Rapid, Sensitive Bioluminescent Reporter Technology for Naphthalene Exposure and Biodegradation

TLDR

A bioluminescent reporter plasmid (pUTK21) was created by inserting a Vibrio fischeri lux cassette into the naphthalene catabolic plasmid of Pseudomonas fluorescens via Tn4431, targeting the nahG salicylate hydroxylase gene. The pUTK21 strains produced rapid, highly responsive bioluminescence upon exposure to naphthalene or salicylate, enabling detection in continuous culture and contaminated soils and suggesting suitability for online environmental monitoring.

Abstract

A bioluminescent reporter plasmid for naphthalene catabolism (pUTK21) was developed by transposon (Tn4431) insertion of the lux gene cassette from Vibrio fischeri into a naphthalene catabolic plasmid in Pseudomonas fluorescens. The insertion site of the lux transposon was the nahG gene encoding for salicylate hydroxylase. Luciferasemediated light production from P. fluorescens strains harboring this plasmid was induced on exposure to naphthalene or the regulatory inducer metabolite, salicylate. In continuous culture, light induction was rapid (15 minutes) and was highly responsive to dynamic changes in naphthalene exposure. Strains harboring pUTK21 were responsive to aromatic hydrocarbon contamination in Manufactured Gas Plant soils and produced sufficient light to serve as biosensors of naphthalene exposure and reporters of naphthalene biodegradative activity. The robust and sensitive nature of the bioluminescent reporter technology suggests that new sensing methods can be developed for on-line process monitoring and control in complex environmental matrices.