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Malachite green and recessive malachite green degrading bacteria and their separation and purification methods and applications

A technology of recessive malachite green and malachite green, which is applied in the direction of chemical instruments and methods, biochemical equipment and methods, and methods based on microorganisms, can solve the problem of unfavorable quality of aquaculture ecosystems and aquatic products, malachite green and hidden malachite green. There are no public reports on the microbial degradation bacteria of malachite green, and it has achieved the effect of protecting the fishery ecological environment, facilitating wide application, and good repeatability

Active Publication Date: 2021-08-06
上海市水产研究所(上海市水产技术推广站)
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] In view of the above-mentioned defects of the prior art, the technical problem to be solved by the present invention is that existing malachite green medicines and recessive malachite green residues are unfavorable to the aquaculture ecosystem and the quality of aquatic products in the fishery aquaculture environment. There are no public reports on microbial degradation bacteria of medium malachite green and recessive malachite green

Method used

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  • Malachite green and recessive malachite green degrading bacteria and their separation and purification methods and applications
  • Malachite green and recessive malachite green degrading bacteria and their separation and purification methods and applications
  • Malachite green and recessive malachite green degrading bacteria and their separation and purification methods and applications

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0045] Embodiment 1, the screening of malachite green and recessive malachite green degrading bacterial strain

[0046] Preparation of media and reagents

[0047] Basic medium: potassium dihydrogen phosphate 3.0g, magnesium sulfate heptahydrate 0.1g, ammonium nitrate 2.0g, anhydrous CaCl 2 Mix 0.01g, 1.5g of dipotassium hydrogen phosphate, and 0.01g of disodium ethylenediaminetetraacetate, add to 1000mL of filtered culture water, adjust the pH to 7.0-7.5, and make it after autoclaving (121°C, 15min);

[0048] Nutrient medium: 10.0g of peptone, 3.0g of beef powder, 5.0g of sodium chloride, and 1.0g of glucose are mixed, added to 1000mL of filtered culture water, adjusted to pH 7.0-7.5, and prepared after high-pressure steam sterilization (121°C, 15min);

[0049] Enrichment culture solution: add malachite green and recessive malachite green to the nutrient medium, so that the concentrations of both malachite green and recessive malachite green are 5mg / L;

[0050] Malachite gr...

Embodiment 2

[0057] Embodiment 2: identification of malachite green and recessive malachite green degrading strain

[0058] The bacterial strain D6 that above-mentioned embodiment 1 is obtained carries out morphological characteristic and molecular biological identification, and the bacterium colony morphology of this bacterial strain is as follows figure 1 As shown, the scanning electron micrograph of the strain is shown in figure 2 with image 3 shown.

[0059] After identification, the main biological characteristics of the strain are:

[0060] Gram stain negative, bacillus; arginine dihydrolysis test, lysine decarboxylase test, β-galactosidase, ornithine decarboxylase reaction negative; sorbitol, lactose, rhamnose, glucose test positive; Sucrose can be used; the strain grows in fish farming environment, the optimum growth temperature is 28-35°C, and the optimum growth pH is 7.0-7.5. On solid medium, the colonies are round, thin, white and translucent.

[0061] The strain was ident...

Embodiment 3

[0062] Embodiment 3: Degradation test of malachite green and recessive malachite green degrading bacteria to malachite green and recessive malachite green in fishery water body

[0063] The bacterial strain D6 obtained after the separation and purification of Example 1 was measured as bacterium amount OD 600nm =0.2 Inoculate into 100mL 250mL Erlenmeyer flask containing fishery water body with malachite green and recessive malachite green drug concentration of 10mg / L, take the fishery water body without inoculation as control, shake on a constant temperature shaker at 30°C and 160rpm Cultivate for 36h. After the cultivation, the culture solution was centrifuged at 7000rpm for 5min, 100μL of the solution was taken, diluted into 10mL of water, 300μL of deuterated malachite green and deuterated recessive malachite green with a concentration of 0.2μg / mL were added, and 1mL of the sample was passed through the PRS solid phase. Extraction column, PRS column after activation (2mL ace...

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Abstract

The invention discloses a malachite green and recessive malachite green degrading bacterium, the degrading bacterium is Enterobacter sp. strain D6, preserved in China Center for Type Culture Collection (CCTCC), and the preservation date is On March 16, 2018, the preservation number is CCTCC No.M 2018133. The bacteria has the ability to simultaneously degrade malachite green and its metabolite recessive malachite green in the fish farming environment. The malachite green and recessive malachite green degrading bacteria of the present invention can safely, efficiently and quickly degrade residual malachite green and recessive malachite medicines in the fishery breeding environment at the same time, and can be used in the protection of fishery ecological environment and the control of drug residue pollution in the breeding environment It has a good application prospect in ensuring food safety.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a malachite green and recessive malachite green degrading bacteria, a separation and purification method and application thereof. Background technique [0002] Malachite green (Malachite green, MG), also known as basic green, base block green, malachite green, is a dark green crystalline solid, and malachite green is widely used as an industrial stain and a cytochemical stain. In 1933, it was found that malachite green has antibacterial effect. After that, many countries began to widely use malachite green as a fungicide, mainly for killing parasites in aquaculture water environment and mold in fish eggs. Malachite green is metabolized into colorless leucomalachite green (LMG) in aquatic animals. Malachite green and colorless leucomalachite green tend to remain in aquatic animals for a long time, resulting in highly teratogenic, carcinogenic and Mutagenic effect, recessive malachite...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/20C12N1/36C02F3/34C12R1/01C02F101/38C02F103/20
CPCC02F3/34C02F2101/308C02F2101/38C02F2103/20C12N1/20C12N1/36C12N1/205C12R2001/01
Inventor 刘菁华孙振中张玉平刘淑梅孙建华晏军黄雪玲
Owner 上海市水产研究所(上海市水产技术推广站)
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