Microbiology laboratories where MALDI-TOF MS is available can benefit from its capacity to identify most clinically interesting non-tuberculous mycobacteria in a rapid, reliable, and inexpensive manner.

Introduction: Ten years after its introduction into clinical microbiology, MALDI-TOF mass spectrometry has become the standard routine identification tool for bacteria in most laboratories. The technology has accelerated analyses and improved the quality of results. The greatest significance has been observed for bacteria that were challenging to be identified by traditional methods. Areas covered: We searched in existing literature (Pubmed) for reports how MALDI-TOF MS has contributed to identification of rare and unknown bacteria from different groups. We describe how this has improved the diagnostics in different groups of bacteria. Reference patterns for strains which yet cannot be assigned to a known species even enable the search for related bacteria in studies as well as in routine diagnostics. MALDI-TOF MS can help to discover and investigate new species and their clinical relevance. It is a powerful tool in the elucidation of the bacterial composition of complex microbiota in culturomics studies. Expert opinion: MALDI-TOF MS has improved the diagnosis of bacterial infections. It also enables knowledge generation for prospective diagnostics. The term 'hard-to-identify' might only be rarely attributed to bacteria in the future. Novel applications are being developed, e.g. subspecies differentiation, typing, and antibiotic resistance testing which may further contribute to improved microbial diagnostics.

We have developed a model for the accurate identification of M. chimaera and M. intracellulare by MALDI-TOF MS. This approach has the potential for routine use in microbiology laboratories, as the model itself can be easily implemented into the software of the currently available systems by MALDI-TOF MS manufacturers.

For the first time, GIM-1 has been demonstrated in A. pittii. This resistance mechanism has previously been reported only in Enterobacteriaceae and Pseudomonas aeruginosa. As GIM-1 was found in strains with diverse clonal backgrounds, but encoded on plasmids of a similar size, further spread among Acinetobacter spp. seems possible. The detection of GIM-1 production might be challenging in some strains due to the low MICs of carbapenems.

The ability of MALDI-TOF for the identification of nontuberculous mycobacteria (NTM) has improved recently thanks to updated databases and optimized protein extraction procedures. Few multicentre studies on the reproducibility of MALDI-TOF have been performed so far, none on mycobacteria. The aim of this study was to evaluate the reproducibility of MALDI-TOF for the identification of NTM in 15 laboratories in 9 European countries. A total of 98 NTM clinical isolates were grown on Löwenstein-Jensen. Biomass was collected in tubes with water and ethanol, anonymized and sent out to the 15 participating laboratories. Isolates were identified using MALDI Biotyper (Bruker Daltonics). Up to 1330 MALDI-TOF identifications were collected in the study. A score ≥ 1.6 was obtained for 100% of isolates in 5 laboratories (68.2–98.6% in the other). Species-level identification provided by MALDI-TOF was 100% correct in 8 centres and 100% correct to complex-level in 12 laboratories. In most cases, the misidentifications obtained were associated with closely related species. The variability observed for a few isolates could be due to variations in the protein extraction procedure or to MALDI-TOF system status in each centre. In conclusion, MALDI-TOF showed to be a highly reproducible method and suitable for its implementation for NTM identification.