The article by Philip S. Stewart discusses the significant role of diffusion in microbial biofilms, explaining how it influences the chemistry and biology of biofilm growth. Diffusion is crucial in biofilms because fluid flow is reduced, increasing the diffusion distance and leading to diffusion limitation. The author addresses five key questions: 1. **Why is diffusion important in biofilms?** Diffusion is critical in biofilms due to reduced fluid flow and increased diffusion distance, which can be orders of magnitude greater than in free aqueous suspensions. This limits the transport of solutes and nutrients. 2. **How fast do solutes diffuse into or out of a biofilm?** The time required for a solute to reach a certain concentration in a biofilm is calculated using the effective diffusion coefficient and the geometry of the biofilm. Examples illustrate the penetration times for different scenarios, such as staining a biofilm or penetration of antimicrobial agents. 3. **Why are biofilm chemistry and biology so spatially heterogeneous?** Local variations in nutrient concentrations and microbial species are common in biofilms, leading to environmental microniches that support diverse species. Chemical gradients created by nutrient depletion influence the distribution of bacteria and their metabolic activities. 4. **Do water channels in biofilms eliminate diffusion limitation?** Water channels can improve solute transport in the immediate lining of the channel but do not guarantee access to the interior of cell clusters, which remain diffusion-limited due to dense bacterial aggregation. 5. **Are there critical biofilm phenomena that diffusion does not explain?** While diffusion limitation explains many phenomena, it may not account for all aspects of biofilm behavior, such as sensing pathways and gene expression triggered by nutrient availability or quorum sensing. The article concludes with a summary of four key points about diffusion in biofilms and acknowledges the support from the W. M. Keck Foundation.The article by Philip S. Stewart discusses the significant role of diffusion in microbial biofilms, explaining how it influences the chemistry and biology of biofilm growth. Diffusion is crucial in biofilms because fluid flow is reduced, increasing the diffusion distance and leading to diffusion limitation. The author addresses five key questions: 1. **Why is diffusion important in biofilms?** Diffusion is critical in biofilms due to reduced fluid flow and increased diffusion distance, which can be orders of magnitude greater than in free aqueous suspensions. This limits the transport of solutes and nutrients. 2. **How fast do solutes diffuse into or out of a biofilm?** The time required for a solute to reach a certain concentration in a biofilm is calculated using the effective diffusion coefficient and the geometry of the biofilm. Examples illustrate the penetration times for different scenarios, such as staining a biofilm or penetration of antimicrobial agents. 3. **Why are biofilm chemistry and biology so spatially heterogeneous?** Local variations in nutrient concentrations and microbial species are common in biofilms, leading to environmental microniches that support diverse species. Chemical gradients created by nutrient depletion influence the distribution of bacteria and their metabolic activities. 4. **Do water channels in biofilms eliminate diffusion limitation?** Water channels can improve solute transport in the immediate lining of the channel but do not guarantee access to the interior of cell clusters, which remain diffusion-limited due to dense bacterial aggregation. 5. **Are there critical biofilm phenomena that diffusion does not explain?** While diffusion limitation explains many phenomena, it may not account for all aspects of biofilm behavior, such as sensing pathways and gene expression triggered by nutrient availability or quorum sensing. The article concludes with a summary of four key points about diffusion in biofilms and acknowledges the support from the W. M. Keck Foundation.