Fiber optic detection of in situ lux reporter gene activity in porous media: system design and performance

Abstract

A luminescence detection system is described that couples a genetically engineered bioluminescent reporter organism and fiber optic technology for monitoring in situ reporter gene activity in porous media under dynamic conditions. The reporter bacterium used was Pseudomonas putida RB1353, which carries plasmids NAH7 and pUTK9 that encode genes for salicylate degradation ( nah) and luminescence ( lux) that are regulated by the same promoter. The system can be used to examine the relationship between microbial activity and the resultant impact on biodegradation and transport of salicylate in porous media. Several batch and column experiments were conducted to analyze the performance of the fiber optic detection system. Batch studies showed that the fiber optic response in liquid cultures was linearly correlated to a calibrated system response using a liquid scintillation counter in the single photon counting mode. In column studies, it was demonstrated that decreases in salicylate and dissolved oxygen concentrations associated with biodegradation were correlated with an increase in luminescence response. The maximum luminescence given off by the reporter organism was linearly related to salicylate concentration. These results indicate that the fiber optic system can be used to monitor microbial activity under dynamic conditions. This system allows rapid, real-time, and non-destructive measurements of luminescence from a specific lux reporter microbial population in porous media.