The use of time-of-flight mass spectrometry for diagnosis of brucellosis and interspecific differentiation of strains of Brucella spp.

• Ulshina D.V.
• Kovalev D.A.
• Tatyana N. Bobrysheva
• Dmitry G. Ponomarenko
• Rusanova D.V.
• Kovaleva N.I.
• Alexander N. Kulichenko

Abstract

The aim is to determine the possibility of using time-of-flight mass spectrometry with matrix-assisted laser desorption/ionization (MALDI-TOF MS) for the diagnosis of acute brucellosis based on the analysis of the protein profiles of patients with human brucellosis and to evaluate the effectiveness of using MALDI-TOF MS for interspecific differentiation of strains of the brucellosis microbe.

Material and methods. The paper presents the results of a study of the blood of patients with a clinical diagnosis of acute brucellosis (n=18) and conditionally healthy people (n=30) using time-of-flight mass spectrometry. The protein profiles of 69 strains of 6 brucella species (B. melitensis, B. abortus, B. suis, B. canis, B. ovis, B. neotomae) have been studied.

Bacteriological, mass-spectrometric, bioinformation, and statistical methods were used in the work.

Results. A method for decontamination and sample preparation of blood is proposed for subsequent analysis using MALDI-TOF MS. As a result of the comparative analysis of the protein profiles of the samples of patients with human brucellosis, 27 common signals were characterized, including 7 specific for Brucella (m/z±5 Da): 2422, 3268, 3336, 3696, 5360, 6672, 7048, which discriminate them from the mass spectra of conditionally healthy people.

69 representative protein profiles of representatives of 6 brucella species were studied, which are included in the electronic database of mass spectra in the environment of the Biotyper v 3.0 program (Bruker Daltonics, Germany). The effectiveness of multidimensional scaling and the RGL package in the R language environment for the interspecies differentiation of strains of the brucellosis pathogen on the basis of mass spectrometric data is shown.

Conclusion. The possibility of identifying specific markers of the causative agent of brucellosis in the blood of patients with a clinical diagnosis of acute brucellosis by the MALDI-TOF MS method without the stage of isolation of pure culture or accumulation of pathogens in the sample at the stage of sample preparation was confirmed. The perspectives of application of applied packages in the environment of the R language for bioinformational processing of time-of-flight mass spectrometry data of protein extracts of cultures for the purpose of interspecies differentiation of strains of brucellosis causative agent of clinical significance are demonstrated.

Keywords:brucellosis, laboratory diagnostics, mass spectrometry, protein profiling, species differentiation

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