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Biomarker for detecting highly pathogenic viral hemorrhagic septicemia virus and diagnostic method

A viral hemorrhage, biomarker technology, applied in the field of diagnosis of VHSV infection

Active Publication Date: 2018-04-17
国立水产物品质管理院
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] However, as a result of an analysis of the total gene sequence information of low-pathogenic VHSV and highly-pathogenic VHSV obtained in Korea, there was no difference in the NV gene sequences reported to be involved in the pathogenicity of VHSV

Method used

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  • Biomarker for detecting highly pathogenic viral hemorrhagic septicemia virus and diagnostic method
  • Biomarker for detecting highly pathogenic viral hemorrhagic septicemia virus and diagnostic method
  • Biomarker for detecting highly pathogenic viral hemorrhagic septicemia virus and diagnostic method

Examples

Experimental program
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Effect test

Embodiment 1

[0042] Confirmation of the mutation in the whole gene of viral hemorrhagic sepsis virus (VHSV)

[0043] In order to diagnose highly pathogenic VHSVs that cause severe mortality in various farmed fish species (including flounder) at an early stage, the whole gene mutation sequences of 13 types of VHSVs that induce fish mortality were obtained in coastal areas of Korea, and then from 2012- It was sequenced in 3 years in 2014.

[0044] 1. Separate VHSV

[0045] In order to obtain VHSV from the intestines of flounder exhibiting VHS symptoms, intestinal tissues obtained from 13 flounder were crushed in MEM medium containing antibiotics and fetal bovine serum, and then crushed at 13,000 rpm at 4°C. The product was centrifuged for 10 minutes and then filtered through a 0.45 μm membrane.

[0046] The pulverized product obtained after filtration was inoculated with EPC flounder cells at a dilution ratio of 1:100-1:1000, and the cells were cultured at 16°C. Then, by microscopic exam...

Embodiment 2

[0064] Confirmation of pathogenicity of VHSV amino acid mutant

[0065] 1. Confirmation of the effect of wild-type VHSV NV protein mutants in soleus cells

[0066] First, in soleus cells, the degree of production of ATP, which is considered as cellular energy, was analyzed to confirm the high pathogenicity of VHSV.

[0067] In order to efficiently express the wild-type VHSV NV protein in cells, the wild-type VHSV NV gene was cloned into a pcDNA3 vector to prepare a recombinant vector.

[0068] The prepared recombinant vector was used to transduce HINAE cells (embryonic cells of flounder) (National Institute of Fisheries Science, National Institute of Fisheries Science) to express VHSV NV protein. 24 hours after transduction, the production of ATP (which is considered as cellular energy) was measured by using the ATP Bioluminescence Assay Kit (Roch Company, Switzerland) according to the manufacturer's instructions. As a result, changes in cellular energy production upon expr...

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Abstract

The present invention is a technique relating to a biomarker composition for detecting viral hemorrhagic septicemia virus (VHSV), comprising as an effective ingredient a polypeptide of 5 to 120 consecutive amino acids containing at least one amino acid substitution selected from the group consisting of leucine (L) for serine (S) at position 56; asparagine (N) for serine (S) at position 8; alanine(A) for threonine (T) at position 81; alanine (A) for valine (V) at position 88; aspartic acid (D) for glycine (G) at position 117; and lysine (K) for glutamic acid (E) at position 119 in the amino acid sequence of VHSV consisting of the amino acid sequence of SEQ ID NO: 1. In the present invention, the six amino acid mutations can be used in a biomarker to detect new VHSV and in a biomarker, capable of early determination of VHSV infection in fishes, for diagnosis of viral hemorrhagic septicemia.

Description

technical field [0001] The invention relates to a biomarker composition for detecting highly pathogenic viral hemorrhagic septicemia virus (viral hemorrhagic septicemia virus, VHSV) and a method for diagnosing VHSV infection. Background technique [0002] Viral hemorrhagic septicemia virus (VHSV) is a viral pathogen that causes severe losses in many farmed fish species, including flounder, and is believed to be the cause of viral hemorrhagic septicemia (VHS). [0003] VHS was first reported as a critical viral disease of rainbow trout. However, VHS has recently been reported to occur in a variety of fish species, not only salmonids (e.g. trout, rainbow trout, coho salmon, brook trout, brown trout) trout (brown trout or steelhead trout), also includes fish in natural water systems (such as cod or herring) and farmed marine fish (such as flounder); Happened in Asia. [0004] In Korea, VHS has continued to be reported in the Gyeongbuk region and Jeju region since it was firs...

Claims

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Application Information

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IPC IPC(8): C12Q1/70C12Q1/6869G01N33/569
CPCC12Q1/701G01N33/56983G01N2333/08C12N2760/20011C12Q1/6869G01N2333/145G01N21/00
Inventor 郑载勋赵炫国成美沼赵眉英朴明爱池宝荣黄晟敦黄智妍
Owner 国立水产物品质管理院
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