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This study proposes to exploit the in vivo metabolism of two probiotics (<i>Bifidobacterium longum</i> subsp. <i>infantis</i> and <i>Lactobacillus reuteri</i>) which, upon adhesion on a solid surface, form a biofilm able to control the growth of pathogenic and food spoilage bacteria. The results showed that pathogenic cell loads were always lower in presence of biofilm (6.5&#8722;7 log CFU/cm²) compared to those observed in its absence. For <i>Escherichia coli</i> O157:H7, a significant decrease (&gt;1&#8722;2 logarithmic cycles) was recorded; for <i>Listeria monocytogenes</i>, <i>Staphylococcus</i> <i>aureus,</i> and <i>Salmonella enterica</i>, cell load reductions ranged from 0.5 to 1.5 logarithmic cycles. When tested as active packaging, the biofilm was successfully formed on polypropylene, polyvinyl chloride, greaseproof paper, polyethylene and ceramic; the sessile cellular load ranged from 5.77 log CFU/cm² (grease-proof paper) to 6.94 log CFU/cm² (polyethylene, PE). To test the potential for controlling the growth of spoilage microorganisms in food, soft cheeses were produced, inoculated with <i>L.</i> <i>monocytogenes</i> and <i>Pseudomonas fluorescens,</i> wrapped in PE pellicles with pre-formed biofim, packed both in air and under vacuum, and stored at 4 and 15 &#176;C: an effective effect of biofilms in slowing the decay of the microbiological quality was recorded.