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Respiratory tract infection pathogen detection method based on metagene/metatranscriptome sequencing

A macrotranscriptome and detection method technology, which is applied in the field of respiratory infection pathogen detection based on metagene/metatranscriptome sequencing, can solve the problems of inability to identify RNA viruses, inability to detect microbial gene expression and analyze microbial activity, etc.

Pending Publication Date: 2022-03-01
SHANGHAI CHILDRENS MEDICAL CENT AFFILIATED TO SHANGHAI JIAOTONG UNIV SCHOOL OF MEDICINE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] Although pathogenic metagenomics is a powerful tool for identification of pathogen species, it can detect the genetic information of pathogens, but it cannot detect the gene expression of microorganisms, analyze the activity of microorganisms, and cannot identify RNA-like viruses, while pathogenic metatranscriptomes can be easily Make up for this shortcoming of mNGS

Method used

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  • Respiratory tract infection pathogen detection method based on metagene/metatranscriptome sequencing
  • Respiratory tract infection pathogen detection method based on metagene/metatranscriptome sequencing
  • Respiratory tract infection pathogen detection method based on metagene/metatranscriptome sequencing

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] Example 1 Sputum Sample Detection of Pathogens of Children's Respiratory Tract Infectious Diseases

[0044] Step S1: Process the collected children's sputum: In the metagenomic sequencing process, take 600 μL of the sputum sample to be tested into a 2 mL grinding tube filled with 500 μL of glass beads, and tighten the cap of the tube to ensure no leakage when the wall is broken. Further, put the 2mL grinding tube into the 2mL grinding tube holder, balance it, turn on the grinding homogenizer, and use the following procedure to break the wall and process the parameters as shown in Table 4.

[0045] Table 4 Broken wall treatment parameters

[0046]

[0047] After breaking the wall, take out the grinding tube and briefly centrifuge for 5s in an AQBD cubee mini centrifuge. The grinding homogenizer is Tiangen Bioprep-24 tissue grinding homogenizer. Then use the DNA extraction kit produced by Guangzhou Weiyuan Medical Instrument Co., Ltd. to extract DNA according to the ...

Embodiment 2

[0068] Example 2 Bubble Lavage Fluid Sample Detects Pathogens of Children's Respiratory Tract Infectious Diseases

[0069] In this embodiment, a sample of alveolar lavage fluid is used, and other steps and parameters are the same as those in Embodiment 1.

Embodiment 3

[0070] Example 3 Swab sample with preservation solution detects pathogens of respiratory tract infectious diseases in children

[0071] In this embodiment, a swab sample with preservation solution is used, and other steps and parameters are the same as in Embodiment 1.

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Abstract

The invention discloses a children respiratory tract infectious disease pathogen detection method based on metagenome / metatranscriptome sequencing, and is characterized in that the children respiratory tract infectious disease pathogen detection method specifically comprises the following steps: treating a to-be-detected sample, and respectively extracting DNA and RNA by using a DNA extraction kit and an RNA extraction kit; constructing a sample DNA sequencing library and an RNA sequencing library; high-throughput sequencing is carried out on the sequencing library, and sequencing is carried out on negative control of DNA and RNA extraction and library establishment on the same lane at the same time; comparing the sample sequence data with the reference database to obtain a sequence of which the comparison ratio is 70% or above and which is not subjected to multiple comparison, then screening pathogenic microorganisms, and detecting pathogens of respiratory tract infectious diseases in the sample according to a screening result. According to the method, pathogens of respiratory tract infectious diseases of children are rapidly and clearly determined, colonization and infection are distinguished, so that a targeted treatment strategy is formulated, and the hospitalization time, the medical cost, the antibiotic use days and the death rate of child patients are reduced.

Description

technical field [0001] The invention belongs to the technical field of microorganism detection, and in particular relates to a method for detecting pathogens of respiratory tract infection based on macrogene / metatranscriptome sequencing. Background technique [0002] Acute respiratory tract infection (ARTI) is the leading cause of outpatient and hospitalization for children of all ages, especially in winter and spring; at the same time, ARTI is also the second leading cause of death for children under 5 years of age, becoming the main public health problem faced by children. Sanitation threat. Taking the single disease of pneumonia as an example, the WHO survey shows that more than 2 million children aged 0-5 die of pneumonia every year, accounting for almost 1 / 5 of the deaths of children under 5 years old in the world; in China, 14.8% of deaths in children under 5 are caused by pneumonia. Therefore, improving the diagnosis and treatment of children's respiratory tract inf...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6869C12Q1/70C12Q1/04
CPCC12Q1/6869C12Q2531/113C12Q2535/122C12Q2525/191C12Q2545/113
Inventor 莫茜陶悦杜白露
Owner SHANGHAI CHILDRENS MEDICAL CENT AFFILIATED TO SHANGHAI JIAOTONG UNIV SCHOOL OF MEDICINE