Formic Acid-Based Direct, On-Plate Testing of Yeast and Corynebacterium Species by Bruker Biotyper Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry

Abstract

An on-plate testing method using formic acid was evaluated on the Bruker Biotyper matrix-assisted laser desorption ionizationtime of flight (MALDI-TOF) mass spectrometry system using 90 yeast and 78 Corynebacterium species isolates, and 95.6 and 81.1% of yeast and 96.1 and 92.3% of Corynebacterium isolates were correctly identified to the genus and species levels, respectively. The on-plate method using formic acid yielded identification percentages similar to those for the conventional but more laborious tube-based extraction. Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry (MS) is a rapid and accurate method for identification of bacteria and fungi (2,3,5,6). This technology employs an ionizing laser to generate isolate-derived spectra, which are compared in real time to a library of reference spectra. Depending on the similarity between the acquired spectra and the library, an identification and a confidence score are generated. Some organisms, such as Gram-negative bacteria, are easily analyzed by directly smearing a colony onto the MS plate. Other types of bacteria and yeasts generally require preparatory tube extraction (2). Using the Bruker Biotyper MALDI-TOF MS system (Billerica, MA) and a preparatory tube extraction method, we have previously shown that most yeast and Corynebacterium species are correctly identified to the genus and species levels (1, 3). The tube extraction process involves immersing the isolate in 70% ethanol in a separate tube and pelleting and drying the organism followed by extraction and spotting onto the MS plate. This is cumbersome and time-consuming for routine clinical use, and it requires additional labeling steps, which may lead to specimen misidentification, compromising patient safety.To overcome these preanalytical processing limitations, we evaluated a rapid, direct on-plate testing method using formic acid for the identification of yeast and Corynebacterium species, and we compared our results to those from our prior studies of the same isolates tested by the preparatory tube extraction method (1, 3). Furthermore, the on-plate extraction method uses smaller volumes of formic acid and fewer laboratory consumables than tube extraction, and it is an overall more environmentally friendly process for isolate preparation.(This material was presented, in part, at the 112th American Society for Microbiology General Meeting, San Francisco, CA, 16 to 19 June 2012.)A total of 90 yeast and 78 Corynebacterium species isolates were cultured as described previously (1, 3) and analyzed using the Bruker Microflex LT/SH Biotyper following direct on-plate extraction. For yeast isolates, on-plate testing was performed by smearing a small amount of the organism from a single colony directly onto a spot on the MALDI-TOF MS steel anchor plate (BigAnchor 96-well plate; Bruker) and overlaying it with 1 l of 70% formic acid (Fluka [SigmaAldrich], St. Louis, MO). The order of addition was reversed (formic acid followed by smearing with organism) for the Co...

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).…”

Evaluation of the Bruker MALDI Biotyper for Identification of Gram-Positive Rods: Development of a Diagnostic Algorithm for the Clinical Laboratory

“…Similar identification rates were reported for Corynebacterium spp. and anaerobic bacteria when on-target formic acid preparation and extraction were compared (19,24). An alternative preparation method with on-target ethanol treatment was described by Farfour et al (25), who analyzed a large collection of GPR using the Andromas system and reported accurate species identification.…”

“…An alternative preparation method with on-target ethanol treatment was described by Farfour et al (25), who analyzed a large collection of GPR using the Andromas system and reported accurate species identification. Few studies have systematically compared different sample preparation methods for the identification of GPR with MALDI-TOF MS (13,24,26).…”

Comparision of matrix-assisted laser desorption ionization-time of flight (MALDI-TOF MS) with API ID 32C and VITEK 2 for the identification of Trichosporon species

“…Mayaların MALDI-TOF MS ile identifikasyonu öncesinde yaklaşık 25 dakika süren bir ekstraksiyon aşaması gerçekleştirilmesi gerekli olmakla birlikte, bazı araştırmacılar buna alternatif olan ve daha kısa süren yöntemler geliştirdiklerini bildirmişlerdir (13)(14)(15)(16)(17)(18)(19) .…”