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Method for detecting pathogenic vibrios by MALDI-TOF-MS (Matrix Assisted Laser Desorption Ionization-Time-Of-Flight-Mass Spectrometry)

A MALDI-TOF-MS, pathogenic Vibrio technology, which is applied in the construction and application of the MALDI-TOF-MS database, can solve the problems of geographical differences of strains, imperfect identification data of pathogenic Vibrio, etc.

Inactive Publication Date: 2014-06-18
麻丽丹 +3
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, most of the spectra in the database are from foreign strains. Due to the fact that the research and application of this technology has just started in China, the mass spectrogram database of various Vibrio strains has not yet been established to identify pathogenic Vibrio pathogens. Data are far from complete, and there may be significant geographical differences in strains from country to country

Method used

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  • Method for detecting pathogenic vibrios by MALDI-TOF-MS (Matrix Assisted Laser Desorption Ionization-Time-Of-Flight-Mass Spectrometry)
  • Method for detecting pathogenic vibrios by MALDI-TOF-MS (Matrix Assisted Laser Desorption Ionization-Time-Of-Flight-Mass Spectrometry)
  • Method for detecting pathogenic vibrios by MALDI-TOF-MS (Matrix Assisted Laser Desorption Ionization-Time-Of-Flight-Mass Spectrometry)

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0056] Embodiment 1, MALDI-TOF-MS detects Vibrio parahaemolyticus

[0057] Use the determined appropriate medium and culture time: 2% NaCl brain heart extract agar, culture 29 strains (strain number 1-29) at 36°C for 24 hours, weigh 8±2 mg colonies each, and pretreat the preparation with mixed solvent treatment. After the samples were formed, the accuracy of the experimental strains was analyzed using the conditions determined by the above research content. Data acquisition software Flexcontrol by MALDI-TOF-MS TM 3.0 Collect data from the sample wells, and then pass the MALDI-TOF-MS identification software Biotyper TM 2.0 Analyze the obtained mass spectra of each strain and match them with the resources in the database. The MALDI-TOF-MS identification results were compared with the biochemical identification results.

[0058] The identification results and identification scores were obtained and compared with the biochemical identification results (Table 2). The MALDI-TOF-...

Embodiment 2

[0063] Example 2. MALDI-TOF-MS detection of Vibrio mazei

[0064] The method established by the present invention was used to carry out 6 repeated analyzes on the standard strain of Vibrio mazei (ATCC700040). After all the strains were detected by MALDI-TOF-MS, the operation steps of the content of the invention were followed, and the 6 mass spectra obtained by each strain were compared, as well as the identification scores and identification results. The 6 mass spectra obtained by the strain are all very close (see figure 2 ), the identification results were completely consistent, and the identification scores were all above 2.3 points. It shows that the MALDI-TOF-MS detection method of the present invention has ideal repeatability.

Embodiment 3

[0065] Embodiment 3, MALDI-TOF-MS detection method specificity analysis

[0066] Using the method and conditions established by the present invention to treat Vibrio parahaemolyticus (ATCC17802), Vibrio vulnificus (ATCC27562), Vibrio mimeticum (ATCC33653), Vibrio meibii (ATCC700040) and other bacterial strains ATCC25922, ATCC25923, ATCC13076, ATCC19111 Identification by MALDI-TOF-MS. The specificity of the MALDI-TOF-MS detection method was evaluated by the obtained identification results, scores and mass spectra.

[0067] MALDI-TOF-MS accurately identified 4 standard strains belonging to the genus Vibrio (ATCC17802, ATCC27562, ATCC33653 and ATCC700040) and 4 standard strains of different genera (ATCC25922, ATCC25923, ATCC13076, ATCC19111). Obtained mass spectrograms through observation, the four standard strains of Vibrio parahaemolyticus, Vibrio vulnificus, Vibrio mimeticus and Vibrio martensili, which belong to the genus Vibrio, have mass-to-charge ratios (m / z) in the range...

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Abstract

The invention discloses a method for detecting pathogenic vibrios by MALDI-TOF-MS (Matrix Assisted Laser Desorption Ionization-Time-Of-Flight-Mass Spectrometry). The method comprises the following steps: acquiring MALDI-TOF-MS data information of a strain to be detected, and comparing the MALDI-TOF-MS data information with acquired information in an MALDI-TOF-MS database so as to obtain an identification result, wherein the MALDI-TOF-MS database is a database including MALDI-TOF-MS data information of selected microorganisms, and the selected microorganisms are microorganisms with strain numbers of 1-128 in a table 1. The invention also provides an MALDI-TOF-MS database, and a newly added base strain comprises 128 common pathogenic strains, which are wide in strain source; the constructed MALDI-TOF-MS strain database is large in amount of information. By using the constructed data, detection and identification of an unknown strain can be realized with high sensitivity, high specificity, high speed, high efficiency and high flux.

Description

technical field [0001] The invention relates to a MALDI-TOF-MS detection method for common pathogenic Vibrio in the environment and food, in particular to the construction and application of the MALDI-TOF-MS database. Background technique [0002] Matrix assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF-MS) technology is a new foodborne pathogenic bacteria identification and traceability technology gradually developed in recent years. The technology has two outstanding features: accurate and rapid identification of pathogenic bacteria and simultaneous tracing of pathogenic bacteria protein typing. In addition to the above two outstanding features, MALDI-TOF-MS technology also has the advantages of high detection sensitivity, high automation of instrument analysis, and low detection cost. Before using MALDI-TOF-MS for identification, the strains can be collected after simple pretreatment. In the subsequent process, high-throughput identificatio...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/33G01N1/28G01N1/38
Inventor 麻丽丹王殿夫高世光闫平平
Owner 麻丽丹
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