Biofilm Characterization by AFM and SEM and Growth Kinetics of Geobacter sulfurreducens in Regional Cheese Whey

Geobacter sulfurreducens is a model bacterium widely used in microbial fuel cell (MFC) research due to its efficient extracellular electron transfer. However, the high cost of synthetic media limits the scalability of these systems, making agro-industrial byproducts like cheese whey a sustainable alternative. This study evaluated cheese whey as a growth medium for G. sulfurreducens and its influence on biofilm development on graphite bars electrodes. Bacterial growth kinetics and biofilm architecture were characterized using Atomic Force Microscopy (AFM) as the primary quantitative tool, supplemented by Scanning Electron Microscopy (SEM). Growth curves revealed a diauxic-like transition within the first 48 h, with high cell viability (94%). AFM analysis demonstrated a non-linear topographical evolution: an initial attachment phase was followed by a peak in structural heterogeneity at 14 days (Sq = 683.08 nm), eventually reaching a mature, confluent state at 21 days with a maximum thickness of ~8 μm. Energy-Dispersive Spectroscopy (EDS) confirmed an organic and mineral matrix consistent with bacterial biomass and whey components. These results demonstrate that cheese whey effectively supports the growth of G. sulfurreducens and the formation of structurally complex biofilms, highlighting its potential as a low-cost substrate for microbial cultivation and dairy waste valorization.