Method for detecting Escherichia coli in water and detection culture solution

A technology of Escherichia coli and culture solution, which is applied in the direction of biochemical equipment and methods, and the determination/inspection of microorganisms, which can solve the problems affecting the detection accuracy and detection time, small sampling volume, undisclosed chromogenic medium, etc. problems, to achieve real-time monitoring of microbial contamination, reduced detection time, and short detection time

Active Publication Date: 2014-10-08
HANGZHOU GREAN WATER SCI & TECH INC
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, the detection object of this method is coliform bacteria, and its detection method is not suitable for the detection of Escherichia coli. At the same time, the most critical chromogenic medium has not been disclosed, and these all affect the detection accuracy and detection time. Key factor
In addition, the existing detection methods based on the enzyme substrate method all have the problem of too little sampling volume, generally only 55.5-100mL can be obtained, and the sampling volume can affect the uncertainty of the detection results

Method used

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  • Method for detecting Escherichia coli in water and detection culture solution
  • Method for detecting Escherichia coli in water and detection culture solution
  • Method for detecting Escherichia coli in water and detection culture solution

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Experimental program
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Embodiment 1

[0037] Embodiment 1: method of the present invention

[0038] Detection medium: Tryptone 10.0g, magnesium sulfate 50mg, manganese sulfate 0.5mg, zinc sulfate 0.5mg, ammonium sulfate 5.0g, sodium chloride 10.0g, calcium chloride 50mg, sodium sulfite 40mg, Tween-802g, NaH 2 PO 4 ·H 2 O6.10g, K 2 HPO 4 ·3H 2 O2.75g, o-nitrobenzene-β-D-galactopyranoside 250mg, 4-methylumbelliferone-β-D-glucuronide 35mg, No. 3 bile salt 3.0g.

[0039] A series of suspensions of Escherichia coli with different concentrations (5.2, 52, 5.2×10 2 , 5.2×10 3 , 5.2×10 4 , 5.2×10 5 , 5.2×10 6 , 5.2×10 7 MPN / 100mL) each 100mL, first filter and concentrate through a cellulose acetate filter membrane with a membrane pore size of 0.45 μm and a membrane diameter of 47 mm, then add the filter membrane to the test culture solution for cultivation, the culture temperature is 37 ° C, and the culture solution is continuously tested during the culture period The 450nm fluorescence intensity produced under...

Embodiment 2

[0042] Embodiment 2: method of the present invention

[0043] Detection medium: Tryptone10.0g, magnesium sulfate 100mg, manganese sulfate 1mg, zinc sulfate 1mg, ammonium sulfate 10.0g, sodium chloride 10.0g, calcium chloride 100mg, sodium sulfite 100mg, Tween-800.5g, NaH 2 PO 4 ·H 2 O6.10g, K 2 HPO 4 ·3H 2 O2.75g, o-nitrobenzene-β-D-galactopyranoside 250mg, 4-methylumbelliferone-β-D-glucuronide 35mg, No. 3 bile salt 1.0g.

[0044] A series of suspensions of Escherichia coli with different concentrations (6.3, 63, 6.3×10 2 , 6.3×10 3 , 6.3×10 4 , 6.3×10 5 , 6.3×10 6 , 6.3×10 7 MPN / 100mL) each 100mL, first filter and concentrate through a cellulose acetate filter membrane with a membrane pore size of 0.45 μm and a membrane diameter of 47 mm, then add the filter membrane to the test culture solution for cultivation, the culture temperature is 37 ° C, and the culture solution is continuously tested during the culture period The 450nm fluorescence intensity produced unde...

Embodiment 3

[0047] Embodiment 3: comparison of detection time

[0048] Dilute with sterile water to prepare a series of Escherichia coli suspensions with different concentrations of 100mL each, first filter and concentrate through a cellulose acetate filter membrane with a membrane pore size of 0.45 μm and a membrane diameter of 47 mm, and add the filter membrane to the detection culture solution Cultivate at a temperature of 37°C. During the cultivation period, continuously detect the 450nm fluorescence intensity of the culture solution under the 365nm excitation light, and record the fluorescence intensity of the 450nm fluorescence produced by the Escherichia coli suspension at each concentration under the 365nm excitation light. 240 hours of time.

[0049] At the same time, according to the linear curve and linear equation of the patent 200910148688.7, the different concentrations of Escherichia coli suspensions prepared in this example were substituted into it, and the detection time ...

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Abstract

The invention relates to the technical field of biology, and discloses a method for detecting Escherichia coli in water and a detection culture solution. The method comprises the following steps: providing a series of Escherichia coli suspensions with different concentrations, filtering, concentrating, adding into the detection culture solution, culturing, recording the characteristic time of the Escherichia coli suspensions with different concentrations, and establishing a linear equation on Escherichia coli suspension concentration and characteristic time; filtering and concentrating a water sample to be detected, adding into the detection culture solution, culturing, recording the characteristic time of the water sample to be detected on the premise that the 410nm absorbance has obvious changes, and substituting the characteristic time in the linear equation to obtain the concentration value. On the basis of the principle of the enzyme substrate process, by using absorbance and fluorescence intensity as the standard and adopting the detection culture solution suitable for Escherichia coli, the method can quickly and accurately detect the concentration of Escherichia coli in water and can monitor the microbial contamination condition in water in real time.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a method and a kit for detecting Escherichia coli in water. Background technique [0002] Total coliform, thermotolerant coliform and Escherichia coli are the three most important bacteriological indicators indicating fecal pollution in water bodies. These bacteria can be detected in human and animal feces, and can also be detected in nutrient-rich water, that is, in the case of non-fecal pollution, it is also possible to detect these bacteria. The composition of the heat-resistant coliform group is the same as that of the total coliform group, but the main composition is Escherichia. In this genus, there is only one species closely related to human life, that is, Escherichia coli. For example, the genera of Citrobacter, Klebsiella and Enterobacter accounted for a small number. As the indicator bacteria of fecal contamination, the detection of Escherichia coli is the most significa...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/06
Inventor 单旭亮毛芳芳崔海松
Owner HANGZHOU GREAN WATER SCI & TECH INC
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