Source: Xiaojudeng.com
Author: quite
Matrix Assisted Laser Desorption Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS) is a new technology applied in the field of clinical microbiological testing. The advent of this technology provides a good tool for the accurate identification of bacteria, fungi and mycobacteria . The European microbiology laboratory first started using MALDI-TOF MS. This technique is widely adopted in microbiology laboratories worldwide due to its simplicity, high accuracy, fast turnaround time, and low cost. Thanks to the rapid development of mass spectrometry in the identification of microorganisms, domestic researchers have gradually paid attention to the application of MALDI-TOF MS in the field of microorganisms.
History and Development
Although the use of mass spectrometry for bacterial identification was first proposed in 1975, the ionization method used at that time would fragment proteins, thus hindering its ability to analyze effectively. In the 1980s, two teams proposed a similar idea to to detect the complete macromolecule of , and together they invented the technology of using MS to study the complete (Koichi Tanaka) and proteins. His colleagues developed a method to use ultrafine metal powder mixed with glycerol to analyze the biomacromolecule without destroying the macromolecules. For this, Koichi Tanaka was awarded the Nobel Prize in Chemistry (shared with John Fenn and Kurt Wiith rich) [2] At the same time that Koichi Tanaka published their research, Franz Hillen kamp and Michael Karas published the soft desorption of niacin using the organic compound substrate Ionization to analyze the study of biological macromolecules [3] Matrix-assisted laser desorption ionization (matrix-assisted laser desorption MALDI) The term is also derived from this.In 2006, Fox published the first article on the application of mass spectrometry in clinical microbiology in the Journal of Clinical Microbiology, which attracted the attention of peers. He believed that there were still many technical difficulties to be overcome in the identification of bacteria by this technology, and it was not easy to become a routine method, but in 2009 After a few years, microbial mass spectrometry technology quickly swept some countries in Europe, North America and Asia. Matrix-assisted laser desorption ionization ion source and matrix-assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF MS) technology meet the expectations of microbiologists: simple operation, rapid identification, low operation Cost and automatically updated database. It is particularly outstanding in the identification of difficult-to-cultivate microorganisms such as microaerophilic bacteria, anaerobic bacteria , fungi and mycobacteria, and the identification time is advanced from days to minutes and seconds. The new development projects are even more encouraging, such as the direct identification of sterile body fluids, the identification of drug resistance phenotypes, the analysis of pathogenic toxins, and the genetic traceability of the nosocomial pathogen .
Related national standards and guidelines
Friends in the industry all know that a major problem of mass spectrometry in the clinical field is the lack of standardization and standardization. But MALDI TOF MS is relatively comprehensive in its standards and consensus.
National Standard "GB/T 33682-2017 General Principles of Matrix-Assisted Laser Desorption Ionization Time-of-Flight Mass Spectrometry for Identification of Microorganisms"
"Expert Consensus on Quality Assurance of MALDI-TOF MS Pathogen Identification" Consensus "
"Expert Consensus on Basic Requirements for Clinical Microbiology Laboratory Construction"
"Expert Consensus on the Application of Nucleic Acid Mass Spectrometry in China" Published the health standard "Basic Technical Requirements for Clinical Microbiology Testing"
Clinical application
Drug resistanceDetection
The microbiological laboratory must not only provide accurate pathogenic bacteria identification results, but also test its sensitivity to provide treatment for clinicians in accordance with. Conventional drug susceptibility testing methods are time-consuming, and some ELISA, agglutination and other methods can only be limited to a few bacteria. MALDI-TOF MS can provide a good platform for the detection of drug resistance genes, and can analyze almost all drug resistance mechanisms. The currently reported methods are mainly based on the following principles: analysis of antibacterial drug and its modified products; Cell components; analysis of ribosomal DNA methylation; detection of mutations, etc.
