Novel application of pseudomonas aeruginosa fluorescene ironophore

A technology of Pseudomonas aeruginosa and fluorescent siderophore, which is applied in the field of environmental microorganisms and can solve problems such as not being disclosed

Active Publication Date: 2015-05-20
NINGBO UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, there are no relevant research reports on the use of Pseudomonas aeruginosa fluorescent siderophores to inhibit Vibrio brilliantis at home and abroad

Method used

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  • Novel application of pseudomonas aeruginosa fluorescene ironophore
  • Novel application of pseudomonas aeruginosa fluorescene ironophore
  • Novel application of pseudomonas aeruginosa fluorescene ironophore

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0022] Inhibition of the supernatant of Pseudomonas aeruginosa on the growth of Vibrio splendidus

[0023] Pseudomonas aeruginosa can secrete fluorescent siderophores in large quantities in inorganic salt medium with succinic acid as carbon source. Among them, the formula of the inorganic salt medium with succinic acid as the carbon source is: dipotassium hydrogen phosphate 6.0 g / L, potassium dihydrogen phosphate 3.0 g / L, ammonium sulfate 1.0 g / L, magnesium sulfate heptahydrate 0.1 g / L , succinic acid 4.0 g / L. Activate the Pseudomonas aeruginosa seeds stored at -80 ℃, insert the overnight cultured seed solution into the inorganic salt medium with succinic acid as the carbon source according to the inoculum amount of 10%, and cultivate it, and the liquid volume of the shake bottle is 30%. The temperature is 28-30 ℃, the cultivation speed is 100-120 rpm / min, and the culture is 48 hours. The bacterial culture was centrifuged at 10,000 rpm and the supernatant collected. The sup...

Embodiment 2

[0027] Purification of fluorescent siderophores

[0028]① Preparation of copper NTA agarose: Nickel NTA agarose was purchased from Shanghai Biotechnology Engineering Co., Ltd., and buffer A was used to wash off the chelated nickel ions on the agar. The composition of buffer A was 0.02 M sodium phosphate, 0.5 M NaCl, 0.05 M EDTA, pH 7.2. Wash the remaining buffer A with 5 volumes of distilled water. Add 0.5 ml of copper sulfate with a concentration of 0.1 M to chelate copper ions, wash off excess copper ions with 5 times the volume of distilled water, and the resulting copper NTA agarose;

[0029] ②Cultivation of Pseudomonas aeruginosa PA1: Activate the Pseudomonas aeruginosa seeds stored at -80 ℃, and insert the overnight cultured seed liquid into the inorganic salt medium with succinic acid as the carbon source according to the inoculation amount of 10% , the filling volume of the shake bottle is 30%, the culture temperature is 28-30 ℃, the culture speed is 100-120 rpm / min,...

Embodiment 3

[0032] Inhibitory effect of fluorescent siderophore on the growth of Vibrio splendidus in 2216E medium

[0033] The pH of the purified fluorescent siderophores in Example 2 above was adjusted to 7.0-7.2 with NaOH, sterilized by filtration with a 0.45 μm filter membrane, and stored for future use.

[0034] Vibrio splendidus preserved at -80 ℃ was inoculated into 2216E medium, and cultivated overnight in an air bath constant temperature shaker at 28-30 ℃ at 100-120 rpm / min. The cultured bacterial solution was inoculated into the sterilized 2216E medium according to the inoculation amount of 10%, so that the bacterial concentration reached 1.0 × 10 7 CFU / mL. Take 8 test tubes, add 5 mL of 2216E culture solution containing Vibrio splendidus to each tube, and add 0, 1, 5, 10, 20, 40, 80 and 160 μL of purified fluorescent siderophores, respectively. After culturing at 30°C for 24 hours, the absorbance of the cultured bacterial solution was measured at 600 nm by a UV spectrophoto...

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Abstract

The invention discloses novel application of pseudomonas aeruginosa fluorescene ironophore, which is characterized in that the fluorescene ironophore can inhibit the growth of vibro splendidus in 2216E culture medium or sea water, and is derived from pseudomonas aeruginosa; the pure fluorescene ironophore is obtained by carrying out affinity chromatography on a copper NTA agarose column; 3 percent volume of purified fluorescene ironophore is added into a 2216E culture medium containing vibro splendidus to culture at 30 DEG C for 24 hours, and the inhibition rate on vibro splendidus can be 50 percent; 1 percent volume of the purified fluorescene ironophore is added into sea water containing vibro splendidus to incubate at 30 DEG C for 24 hours, and the growth of the vibro splendidus in the sea water can be completely inhibited. The pseudomonas aeruginosa fluorescene ironophore is expected to play an important role in inhibiting pathogen vibro splendidus.

Description

technical field [0001] The invention relates to the field of environmental microorganisms, in particular to a new application of Pseudomonas aeruginosa fluorescent siderophore. Background technique [0002] With the rapid development of intensive marine aquaculture in recent years, corresponding aquaculture problems have become increasingly prominent. Intensive marine aquaculture can easily lead to the occurrence of aquaculture biological diseases. Bacterial diseases have caused huge economic losses to the mariculture industry. According to relevant statistics, the annual loss of mariculture industry caused by diseases is as high as 1.25 billion U.S. dollars, which is a serious loss. Vibrio splendidus is an opportunistic pathogen that widely exists in seawater environment. The pathogenicity of Vibrio splendidus is affected by many factors such as host physiological status and environmental conditions. Under normal conditions, Vibrio splendidus does not cause biological di...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C02F3/34C02F103/08
CPCC02F3/34C02F2103/08C02F2303/04
Inventor 张卫卫李成华梁伟康
Owner NINGBO UNIV
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