR-P P1 is a mixture of plasma samples from cows 1-10, bTB PCR-P P2 is a mixture of plasma samples from cows No. 11-20). According to this method, prepare bTB PCR-N (bTB PCR-N P1 and bTB PCR-N P2) and pooled plasma samples of the NC groups (NC P1 and NC P2).

[0056] 2. Removal of high-abundance proteins: Since IgG and BSA in plasma account for more than 85% of the total proteins in the sample, which will affect the detection of low-abundance proteins, high-abundance protein removal kits (purchased from Bio-Rad) were used to remove Highly abundant protein in...

Embodiment 3

[0062] Example 3 Verification of molecular markers in the present invention

[0063] 1. PRM identification of differential proteins: refer to the method of Example 2 to process the sample, adopt the targeted analysis strategy of mass spectrometry PRM, and after passing the DDA detection, add the peptide of the identified target protein to the inclusion list of the mass spectrometry acquisition method, so that the mass spectrometry Data collection was carried out for these specific peptides, and relative quantitative analysis was carried out by extracting fragment ion information (see Table 4 for results). According to the identification results of iTRAQ and PRM (see Table 3), IL-8, agp and TF in tuberculosis bovine plasma The content of CRP is more than 1.3 times that of healthy bovine plasma, and the content of CRP in tuberculosis bovine plasma in the excretion period is more than two times that in non-excretion period tuberculosis bovine plasma, and the results of iTRAQ and P...

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Abstract

The invention provides a bovine tuberculosis diagnosis marker and application thereof. A marker IL-8, CRP for distinguishing negative from positive of bovine tuberculosis is obtained by using a non-targeting proteomics technology for screening and a targeting proteomics technology for verification. The provided bovine tuberculosis diagnosis marker can be used to determine positive / negative state of the bovine tuberculosis as well as whether a cattle with the positive bovine tuberculosis state is in an infective period. The diagnosis marker can be used to prepare a kit or reagent for detectingthe bovine tuberculosis, provides a new detection target point for diagnosis of the bovine tuberculosis, helps timely detection and weed-out of the cattle with tuberculosis, and provides guarantee forintegrated control for the bovine tuberculosis.

Description

technical field [0001] The invention relates to a bovine tuberculosis diagnostic marker and its application in preparing a bovine tuberculosis diagnostic kit. Background technique [0002] Bovine Tuberculosis (Bovine Tuberculosis) is mainly caused by infection of Mycobacterium bovis (M.bovis), a member of Mycobacterium tuberculosis complex (MTBC). Compared with Mycobacterium tuberculosis, Mycobacterium bovis has a wider host range, and it is most likely to infect cattle. It can also infect other livestock, primates, cats, dogs, and wild ruminants. Human-to-animal and human-to-human transmission. Tuberculosis cows excrete pathogenic bacteria in the form of aerosols, and aerosols containing Mycobacterium bovis adhere to pastures, water tanks, etc. Healthy animals can inhale 6 to 10 bacteria to cause infection. In addition, a milk cow with tuberculosis mastitis Bovis releases large amounts of Mycobacterium bovis, enough to contaminate the total milk produced by 100 healthy co...

Claims

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Application Information

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IPC IPC(8): G01N33/569G01N33/68
CPCG01N33/6869G01N2333/5421G01N2333/4737G01N2333/35G01N33/54306G01N2800/52G01N2333/705G01N2800/12
Inventor 鑫婷贾红高新桃郭晓宇姜一曈朱鸿飞侯绍华林伟东
Owner INST OF ANIMAL SCI OF CHINESE ACAD OF AGRI SCI
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