Pseudomonas fluorescens HK44: lessons learned from a model whole-cell bioreporter with a broad application history

Abstract

Initially described in 1990, Pseudomonas fluorescens HK44 served as the first whole-cell bioreporter genetically endowed with a bioluminescent (luxCDABE) phenotype directly linked to a catabolic (naphthalene degradative) pathway. HK44 was the first genetically engineered microorganism to be released in the field to monitor bioremediation potential. Subsequent to that release, strain HK44 had been introduced into other solids (soils, sands), liquid (water, wastewater), and volatile environments. In these matrices, it has functioned as one of the best characterized chemically-responsive environmental bioreporters and as a model organism for understanding bacterial colonization and transport, cell immobilization strategies, and the kinetics of cellular bioluminescent emission. This review summarizes the characteristics of P. fluorescens HK44 and the extensive range of its applications with special focus on the monitoring of bioremediation processes and biosensing of environmental pollution.

Keywords: Pseudomonas fluorescens HK44; bioluminescence; bioreporter; biosensors; lux genes.

Figure 1.

Transmission electron micrograph of Pseudomonas fluorescens HK44 encapsulated in a silica gel (reprinted from [38] with permission).

Figure 2.

Bioluminescence emission from P. fluorescens HK44 in a flowcell exposed to cyclic perturbations of naphthalene. Adapted from [39].

Figure 3.

Plasmid pUTK21 contains a transposon-based luxCDABE insert positioned within the nahG gene. This permits direct observation of naphthalene catabolic activity via emission and real-time measurement of bioluminescence. Tc^r, tetracycline resistance gene; Tnp, transposase.

Figure 4.

Physical organization of the gene clusters in three plasmids pDTG1, pNAH20, and pUTK21 belonging to Pseudomonas strains. Genes of the same color indicate corresponding orthologous genes with high homology (>80%) at the nucleic acid level and arrows indicate direction of transcription (Adapted from [45]).

Figure 5.

Effect of methanol on bioluminescence emission from non-induced P. fluorescens HK44. Values are expressed relative to a control not exposed to methanol. Reprinted from [38] with permission.

Figure 6.

Scheme of the lysimeter facility used in long-term field biodegradation experiment (A) The lysimeter facility consisted of six replicate and control soil ecosystems; (B) Inside view of one of the six soil packed lysimeters; (C) Representative schematic of one of the 4 m deep × 2.5 m diameter lysimeters used for the release of P. fluorescens HK44.