Bacterial Identification via MALDI-TOF MS

Initially, the positive blood cultures were subjected to Gram staining to determine the presence of gram-positive or gram-negative bacteria. Based on these results, appropriate agar plates, including blood, Maconkey, chocolate, and anaerobic blood agar, were used for further culturing. The plates were grown in an incubator (Thermo Fisher Scientific, USA) at 35°C in a 5% CO₂ or anaerobic atmosphere until visible colonies appeared. Identification was then performed with the Bruker microflex MALDI-TOF MS system using the MALDI Biotyper 3.0 Realtime classification (RTC) database (Bruker Daltonics, Bremen, Germany) as described previously (Chen et al., 2013 (link); Tian et al., 2016 (link); Zhou et al., 2017 (link)). In brief, a pure bacterial colony was smeared onto a steel target plate with a wood toothpick, and 70% formic acid solution was added to lyse the bacterial cells. Once the formic acid solution dried, 1 μL of α-cyano-4-hydroxycinnamic acid (HCCA) matrix (Bruker Daltonics) solution was added for subsequent MALDI-TOF MS (Bruker Daltonics) identification. The calibration and validation of MALDI-TOF MS was carried out once a week with a bacterial test standard (BTS) according to the manufacturer's instructions (Bruker Daltonics).