Foodborne diseases are important to determine bacteria in strain level, which are analyzed by library‐based devices and bioinformatics‐enabled. The aim of this study was to investigate the contribution of principal component analysis (PCA) with matrix‐assisted laser desorption ionization time‐of‐flight mass spectrometry (MALDI‐TOF‐MS) to distinguish according to the differences of bacterial strains as rapidly screening of foodborne bacteria. The MALDI‐TOF‐MS‐based PCA analysis was used for differentiating bacterial strains isolated from ready‐to‐eat foods. According to the results of PCA analysis, the percentages of distance and proximity between species were evaluated by composite correlation indexes (CCI). Bacillus cereus were detected in burghul salad (BC1) and macaroni salad (BC2) taken from the SB2‐snack bar, and the similarity rate was determined as 97%. Three other B. cereus bacteria (BC3, BC4, and BC5) in the same cluster were also isolated from salads collected from SB4‐snack bar. The similarity of Klebsiella pneumoniae bacteria, which have the codes KP1 and KP2, isolated from macaroni salad and burghul salad taken from the SB2 snack bar respectively were 96%. Additionally, the CCI values of two E. coli strains in burghul (EC1) salad and Russian salad (EC2) in the same sampling point (SB1) were determined as 97%. In conclusion, this analysis with MALDI‐TOF‐MS based PCA has revealed the relationships between bacteria genera and species, beyond just the identification of bacteria and the rapid screening of bacteria in perishable foodstuffs. Similarities between bacterial strains identified for different samples from the same sampling point suggested that there were not adequate hygiene rules and storage requirements were not followed.