Establishment of a Rapid Identification Method for Three Common Foodborne Gram-Negative Bacteria Based on Matrix-Assisted Laser Desorption Ionization Time-of-Flight Mass Spectrometry

This study aimed to establish a rapid method for identifying three common foodborne pathogenic Gram-negative bacteria (Vibrio parahaemolyticus, Enterobacter cloacae, and Acinetobacter baumannii). We explored the effects of different pretreatment methods, media, and culture durations on the results of bacterial identification using matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). We optimized the MALDI-TOF/TOF MS (tandem TOF MS) method with respect to culture medium and time. Formic acid-acetonitrile extraction was found to be the most suitable pretreatment method. The single-stage TOF MS spectra showed significant differences in identification scores, the number of characteristic peaks, and morphology among strains grown on different media. The number of major ion peaks decreased with culture time, leading to reduced matching accuracy. The tandem TOF MS spectra showed no significant changes in the characteristic peaks of strains at different culture times, indicating that the tandem TOF MS method was less affected by the external environment, exhibiting higher specificity. The single-stage TOF MS method had high accuracy for identifying strains cultured in specific environments for short durations. However, as the culture environment changed and the culture time increased, the results of single-stage TOF MS became unstable, while those of tandem TOF MS changed little. When they were applied to identify Gram-negative bacteria, compared with single-stage TOF MS, tandem TOF MS gave more stable results with fewer interfering peaks, thereby serving as an effective supplement to the existing microbial identification methods.