Due to the physicochemical properties of nanoparticles, the use of nanomaterials increases over time in industrial and medical processes. We herein report the negative impact of nanoparticles, using solid growth conditions mimicking a biofilm, on the ability ofBacillus subtilisto fight against a stress. Bacteria have been exposed to sublethal doses of nanoparticles corresponding to conditions that bacteria may meet in their natural biotopes, the upper layer of soil or the gut microbiome. The analysis of the proteomic data obtained by shotgun mass spectrometry have shown that several metabolic pathways are affected in response to nanoparticles, n-ZnO or n-TiO2, or zinc salt: the methyglyoxal and thiol metabolisms, the oxidative stress and the stringent responses. Nanoparticles being embedded in the agar medium, these impacts are the consequence of a physiological adaptation rather than a physical cell injury. Overall, these results show that nanoparticles, by altering bacterial physiology and especially the ability to resist to a stress, may have profound influences on a "good bacteria",Bacillus subtilis, in its natural biotope and moreover, on the global equilibrium of this biotope.