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Non-tuberculous mycobacteria (NTM) are difficult to identify by biochemical and genetic methods due to their microbiological properties and complex taxonomy. The development of more efficient and rapid methods for species identification in the veterinary microbiological laboratory is, therefore, of great importance. Although MALDI-TOF Mass Spectrometry (MS) has become a promising tool for the identification of NTM species in human clinical practise, information regarding its performance on veterinary isolates is scarce. This study assesses the capacity of MALDI-TOF MS to identify NTM isolates ( n  = 75) obtained from different animal species. MALDI-TOF MS identified 76.0% ( n  = 57) and 4% ( n  = 3) of the isolates with high and low confidence, respectively, in agreement with the identification achieved by Sanger sequencing of housekeeping genes (16S rRNA,  hsp65 , and  rpoB ). Thirteen isolates (17.3%) were identified by Sanger sequencing to the complex level, indicating that these may belong to uncharacterised species. MALDI-TOF MS approximated low confidence identifications toward closely related mycobacterial groups, such as the  M. avium  or  M. terrae  complexes. Two isolates were misidentified due to a high similarity between species or due to the lack of spectra in the database. Our results suggest that MALDI-TOF MS can be used as an effective alternative for rapid screening of mycobacterial isolates in the veterinary laboratory and potentially for the detection of new NTM species. In turn, Sanger sequencing could be implemented as an additional method to improve identifications in species for which MALDI-TOF MS identification is limited or for further characterisation of NTM species.

MALDI-TOF Mass Spectrometry as a Rapid Screening Alternative for Non-tuberculous Mycobacterial Species Identification in the Veterinary Laboratory