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Method and kit for detecting streptococcus agalactiae

A technology of Streptococcus agalactiae and a kit is applied in the field of detection methods and detection kits of Streptococcus agalactiae, can solve problems such as detection of Streptococcus agalactiae, etc., and achieves the effects of rapid detection, short time consumption, and prevention of fish diseases

Active Publication Date: 2011-11-16
TONGWEI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] No method for detection of S. agalactiae by amplification of the S. agalactiae cpsF gene has been reported

Method used

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  • Method and kit for detecting streptococcus agalactiae
  • Method and kit for detecting streptococcus agalactiae
  • Method and kit for detecting streptococcus agalactiae

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] The isolation and identification of embodiment 1 pathogenic bacteria

[0038] 1. Experimental animals

[0039] Diseased tilapia: tilapia disease materials collected from many diseased areas in Hainan;

[0040] Healthy tilapia: Chongzhou Tongwei Seed Breeding Farm.

[0041] 2. Method

[0042] 2.1 Disease data collection

[0043] Eight batches of disease materials collected from Fengpo Town, Wenjiao Town, Overseas Chinese Farm in Wenchang City, Hainan Province, and Xinglong Town in southwestern Hainan Province were used for etiological research. Disease materials were collected from affected fish ponds, and 10 to 30 live tilapias with typical symptoms were collected from each fish pond. The typical symptoms of the disease were swimming, rollover, eye protruding, brain congestion, and gallbladder enlargement. , Liver and kidney hemorrhage, part of the diseased fish collected was used for on-site necropsy and organ detection, part of the diseased material was frozen quick...

Embodiment 2

[0099] Example 2 Primer Design

[0100] 1. Experimental method

[0101] 1. PCR primer design and synthesis

[0102] According to the cpsF gene sequence and cfb gene sequence in the S. agalactiae genome in GeneBank, two pairs of primers were designed with oligo6.0 software, as shown in Table 4, cpsF-F and cpsF-R are primer pair 1, respectively for amplifying the cpsF gene Forward and reaction primers are sequences shown in SEQ ID NO: 1 and SEQ ID NO: 2; cfb-F and cfb-R are primer pairs 2, which are forward and reverse primers for amplifying the cfb gene, respectively. Sequences shown in SEQ ID NO:3 and SEQ ID NO:4. The above primers were synthesized by Shanghai Sangon Bioengineering Technology Service Co., Ltd.

[0103] Table 4PCR primer information

[0104]

[0105] 2. Template preparation

[0106] According to the requirements of aseptic operation, the standard strain of Streptococcus agalactiae was inoculated in BHI broth, cultured by shaking at 37°C (120r / min) for 2...

Embodiment 3

[0132] Embodiment 3 primer specificity test

[0133] 1. Test method

[0134] 1. PCR primers

[0135] Take primer pair 1 and primer pair 2 in Example 2.

[0136] 2. Template preparation:

[0137] According to the requirements of aseptic operation, the standard strain of Streptococcus agalactiae, Streptococcus agalactiae HN0401 strain and Streptococcus iniae were inoculated in BHI broth, shake cultured at 37°C (120r / min) for 24 hours, and 2mL of the bacterial liquid was taken to extract DNA; Staphylococcus aureus, Edwardsiella spp., Aeromonas hydrophila, Aeromonas salmonicida, Aeromonas caviae and Stenotrophomonas maltophilia were respectively inoculated in LB broth, shaken at 28°C (120r / min) After culturing for 24 hours, take 2 mL of bacterial liquid to extract DNA. The bacterial DNA extraction procedure was carried out according to the instructions of the kit (No. DV810A) of Takara Biochemical Reagent Co., Ltd.

[0138] 3. PCR amplification

[0139] The standard strain ...

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Abstract

The invention discloses a method and a kit for detecting pathogenic germs of fish, and relates to detection technology for streptococcus agalactiae. The invention provides a kit for detecting detecting pathogenic germs of fish. The kit comprises a primer pair 1 with sequences as represented by SEQ ID NO.1-2 for amplifying genes from streptococcus agalactiae and / or a primer pair 2 with sequences as represented by SEQ ID NO. 3-4 for amplifying genes from streptococcus agalactiae. The invention also provides a method for detecting detecting pathogenic germs of fish with the above-mentioned kit. The detection kit and method provided in the invention can accurately and effectively detect streptococcus agalactiae, have the characteristics of high sensitivity, strong specificity, short time and rapid detection, are applicable to rapid detection, prediction and prevention of fish diseases, and can increase economic benefits.

Description

technical field [0001] The invention relates to a detection technology for fish pathogenic bacteria, in particular to a detection method and a detection kit for Streptococcus agalactiae. Background technique [0002] Tilapia (Tilapia) is a small and medium-sized fish. It is a freshwater farmed fish that is mainly cultivated in the world's aquaculture industry, and is known as one of the main sources of animal protein in the future. However, with the high-density farming of Tilapia , the occurrence of bacterial diseases is becoming more and more serious. [0003] Streptococcus agalactiae is a group B Streptococcus (GBS), which has a wide range of hosts and can infect humans and a variety of animals. It is highly infectious to aquatic animals and can cause tilapia and rainbow trout. Fish sepsis and meningitis, such as sea carp and mullet, have a high morbidity and mortality rate. In recent years, outbreaks of tilapia disease caused by Streptococcus agalactiae have occurred f...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12Q1/04C12R1/46
Inventor 肖丹汪开毓阳涛刘天强康琦王均
Owner TONGWEI
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