Food safety is a major concern for suppliers in the food chain to ensure the safety of their products. The identification procedure requested by norms is tedious, and it often requires systematic controls and qualified staff to perform the necessary analyses. Raman spectroscopy offers new opportunities to rapidly and efficiently ascertain the presence of pathogens in samples. Nevertheless, this technique requires a standardized procedure to be applied in the industrial context. Our study shows that the variability between spectral fingerprints is related to the physiological state of the microbial species and the growth phase of the bacteria plays a crucial role in its identification by Raman spectroscopy. To improve the discrimination between closely related bacterial species, a procedure based on the selection of bacterial spectra in the exponential growth phase was proposed. Different ways to introduce Raman spectroscopy in the ISO 6579:2002 standards are also proposed from the entire process to a shorter protocol. In the latter case, the identification of bacterial colonies after the selective enrichment step was proposed with the advantages of this path in terms of simplicity and rapidity (analysis time is reduced up to 50 h from the 100 h required by the standard). The protocol validated using six food categories from industrial partners have presented a good correlation by confirmation with other laboratory classical methods. In the future, this procedure could be introduced to the control system of the food production chain with a reliable database for various microorganisms encountered in this field.