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Endophytic fungus strain and metabolite extraction method and purpose thereof

A technology of endophytic fungi and metabolites, applied in microorganism-based methods, biochemical equipment and methods, botanical equipment and methods, etc., can solve the problems of no drug availability, difficult treatment, high mortality, and achieve high development value. and application prospects, easy cultivation, and low fermentation cost.

Active Publication Date: 2017-03-01
XUZHOU NORMAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In the face of these drug-resistant bacterial infections, currently available treatment options and drugs are very limited, or even no drugs are available, especially for immunodeficiency, the elderly and frail, after major surgery, long-term hospitalization, serious underlying diseases, long-term Patients with extensive use of broad-spectrum antibacterial drugs, admission to ICU wards, multiple organ insufficiency, chemotherapy or radiotherapy are difficult to treat and have a high mortality rate

Method used

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  • Endophytic fungus strain and metabolite extraction method and purpose thereof
  • Endophytic fungus strain and metabolite extraction method and purpose thereof
  • Endophytic fungus strain and metabolite extraction method and purpose thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] Example 1: Isolation, purification and preservation of endophytic fungal strain PF1167.

[0027] The experimental method and steps are as follows:

[0028] 1. Collection of samples: Collect the healthy leaves of the top ten fruits and immediately bring them back to the laboratory for the isolation of endophytic fungi.

[0029]2. Isolation of endophytic fungi: Rinse the collected leaves under running water to remove surface dust, cut the leaves into small squares about 0.5×0.5 cm in size after drying, and sterilize the surface as follows: Soak the small sample of leaves in 70% ethanol for 1 min, then transfer to 84 disinfectant (Eitefu group) diluted 40 times for 10 min, and finally rinse with sterile water 3 times (1 min each time). Connect surface-sterilized leaf pieces to plates containing PDA medium, place 5 small pieces evenly on each plate, place the above plate at 25°C for 3-7 days, and observe whether there are bacteria on the edge of the leaves every day silk ...

Embodiment 2

[0032] Example 2. Identification of endophyte strain PF1167.

[0033] 1. Morphological identification: Inoculate the endophytic fungal strain PF1167 on a new PDA medium, culture it at 25°C for 7-30 days, and observe the characteristics of the front and back of the colony every day. After it spores, take a glass slide, drop a drop of lactophenol oil in the center (recipe: 20 mL of heated and melted phenol; 20 mL of lactic acid; 40 mL of glycerin; 20 mL of water), pick a little mycelia, Place it in milk phenol oil, use an inoculation needle to disperse the mycelium, cover it with a cover glass, and observe its spores and sporulation structure and morphological characteristics under a microscope. of imeprfect fungi" (Barnett HL, Hunter BB 1998) for morphological identification. For the morphology of the front and back of the colony, see the appendix figure 1 , 2 , see attached image 3 , 4 . According to its morphological characteristics, it is identified as: Fusarium Ba...

Embodiment 3

[0060] Example 3. In vitro antibacterial experiment of endophytic fungus PF1167 metabolites.

[0061] 1. Experimental method (twice dilution method in micro broth)

[0062] 1) Bacterial strains involved: ① 20 strains of Escherichia coli or Klebsiella pneumoniae carrying β-lactamase gene recombinant plasmids; ② 3 clinical isolates carrying NDM-1 and other β-lactamase genes strains; ③ MRSA; ④ wild-type Escherichia coli, Staphylococcus aureus and Bacillus subtilis.

[0063] 2) Activation of strains: Take out each strain from the -80 ºC refrigerator, and inoculate them on Luria-Bertani (LB) medium plates respectively. For the bacterial strain carrying the recombinant plasmid of the β-lactamase gene, the medium used contains corresponding antibiotics, and the type and concentration of the antibiotics contained are consistent with the selection markers of the plasmid, as shown in Table 1 for details. For clinical isolates carrying multiple β-lactamase genes such as NDM-1, the medi...

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Abstract

The invention relates to an endophytic fungi strain and a metabolite extraction method and a purpose thereof, which belong to the field of microbe and microbe medicines. Through morphological characteristic observation and its rDNA ITS sequence analysis and identification, the strain is Fusarium sp, and has obvious bacteriostatic activity. A metabolite extract product of the strain has obvious bacteriostatic activity for wild bacterial strains such as escherichia coli, staphylococcus aureus, bacillus subtilis as well as drug-resistant strains such as beta-lactamase gene recombinant plasmid-carried escherichia coli or klebsiella pneumonia, and has obvious bacteriostatic activity for clinically separated superbacteria simultaneously carrying with a plurality of beta-lactamase gene escherichia coli, enterobacter cloacae and klebsiella pneumonia as well as methicillin-resistant staphylococcus aureus.

Description

technical field [0001] The invention relates to an extraction method and application of endophytic fungal strains and metabolites thereof, belonging to the field of microorganisms and microbial medicines. Background technique [0002] Bacterial drug resistance is becoming more and more serious around the world, and some "super bacteria" with high multi-drug resistance have emerged. In 2010, the world-renowned medical journal "The Lancet Infectious Diseases" ("Lancet-Infectious Diseases") reported a new Delhi metallo-β-lactamase-1 (New Delhi metallo-β-lactamase-1, Abbreviated as NDM-1) a new type of "super bacteria" with drug resistance genes. This "super bacteria" can tolerate almost all antibiotics and is only sensitive to colistin and tigecycline. Some The strains were even able to tolerate both drugs. The drug-resistant gene can be passed between strains through the plasmid, so that it can spread and spread rapidly. Such bacteria have been discovered in many countries ...

Claims

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Application Information

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IPC IPC(8): C12N1/14A01N63/02A01P1/00C12R1/645
CPCA01N63/10C12N1/14
Inventor 周生亮陈才法
Owner XUZHOU NORMAL UNIVERSITY
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