Rapid Identification of Mycobacterial Whole Cells in Solid and Liquid Culture Media by Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry

Abstract

Mycobacterial identification is based on several methods: conventional biochemical tests that require several weeks for accurate identification, and molecular tools that are now routinely used. However, these techniques are expensive and time-consuming. In this study, an alternative method was developed using matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). This approach allows a characteristic mass spectral fingerprint to be obtained from whole inactivated mycobacterial cells. We engineered a strategy based on specific profiles in order to identify the most clinically relevant species of mycobacteria. To validate the mycobacterial database, a total of 311 strains belonging to 31 distinct species and 4 species complexes grown in Löwenstein-Jensen (LJ) and liquid (mycobacterium growth indicator tube [MGIT]) media were analyzed. No extraction step was required. Correct identifications were obtained for 97% of strains from LJ and 77% from MGIT media. No misidentification was noted. Our results, based on a very simple protocol, suggest that this system may represent a serious alternative for clinical laboratories to identify mycobacterial species.

Advantages of Using Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry as a Rapid Diagnostic Tool for Identification of Yeasts and Mycobacteria in the Clinical Microbiological Laboratory

“…The use of matrixassisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) for the rapid identification of clinical yeast and mycobacterial isolates has been evaluated in some studies (13)(14)(15)(16)(17)(18)(19)(20)(21)(22)(23)(24)(25). However, there has been a limited number of direct comparison studies focusing on the performance of the two MALDI-TOF systems to rapidly identify yeast and mycobacterial clinical isolates (26)(27)(28).…”

Comparison of Heat Inactivation and Cell Disruption Protocols for Identification of Mycobacteria from Solid Culture Media by Use of Vitek Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry

“…he Mycobacterium genus consists of over 100 species of rapidly growing and slow-growing acid-fast bacilli (AFB) (1)(2)(3)(4). Rapid and accurate diagnosis of mycobacteria infections is important to patient care and public health (4).…”

“…Rapid identification (ID) of mycobacteria has proven difficult due in part to their fastidious growth requirements and low growth rate (3, 6). Molecular probes and DNA hybridization are relatively fast and simple but are available only for a limited number of clinically common species (2,3,5,7,8). High-performance liquid chromatography (HPLC) (2, 3, 9) and electrospray ionizationtandem mass spectrometry analysis (2) have recently been used to analyze mycolic acid but are labor-intensive and require technical expertise (1, 2).…”

Identificación bacteriana basada en el espectro de masas de proteínas: Una nueva mirada a la microbiología del siglo XXI

“…Protocolos de extracción más sencillos han sido evaluados por otros grupos eliminando el paso de inactivación por calor y agitación con perlas con resultados satisfactorios 19,20 . Una vez finalizada la extracción, se toma una muestra del sobrenadante la que se deposita sobre la placa; una vez seca se procesa de la misma manera que una muestra de colonia directa.…”