Technical Abstract: This work presents the development of a method for rapid bacterial identification based on the fluorescence spectroscopy combined with multivariate analysis. Fluorescence spectra of pure three different genera of bacteria (Escherichia coli, Salmonella, and Campylobacter) were collected from 200 to 700 nm. To find the optimum excitation and emission wavelengths for each bacteria sample, synchronous scan technique was employed and resulted in two maximum Ex at 225 nm and 280 nm and one maximum Em at 335-345 nm for all three bacteria. Due to the similarity in spectral pattern among three bacteria, it was difficult to classify them based on the raw spectra, however, the application of principal component analysis (PCA) to the spectral data resulted in successful classification of the bacteria by their genus as well as concentration. Detecting salmonella on the surface of shell eggs was affected by the contaminants such as feces and cuticle, thus it was difficult to directly assay salmonella on the surface of shell eggs. However, distinguishing between unwashed eggs and washed eggs was successfully performed using PCA. The results demonstrated that fluorescence spectroscopy, when coupled with chemometrics, has the potential to detect and to differentiate bacterial pathogens. This method is rapid and easy to perform compared to standard methods, and requires minimum sample preparation.