Biotyper version 3.0 software

Samples were aseptically removed from containers for subsequent extraction by grinding in sterile phosphate-buffered saline (pH 7.4) at a 1:10 ratio. Then the extracts were inoculated into a liquid MRS medium. Inoculates were incubated at 37 °C for 24 hours, then serial tenfold dilutions in sterile PBS (pH 7.4) were made. Each dilution was plated on MRS agar medium (6.4), and the cultures were incubated in a microaerophilic chamber at 37 °C for 24 hours. The selection of colonies for further research was carried out based on colony morphology, microscopy of smears with Gram stain, and catalase activity.
Pure overnight bacterial cultures were used for mass spectrometric biotyping. Sample preparation for the mass MALDI-TOF spectrometry was carried out by the direct deposition method. The study was performed on a Microflex LT instrument (Bruker Daltonics GmbH, Leipzig, Germany) using Biotyper (version 3.0) software (Bruker Daltonics GmbH, Leipzig, Germany).

The mass spectra were baseline-corrected and smoothing-processed with Biotyper version 3.0 software (Bruker Daltonics, Bremen, Germany). Mass spectral patterns (MSPs) were created from the spectra that were acquired multiple times for each strain by selecting the main peaks and removing very minor peaks and noise using the MALDI Biotyper. An in-house MSP database of wood-rotting fungi was constructed. The identification scores of the newly acquired mass spectra of the MSPs in the database were calculated by the MALDI Biotyper to evaluate the reproducibility. The scores were presented between 0 and 3: a score ≥ 2.0 was accepted for a reliable identification to the species level; a score ≥ 1.7 and < 2.0 was accepted for an identification to the genus level; and a score < 1.7 indicated an unreliable identification of the microorganisms [29 (link),30 (link)].