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MALDI in microbiology



Until recently, clinical diagnostic laboratories relied predominantly on conventional phenotypic pathways for the diagnosis of infections and sometimes on gene sequencing techniques. The latest advances in technology include matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry, which has entered daily microbiological practice.

MALDI-TOF mass spectrometry generates specific mass spectral fingerprints that can be considered to be unique signatures of microorganisms that can help in their precise identification to the genus and species levels – with great potential for strain typing.

Bacterial identification

Multiple approaches to using MALDI TOF mass spectrometry is now used in microbiological diagnostic laboratories. One approach compares results with databases of commercially available mass spectrometry signatures to identify the sample bacteria, and a second uses a proteome database to identify biomarker masses in the bacteria from sequenced genomes. The second bioinformatics approach allows for variations in culture growth and sample treatment conditions, while the first is particularly useful in routine laboratory methods such as diagnostics and can distinguish between species and subspecies.

Given its accuracy, the technology can also be applied directly to various clinical samples, especially blood, cerebrospinal fluid, urine, pleural fluid, and peritoneal fluid. The main inhibition is the amount of bacteria present in the samples due to the detection limit of the current MALDI-TOF protocols. To overcome this disadvantage, large volumes of blood and urine samples are normally needed and cultures used as additional blood accumulation.

Results observed for bacterial identification with MALDI-TOF mass spectrometry using either of the two approaches mentioned above, the diagnostic yield and accuracy are highly dependent on the bacterial taxonomy and the quality of the databases used.

One of the key benefits of using MALDI-TOF technology to identify bacteria is the rapid availability of results, typically in less than a second of an hour. In addition, MALDI-TOF mass spectrometry allows the accurate identification of a wide variety of bacteria with rare phenotypic traits that required 16S rRNA gene sequencing prior to the MALDI-TOF era.

19659004] MALDI-TOF mass spectrometry has been rapidly and successfully adapted to the identification of fungi. At present, this method is mainly used for the routine identification of yeasts, but further development is needed (especially in sample preparation protocols and database libraries) to use this identification approach for other groups of fungi (such as dermatophytes and filamentous fungi).

Similar to bacteria, the misidentification or non-identification of fungal species and species by MALDI-TOF mass spectrometry is largely due to errors, absences, or incomplete reference spectra in databases. The disadvantage is that reference spectra currently contained in databases of commercially available MALDI-TOF mass spectrometry systems are incomplete.

In addition, the spectral signal for filamentous fungi depends on the fungal phenotype – the basidiospores, fruiting bodies, surface mycelium and substrate include mycelium. When vegetative mycelium is grown on agar plates, it also exhibits multiple zones corresponding to different ages or stages of development. This can lead to misidentifications and thus to different results in the same sample. For this reason, it is important that databases contain a comprehensive database of multiple MS fingerprints from different stages of development of filamentous fungi to ensure proper identification.

Accuracy, Time, and Cost Efficiency

19659004] Compared to conventional methods for microbial identification, MALDI-TOF mass spectrometry provides in the majority of cases a substantial gain in both engineer / technician working time (sample preparation) also the processing time (obtaining the results with automated analytical method) [19659002] However, the purchase of a MALDI-TOF mass spectrometer is undoubtedly one of the most expensive investments for the clinical microbiology laboratory. This means that reasonable cost justifications and cost analyzes should be sought which should include mandatory quality control steps.

Applications

MALDI-TOF MS is used in a variety of industries, including biopharmaceuticals, organic chemistry, metabolomics and genomics, as well as clinical and diagnostic and therapeutic applications. In organic chemistry, MALDI-TOF is used to analyze nucleic acid, protein and polymer masses as well as to identify complex mixtures of oligonucleotides and small proteins to provide biochemical and chemical researchers with useful information. MALDI-TOF MS plays a crucial role in improving the rapid diagnosis of patients and improves health outcomes. A good example of this is the routine classification of microorganisms in patient samples for clinical microbiology. Conclusion

In conclusion, MALDI-TOF mass spectrometry is a fascinating new technology for microbial identification that is fast, efficient, cost effective, and easy to use. Soon, this instrument will be widely used in diagnostic laboratories because, despite significant instrument cost and maintenance, consumables and running costs are much lower than traditional methods.

Rev. Chloe Barnett, BSc [19659012] Further reading


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