Abstract
Correct identification of atypical Brucella spp. and their discrimination against former Ochrobactrum species remains difficult with current routine diagnostic procedures. Matrix-assisted time-of-flight mass spectrometry (MALDI ToF MS) is a fast, reliable, cost-effective and high-throughput technique, increasingly used to identify microorganisms by their proteomic fingerprint. However, successful identification of a specific pathogen is only possible with an accurate reference database that covers the respective genus. Unfortunately, none of the major manufacturers of MALDI ToF MS systems offers databases comprising all currently known Brucella species. To overcome this drawback, various internal pipelines and databases have been established over the last years. Yet, the newly emerging species require an extension of these pipelines due to the lack of reference spectra and biomarkers. We describe the identification of several atypical Brucella strains,including former Ochrobactrum anthropi and Ochrobactrum intermedium, using their MALDI ToF MS spectra by taking advantage of unique, species-specific peak pattern profiles. These specific mass spectral peaks have been identified by comparing spectra of 110 different strains and roughly 2,600 spectra belonging to the six classical and two marine Brucella species, the novel species B. microti, B. inopinata, B. papionis and B. vulpis as well as various atypical Brucella isolates from frogs and rodents. The proteins corresponding to the species-specific peaks were identified using our in-house pipeline combined with data from NCBI. The identification of these species-specific spectral peaks and their subsequent assignment to Brucella biomarkers finally highlights the reliability of our method. In summary, classical and novel Brucella spp. can be quickly and easily identified using the here presented MALDI ToF MS pipeline, avoiding the commonly described misidentifications with former O. anthropi, O. intermedium and atypical Brucella strains. The ability of MALDI ToF MS to discriminate novel atypical Brucella and former Ochrobactrum species from classical Brucella and B. inopinata(-like) is especially important in the context of biological safety regulations.
