Specific and Quantitative Assessment of Naphthalene and Salicylate Bioavailability by Using a Bioluminescent Catabolic Reporter Bacterium

Abstract

A bioassay was developed and standardized for the rapid, specific, and quantitative assessment of naphthalene and salicylate bioavailability by use of bioluminescence monitoring of catabolic gene expression. The bioluminescent reporter strain Pseudomonas fluorescens HK44, which carries a transcriptional nahG-luxCDABE fusion for naphthalene and salicylate catabolism, was used. The physiological state of the reporter cultures as well as the intrinsic regulatory properties of the naphthalene degradation operon must be taken into account to obtain a high specificity at low target substrate concentrations. Experiments have shown that the use of exponentially growing reporter cultures has advantages over the use of carbon-starved, resting cultures. In aqueous solutions for both substrates, naphthalene and salicylate, linear relationships between initial substrate concentration and bioluminescence response were found over concentration ranges of 1 to 2 orders of magnitude. Naphthalene could be detected at a concentration of 45 ppb. Studies conducted under defined conditions with extracts and slurries of experimentally contaminated sterile soils and identical uncontaminated soil controls demonstrated that this method can be used for specific and quantitative estimations of target pollutant presence and bioavailability in soil extracts and for specific and qualitative estimations of napthalene in soil slurries.

Non-exhaustive extraction techniques (NEETs) for the prediction of naphthalene mineralisation in soil

“…To evaluate this, a catabolic luminescent bacterial biosensor was used. The bacterial biosensor Pseudomonas fluorescens HK44 pUTK21 responds sensitively and quantitatively to naphthalene [10] and has been used previously in the assessment of contaminated soils [11]. If the sensor induction is correlated to the concentration of naphthalene in soil solution, it may be interpreted that there is no constraining factor to the biodegradation.…”

Kinetic analysis of bacterial bioluminescence

“…Sensors using biological components are increasingly utilized in the analyses of biological processes and environmental conditions. Bioluminescent whole cell sensors are a promising type of bioreporter that have been developed for assaying gene expression (Stewart and Williams, 1992;Steinberg et al, 1995), metabolic activity (Unge et al, 1999;Porteous et al, 2000), viability (Griffiths et al, 2000), bioavailability (Heitzer et al, 1992;Tibazarwa et al, 2001), spatial distribution of organisms in the environment (Ramos et al, 2000), contaminant detection (Hay et al, 2000), and toxicity assessment (Kelly et al, 1999;Choi and Gu, 2001). Bacterial bioluminescent bioreporters are constructed by cloning the genes (lux) that encode for enzymes that catalyze the bioluminescent reactions into a specific host.…”

Measuring mass transfer processes of octane with the help of an alkSalkB::gfp‐tagged Escherichia coli

“…1 nM Hg 21 Selifonova et al, 1993), but not for organic compounds [e.g. 0.3 mM naphthalene (Heitzer et al, 1992) or 1 mM toluene (Applegate et al, 1997)]. This sensitivity appears to be erratic, as one would expect that the response range is closer to the range of octane concentrations actually possible in aqueous systems, i.e.…”