Method for auxiliary identification of comma bacillus by matrix-assisted laser desorption ionisation-time-of-flight mass spectrometry (MALDI-TOF MS)

A MALDI-TOF-MS and Vibrio cholerae technology, applied in the field of assisted identification by matrix-assisted laser desorption ionization time-of-flight mass spectrometer, can solve the problems of insufficient database resources, lack of characteristic spectra of Vibrio cholerae, etc., and achieve simple sample preparation Effect

Inactive Publication Date: 2011-11-23
曹际娟
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, for the detection of specific microorganisms, the popularization of this technology has the problem of insufficie

Method used

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  • Method for auxiliary identification of comma bacillus by matrix-assisted laser desorption ionisation-time-of-flight mass spectrometry (MALDI-TOF MS)
  • Method for auxiliary identification of comma bacillus by matrix-assisted laser desorption ionisation-time-of-flight mass spectrometry (MALDI-TOF MS)
  • Method for auxiliary identification of comma bacillus by matrix-assisted laser desorption ionisation-time-of-flight mass spectrometry (MALDI-TOF MS)

Examples

Experimental program
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Effect test

Embodiment 1

[0055] (1) Select Vibrio cholerae strains

[0056] 42 strains of Vibrio cholerae collected by Liaoning Entry-Exit Inspection and Quarantine Bureau were selected, including 23 strains of Vibrio cholerae O1 group, 11 strains of Vibrio cholerae O139 group and 8 strains of Vibrio cholerae non-O1 / non-O139 group Vibrio cholerae, specific information See Table 1. Vibrio cholerae involved in Table 1 was further confirmed to be Vibrio cholerae through API-20E biochemical identification and automatic bacterial biochemical analyzer identification, which can be used to establish a standard map for MALDI-TOF MS identification.

[0057] Table 1

[0058]

[0059]

[0060] (2) Sample processing:

[0061] The isolates of Vibrio cholerae were respectively inoculated on NA medium, and cultured at 37° C. for 24 hours to obtain fresh strain cultures. Take a fresh culture of Vibrio cholerae, add 300 μL of pure water to an Eppendorf tube, pick an appropriate amount (5-10 mg) of bacteria, mi...

Embodiment 2

[0082] MALDI-TOF-MS clustering analysis of 42 strains of Vibrio cholerae

[0083] On the basis of Example 1, the similarity of the main spectra of 42 strains of Vibrio cholerae in the obtained Vibrio cholerae MALDI-TOF-MS database was calculated by pattern matching, and the similarity score was used to construct a phylogenetic tree, and the 42 strains were clustered Type analysis, the resulting cluster type dendrogram is attached figure 2 shown. The level of difference represents the relatedness to each other with a value between 0 and 1.6. A difference of 0 means that the two MALDI-TOF-MS mass spectra are identical and can be assigned to the same class; a difference of 1.6 means that the cluster analysis results of the existing data show that the relationship between the two strains is the farthest. The number of types of Vibrio cholerae is different when the selected difference level is different. When the difference level was 1, the MALDI-TOF-MS typing results divided t...

Embodiment 3

[0086] MALDI-TOF-MS clustering analysis of 42 strains of Vibrio cholerae

[0087] Among the 42 strains of Vibrio cholerae in Example 1, 23 strains belonged to O1 group Vibrio cholerae, 11 strains belonged to O139 group Vibrio cholerae, and the remaining 8 strains were all non-O1 non-O139 group Vibrio cholerae. Get the mass-to-charge ratio data of 42 strains of Vibrio cholerae in the database, further utilize Matlab R-2007a (Matlab Com., USA) software, carry out principal component analysis (PCA) to the spectrogram of these 42 strains of Vibrio cholerae, PCA analysis result as attached image 3 shown. The results showed that 23 strains of O1 group Vibrio cholerae were relatively concentrated (the box A area in the figure), while the other 19 strains of O139 and non-O1 / non-O139 group Vibrio cholerae were relatively scattered. This indicated that the major constituents were basically the same among O1 V. cholerae and significantly different from those of O139 and non-O1 / non-O139...

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Abstract

The invention relates to a method for the auxiliary identification of comma bacillus by matrix-assisted laser desorption ionisation-time-of-flight mass spectrometry (MALDI-TOF MS). The method comprises the following steps of: 1, selecting 40 to 45 plants of fresh comma bacillus cultures, and pretreating the samples; 2, collecting maps of all bacterial strain samples; 3, analyzing and unitizing the obtained maps by using BioTyper software to obtain a comma bacillus standard map; 4, preparing samples of microbes to be detected by the step 1; 5, collecting MALDI-TOF MS maps of the samples to be detected according to the step 2; and 6, comparing the MALDI-TOF MS maps, which are obtained in the step 5, of the samples to be detected with the comma bacillus standard map, and determining a detection result according to matching fractions. By the method, a comma bacillus identification mass spectrogram database and the comma bacillus standard map are established successfully, so the simple, convenient, microscale, supermatic and accurate identification of the comma bacillus can be realized.

Description

technical field [0001] The invention relates to a new detection method for Vibrio cholerae, in particular to a method for assisting identification by using a matrix-assisted laser desorption ionization time-of-flight mass spectrometer (MALDI-TOF MS). Background technique [0002] Vibrio cholerae is the pathogenic bacteria of cholera, a severe intestinal infectious disease. Cholera has an acute onset and rapid spread. The main symptoms are severe diarrhea and vomiting, which can cause severe dehydration, peripheral circulatory failure and acute renal failure. If it is not treated in time, it is easy to die . Vibrio cholerae has been found to have more than 200 serotypes, and it has been reported that only two serotypes, O1 and O139, can cause outbreaks of cholera (Epidemiology, genetics, and ecology of toxic Vibrio cholera, Mol.Biol.1998.Rev.62 : 1301-1314.). Vibrio cholerae exists widely in rivers, lakes, and sea water, and even exists on the surface and body of aquatic an...

Claims

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Application Information

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IPC IPC(8): G01N27/64G01N1/28
Inventor 曹际娟
Owner 曹际娟
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