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Growing consumer demand for high-quality products coupled with minimally processed products and a minor use of synthetic food additives have increased the need to search for new sources of natural antimicrobials to ensure product safety. This study aimed to evaluate the antimicrobial activity of extracts from the brown algae Ericaria selaginoides against Bacillus cereus in typical Catalan fresh cheese (“mató”) by means of challenge testing. Three concentrations of a crude extract and its corresponding two subfractions (non-polar and mid-polar) obtained after purification showed an antimicrobial dose-dependent effect on B. cereus, from inhibition to inactivation. The best results were obtained with higher concentrations of the non-polar subfraction that caused a total inactivation of the inoculated pathogen after 2 or 4 days, followed by the mid-polar that inactivated B. cereus after 2 or 6 days. The results showed an improvement in the antimicrobial effect after purification compared with the effect observed when the crude extract was tested. Moreover, compounds of different chemical natures may be involved in this antimicrobial activity since it remained in both subfractions after purification. The results obtained in this work show the great potential of macroalgae extracts as natural food preservatives against B. cereus in fresh cheese.