Identification of Brucella spp. isolates and discrimination from the vaccine strain Rev.1 by MALDI-TOF mass spectrometry.

Brucellosis' surveillance and control programs require robust laboratory techniques that can reliably identify and biotype Brucella strains and discriminate between vaccine and field infection. In the recent years, Matrix Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry (MALDI-TOF MS) has revolutionized the routine identification of several microorganisms in clinical microbiology laboratories. Nevertheless, its application on Brucella spp. identification is limited since there are no reference spectra in the commercial databases, due to the microorganism's potential bioterrorist use. In this study, a custom MALDI-TOF MS reference library was constructed and its performance on identification at species level was evaluated using 75 Brucella spp. isolates. Furthermore, distinct peak biomarkers were detected for biovar assignment and discrimination from vaccine strain Rev.1. Analysis of mass peak profiles allowed Brucella accurate identification at genus and species level (100%) with no misidentifications. Despite the high intrageneric similarity, MALDI-TOF MS database succeeded in classifying at biovar level, 47 out of 62 B. melitensis bv. 3 isolates (75.81%), whereas all B. melitensis strains, except for one, were correctly discriminated from vaccine strain Rev.1. MALDI-TOF MS appeared to be a rapid, cost-effective and reliable method for the routine identification of brucellae which reduces time consumption in pathogen identification and could replace in the near future the current conventional and molecular techniques. Its ability to differentiate vaccine from field infection could facilitate brucellosis' monitoring systems contributing in the effective control of the disease.

Christoforidou S ，Kyritsi M ，Boukouvala E ，Ekateriniadou L ，Zdragas A ，Samouris G ，Hadjichristodoulou C ... -  《-》

A MALDI-TOF MS database with broad genus coverage for species-level identification of Brucella.

Mesureur J ，Arend S ，Cellière B ，Courault P ，Cotte-Pattat PJ ，Totty H ，Deol P ，Mick V ，Girard V ，Touchberry J ，Burrowes V ，Lavigne JP ，O'Callaghan D ，Monnin V ，Keriel A ... -  《PLoS Neglected Tropical Diseases》

Protein Biomarker Identification for the Discrimination of Brucella melitensis Field Isolates From the Brucella melitensis Rev.1 Vaccine Strain by MALDI-TOF MS.

Brucella melitensis Rev.1 is a live attenuated vaccine strain that is widely used to control brucellosis in small ruminants. For successful surveillance and control programs, rapid identification and characterization of Brucella isolates and reliable differentiation of vaccinated and naturally infected animals are essential prerequisites. Although MALDI-TOF MS is increasingly applied in clinical microbiology laboratories for the diagnosis of brucellosis, species or even strain differentiation by this method remains a challenge. To detect biomarkers, which enable to distinguish the B. melitensis Rev.1 vaccine strain from B. melitensis field isolates, we initially searched for unique marker proteins by in silico comparison of the B. melitensis Rev.1 and 16M proteomes. We found 113 protein sequences of B. melitensis 16M that revealed a homologous sequence in the B. melitensis Rev.1 annotation and 17 of these sequences yielded potential biomarker pairs. MALDI-TOF MS spectra of 18 B. melitensis Rev.1 vaccine and 183 Israeli B. melitensis field isolates were subsequently analyzed to validate the identified marker candidates. This approach detected two genus-wide unique biomarkers with properties most similar to the ribosomal proteins L24 and S12. These two proteins clearly discriminated B. melitensis Rev.1 from the closely related B. melitensis 16M and the Israeli B. melitensis field isolates. In addition, we verified their discriminatory power using a set of B. melitensis strains from various origins and of different MLVA types. Based on our results, we propose MALDI-TOF MS profiling as a rapid, cost-effective alternative to the traditional, time-consuming approach to differentiate certain B. melitensis isolates on strain level.

Kornspan D ，Brendebach H ，Hofreuter D ，Mathur S ，Blum SE ，Fleker M ，Bardenstein S ，Al Dahouk S ... -  《Frontiers in Microbiology》

Rapid and safe one-step extraction method for the identification of Brucella strains at genus and species level by MALDI-TOF mass spectrometry.

Sali M ，De Maio F ，Tarantino M ，Garofolo G ，Tittarelli M ，Sacchini L ，Zilli K ，Pasquali P ，Petrucci P ，Marianelli C ，Francia M ，Sanguinetti M ，Adone R ... -  《PLoS One》

Reliable identification at the species level of Brucella isolates with MALDI-TOF-MS.

Lista F ，Reubsaet FA ，De Santis R ，Parchen RR ，de Jong AL ，Kieboom J ，van der Laaken AL ，Voskamp-Visser IA ，Fillo S ，Jansen HJ ，Van der Plas J ，Paauw A ... -  《BMC MICROBIOLOGY》