Optical biosensor for environmental on-line monitoring of naphthalene and salicylate bioavailability with an immobilized bioluminescent catabolic reporter bacterium

Abstract

An optical whole-cell biosensor based on a genetically engineered bioluminescent catabolic reporter bacterium was developed for continuous on-line monitoring of naphthalene and salicylate bioavailability and microbial catabolic activity potential in waste streams. The bioluminescent reporter bacterium, Pseudomonas fluorescens HK44, carries a transcriptional nahG-"uxCDABE fusion for naphthalene and salicylate catabolism. Exposure to either compound resulted in inducible bioluminescence. The reporter culture was immobilized onto the surface of an optical light guide by using strontium alginate. This biosensor probe was then inserted into a measurement cell which simultaneously received the waste stream solution and a maintenance medium. Exposure under defined conditions to both naphthalene and salicylate resulted in a rapid increase in bioluminescence. The magnitude of the response and the response time were concentration dependent. Good

A new green fluorescent protein‐based bacterial biosensor for analysing phenanthrene fluxes

“…The reasons for the absence of a detectable GFP signal in EFM even at nominally saturated aqueous phenanthrene concentrations are not immediately clear. In comparison, naphthalene biosensors based on Pseudomonas putida and the nahG promoter have a lower detection limit for naphthalene of ∼0.5 µM (Heitzer et al, 1994;Werlen et al, 2004), although these were equipped with the luxAB bacterial luciferase as reporter protein and the detection limit was derived from a global response of a larger population of cells simultaneously. Actually most whole-cell living bioreporters responsive to organic compounds start giving off measurable reporter signals above 100 nM target compound concentration, but then again this is usually based on a 'global' measurement of cells in aqueous suspension, i.e.…”

“…However, for most PAHs, the bioavailability for microbial degradation is in most realistic cases not appreciable thus presenting an interesting test-case to apply a bacterial biosensor. For PAHs, bacterial biosensors have only been constructed for naphthalene (Heitzer et al ., 1994;Dorn et al ., 2003;Werlen et al ., 2004) and for fluorene (Bastiaens et al ., 2001), although toxicity-responsive bacterial biosensors have been used for the (unspecific) detection of PAHs with higher molecular weight and lower solubility, such as phenanthrene and anthracene (Lee et al ., 2003).…”

Fluorescence‐based sensing system for copper using genetically engineered living yeast cells

“…These sensing systems employ genetically modified whole cells containing novel genetic elements, such that a regulatory protein controls transcription of a reporter gene that subsequently leads to a detectable signal. Sensing systems that use this approach have been employed for detection of metal ions, organics, and other global parameters (Applegate et al, 1998;Corbisier et al, 1999;Guan et al, 2000;Heitzer et al, 1994;Kobatake et al, 1995;Layton et al, 1998;Peitzsch et al, 1998;Ramanathan et al, 1997Ramanathan et al, , 1998Shetty et al, 1999;Sticher et al, 1997;Tauriainen et al, 1998;Virta et al, 1995;Miller, 1998, 1999). In this work, we developed a fluorescence-based sensing system for copper employing genetically designed yeast.…”

SMET: Systematic multiple enzyme targeting – a method to rationally design optimal strains for target chemical overproduction

“…Since the inception of metabolic engineering more than two decades ago [1–3], the field has played a significant role in optimizing microbial biocatalysts and has had a significant impact on biotechnological applications related to health [4, 5], food [2, 6], energy [7, 8], and environment [9, 10]. One of the key questions that metabolic engineers face is to identify which genes should be targeted to develop a robust and efficient strain to achieve desirable phenotypes, e.g.…”