On line, non-destructive biomass determination of bacterial biofilms by fluorometry

The lack of on-line methodology for the determination of microbial biomass and activity of attached bacteria has severely limited the study of biofilm physiology. This study showed that the fluorescent emission of aromatic amino acids in microbial biofilms can be used to determine the biomass formed on 316 stainless steel coupons. Cells resuspended from the substratum were enumerated by viable and acridine orange counts showing correlation coefficients of 0.77 and 0.98, respectively, when compared to the tryptophane fluorescence. Substrata treated with a fluorescent epoxy coating (F-150) showed no fluorescence that could be attributed to the microorganisms. Bioluminescent emission of an actively growing bioluminescent bacterium, Vibrio harveyi, was correlated with acridine orange counts (r 2 = 0.95) and fluorescence (r 2 = 0.93). The results of these studies suggest that fuorescence measurements can be used to monitor microbial biomass associated with various substrata. Coupled with bioluminescence measurements, this method provides information on both biomass constituents and metabolic activity, and therefore possibly an indicator of sub-lethal toxicity.