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Cupriavidus and application thereof

A technology of copper greedy bacteria and strains, applied in the field of microorganisms, can solve the problem of inability to degrade 2-chloro-4-nitrophenol, etc., and achieve a good effect of bioremediation

Active Publication Date: 2019-06-11
YANTAI INST OF COASTAL ZONE RES CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, studies have reported that none of the above bacteria can degrade 2-chloro-4-nitrophenol when the concentration of 2-chloro-4-nitrophenol reaches or exceeds 0.5mM

Method used

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  • Cupriavidus and application thereof
  • Cupriavidus and application thereof
  • Cupriavidus and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0020] Embodiment 1: Isolation, purification and identification of bacterial strains

[0021] (1) Configure inorganic salt medium:

[0022] The inorganic salt medium is composed of disodium hydrogen phosphate (Na 2 HPO 4 12H 2 O) 14.3g; Potassium dihydrogen phosphate (KH 2 PO 4 ) 3g; magnesium sulfate (MgSO 4 ·7H 2 O) 0.06mg; ferrous sulfate (FeSO 4 ·7H 2 O) 0.3mg; manganese sulfate (MnSO 4 ·H 2 O) 0.28mg; copper sulfate (CuSO 4 ) 0.05mg; Zinc sulfate (ZnSO 4 ) 0.05mg; boric acid (H 3 BO 3 ) 0.05 mg. The above ingredients were dissolved in double distilled water and the volume was adjusted to 1000ml, adjusted to pH 7.0, and autoclaved at 121°C for 20 minutes.

[0023] (2) Enrichment of 2-chloro-4-nitrophenol-degrading bacteria:

[0024] Sampling soil contaminated by a pesticide manufacturing plant, adding 5 g of soil to a 250 ml Erlenmeyer flask containing 100 mL of inorganic salt medium, and adding 0.3 mM 2-chloro-4-nitrophenol as the only carbon and nitrogen ...

Embodiment 2

[0029] Example 2: Analysis of bacterial strains' ability to degrade 2-chloro-4-nitrophenol and identification of metabolic intermediates

[0030] (1) Analysis of degradability

[0031] Inoculate the above single colony in inorganic salt medium + LB (volume ratio = 4:1), add 0.3mM 2-chloro-4-nitrophenol, collect the bacteria after overnight culture, and wash with inorganic salt medium 2 times, then press initial OD 600 =0.05 inoculum amount was inoculated into fresh inorganic salt medium (adding 0.3mM 2-chloro-4-nitrophenol), placed in a shaker at 180rpm, cultivated at 30°C and sampled at intervals, centrifuged at 12000rpm, and the supernatant was collected. Use HPLC to detect the concentration of 2-chloro-4-nitrophenol in the supernatant, and use nitrite reagent: 1% sulfanilic acid and 0.02% N-(1-naphthalene) ethylenediamine to detect nitrite , use a chloride ion selective electrode to detect chloride ions, and use a spectrophotometer to detect the bacterial concentration OD...

Embodiment 3

[0038] Embodiment 3: Copper greedy bacteria CNP-8 restores 2-chloro-4-nitrophenol polluted soil

[0039] (1) Soil collection and soil physical and chemical index determination

[0040] Soil samples were collected in the campus of China Agricultural University (Yantai), and there was no history of 2-chloro-4-nitrophenol pollution. The 0-15cm soil was collected to remove the topsoil. Collect after passing through a 2mm sieve and store at 4°C. After testing, the total carbon, total nitrogen and water content of the soil were 0.86%, 0.074% and 10.64% respectively, and the pH value was 7.38.

[0041] (2) Preparation of Inoculated Bacteria

[0042] The strains were cultured with liquid LB medium or inorganic salt + LB (volume ratio = 4:1, and 0.3mM 2-chloro-4-nitrophenol was added at the same time) to the late logarithmic period, and the bacteria were collected by centrifugation at 6000×g for 8 minutes. Then wash twice with 0.85% sterile physiological saline, and set aside.

[0...

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Abstract

The invention relates to the field of microorganisms, in particular to cupriavidus and application of the cupriavidus. The cupriavidus is preserved in China Center for Type Culture Collection, the address is Wuhan University, Wuhan, China, the preservation date is September 25, 2017, and the preservation number is CCTCC M 2017546. Bacterial strain obtained by the invention can degrade 2-chloro-4-nitrophenol by taking phloroglucinol as a ring-opening substrate, and the strain provided by the invention is gram-negative bacteria capable of metabolizing 2-chloro-4-nitrophenol by a 1,2,4-benzenetriol way. On the other hand, the prepared remediation microbial inoculum can completely degrade 100 ppm of 2-chloro-4-nitrophenol in contaminated soil within 12 days. Therefore, the strain has good application prospects in the biological treatment of 2-chloro-4-nitrophenol pollution.

Description

technical field [0001] The invention relates to the field of microorganisms, in particular to a cupriavidus and its application. Background technique [0002] At present, chloronitroaromatic compounds are produced in large quantities and are widely used in industries such as agriculture, industry and pharmaceutical industry. Most of these compounds are toxic to the human body, and have enrichment and amplification effects in the food chain. They have blood toxicity, immunotoxicity, liver toxicity, kidney toxicity, teratogenicity and carcinogenicity, which seriously threaten human health and survival. Artificially synthesized chloronitroaromatic compounds will inevitably leak into the environment during use, and a large amount of leakage will occur in the event of an accident, causing serious environmental pollution. 2-Chloro-4-nitrophenol belongs to a typical chloronitroaromatic compound and is an important chemical raw material. It is often used as a precursor for the synt...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20B09C1/10C02F3/34C02F101/34C02F101/36C02F101/38C12R1/01
Inventor 闵军胡晓珂陈卫卫
Owner YANTAI INST OF COASTAL ZONE RES CHINESE ACAD OF SCI
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