Results within seconds

Fig. 1

Scanning the barcode digitally captures the sample.

Source: bioMérieux company

Enterococcus faecium

Staphylococcus aureus

Pseudomonas aeruginosa

In MALDI TOF mass spectrometry (Matrix-assisted Laser / Desorption Ionization Time Of Flight), small amounts of a sample are introduced into a low-molecular-weight matrix. Laser irradiation and absorption of the laser radiation enable the detection of ribosomal proteins and their representation as a "fingerprint." The resulting mass spectra are automatically evaluated by the database, and the results are listed.

The MALDI TOF method offers several practical advantages over conventional biochemical or molecular biological methods for identification. The primary factor here is time: after inputting the prepared sample into the device, the desired results are available within seconds. Since a preliminary suspicion diagnosis regarding the microorganism identification is not necessary, the process can be carried out even more quickly. The sample application is the same for most microorganisms, only the identification of yeasts and molds is more laborious. With MALDI TOF, regardless of incubation time and nutrient medium from which the sample material is taken, equally informative and accurate results can be achieved.

Operational Hygiene Identifications

The MALDI TOF mass spectrometer (Company bioMerieux, VITEK MS) is validated for identifying microorganisms from pure cultures incubated for 24 to 72 hours. However, it is also conceivable that, due to the advantages of the method, identifications can be performed directly from the original product. The focus here is on cultures on solid nutrient media such as imprint and sedimentation plates, as well as media from airborne microbial counts and broths.

The goal is to obtain at least a rough identification in as short a time as possible. Especially when evaluating samples from hygiene monitoring, this can save time and costs regarding the production process.

Prompt processing of hygiene samples generally leads to reduced downtime of production facilities. Furthermore, targeted investigation of causes can be conducted when action limits are reached, which may influence cleaning processes. Faster results of identification allow for timely release of the investigation subject, resulting in lower or no storage costs. Additionally, cleaning validations can be completed more quickly, enabling faster clearance of rooms.

Project: Direct Identification from Original Samples

In a project, microorganisms present on original samples from hygiene monitoring (imprint plates and nutrient media for airborne microbial counts) were used for identification with VITEK MS without further preparation. The results of the molecular method were compared with those of the parallel biochemical identification and summarized in a table (see Fig. 1).

82 percent of the results matched those of traditional biochemical methods. In 18 percent of the measurements, no identification result was provided. The results reflect the existing experience with MALDI TOF MS.

Staphylococci and micrococci—as well as Pseudomonads and Enterococci—are identified with good confidence values, provided that the respective microorganism is stored in the used database. However, identification of some bacteria still poses difficulties, for example due to their specific growth characteristics.

For example, if they form slimy, viscous colonies (Bacillus, Lactobacillus) or only show very fine microbial growth (coryneform bacteria, Listeria), too few masses are detected. The device then does not provide a result.

Mixed cultures are not recognized as such, so a fresh subculture on a complete medium is indispensable. This is also needed for the final plausibility check. For this reason, direct identification from a broth is also problematic.

Additional conditions must also be met. For example, a microbial count of at least 10^4 CFU/ml is necessary to use the pellet obtained by centrifugation for processing. There should be no other components, such as gels or solids, that could interfere with the measurement.

It is generally possible to perform direct identifications from original samples, especially for cultures on solid nutrient media.