Epidemiology
One of the challenges faced by microbiology laboratories, infection control personnel and clinicians is to obtain specific data on the representative classification of a clinical strain in the event of an outbreak, such as Salmonella, Streptococcus Etc., in order to accurately obtain the serotype, subtype or other classification, the laboratory must spend more time for further testing, and many strain types require special test methods, instruments, consumables, etc. Therefore, laboratories with limited conditions need to send out testing, which may lead to prolonged testing time, loss of important epidemiological data, increased testing costs, and inaccurate results. However, if the results of important data are completed quickly and accurately in the microbiology laboratory, it will help clinicians, nurses, infection control personnel and health institutions to process, track and control the prevalence of infectious pathogens.
At present, the ability of MALDI-TOF MS to identify these highly related strains will increase the influence of clinical microbiology laboratories through group, genus, and species analysis of a large number of strains, as well as in-depth research on the typing of many strains. More sophisticated MALDI-TOF MS instruments may appear in the future, which can not only identify pathogenic bacteria, but also provide rapid and accurate epidemiological data of strains, which will enable clinical microbiology laboratories to control infections in hospitals, outbreaks of pathogenic bacteria, and -resistant bacteria. plays an important role in the fields of national surveillance and biological defense.
Identification of common bacteria
Conventional microbial identification is mainly based on biochemical reactions, microscopy, immunology or molecular biology. These analytical methods require high expertise and are time-consuming and labor-intensive. Antibiotics are used empirically by clinicians while awaiting results, but are sometimes inappropriate or wrong.Modern clinical microbiology laboratories require rapid, reliable and economical methods to identify bacteria for early appropriate antibacterial drug therapy. Mass spectrometry shortens the identification of microorganisms from 24~48h to minutes, and only needs to analyze a very small amount of bacteria (104~106 CFU/mL). Taking 16SrRNA gene sequencing results as the standard, the correct identification rate of by mass spectrometer was 90.0%~95.0%, which is much higher than the previous commercial automated identification system.
List of NMPA-certified companies
In recent years, the R&D and production of MALDI-TOF MS instruments in China has started rapidly, and a number of scientific researchers and enterprises have emerged, which has greatly promoted the process of localization of MALDI-TOF MS. MALDI-TOF MS will likely become the first mass spectrometer category in which Chinese companies have mastered the most advanced core technology and led the development of technology, which will greatly promote my country's research on biological macromolecular analysis and clinical molecular diagnostic applications. . It can be seen from the data that from 2020 to the present, there are 9 newly-approved domestic time-of-flight mass spectrometry .
challenge
The accuracy of mass spectrometry identification depends on the quality of the strain database. Incomplete information in the strain database will limit the accuracy of the identification. Enriching the strain database will help the accuracy of the identification results. Although self-built database can improve the identification rate, it is time-consuming and requires the guidance of professional technicians and mycology experts. In addition, the database of IVD certification is not allowed to be increased at will, so it can only be used for scientific research. Second, although MALDI-TOF MS can quickly and effectively identify bacteria, its guiding significance for subsequent antimicrobial susceptibility testing remains controversial.
Although the results obtained by MALDI-TOF MS are generally reproducible, variations are possible.Mass spectrometers (different types and individual instruments, instrument age, instrument configuration), matrix and solvent composition, preparation methods, technician training and competence, culture conditions (eg, culture medium, time and temperature of strain culture), and biological variability are all possible affect its results. Therefore, its standardized quality control strategy still needs to be formulated. In addition, the isolated colony must be tested. If the colony is not well separated, there may be a variety of microorganisms, and the bacterial species with a small proportion may not be found.
Reference:
[1] FoxA. Mass spectrometry for species or strain identification after culture or without culture: past, present, and future[J]. J Clin Microbiol, 2006, 44:2677-2680.
[2] Cao Shujuan, Liu Bin, Zhu Anna. Application and prospect of MALDI-TOF MS in pharmacogenomics detection [J]. Journal of Molecular Diagnosis and Therapy, 2020, v.12;No.65(01):121-125.
[3 ] Luo Yanping. The application prospect of mass spectrometry in clinical microbiology laboratory [J]. Laboratory Medicine , 2015(2):97-101.
[4] Tian Yueru, Guan Ming, Li Min. MALDI-TOF MS Challenges of technology in the application of clinical microbiology diagnosis [J]. Chinese Journal of Laboratory Medicine , 2018(8):559-562.
