Assessment of PFAS toxicity using lux-marked bioluminescent Pseudomonas biosensors

Per- and polyfluoroalkyl substances (PFAS) are persistent fluorinated contaminants of growing environmental concern, yet rapid methods for assessing their biological effects remain limited. This study evaluated lux-marked Pseudomonas putida F1 Tn5 luxCDABE and P. putida 12708 Tn5 luxCDABE as whole-cell biosensors for detecting sublethal PFAS toxicity. Fluoride bioassays confirmed both strains tolerated up to 1,000 mg L-1 fluoride without measurable bioluminescence inhibition, supporting their suitability for PFAS exposure studies. Growth and bioluminescence responses were then assessed in M9 minimal medium following exposure to PFOA, PFNA, and PFHxS. Conventional growth measurements showed limited effects across tested concentrations, with strain-specific reductions observed mainly in P. putida F1 Tn5 luxCDABE. Bioluminescence revealed clearer compound- and concentration-dependent responses. PFOA and PFNA caused significant inhibition of peak bioluminescence in P. putida F1 Tn5 luxCDABE, while P. putida 12708 Tn5 luxCDABE showed a variable response, including increased luminescence following PFOA exposure and inhibition at higher concentrations of selected PFAS. These findings demonstrate that lux-marked P. putida biosensors are able to detect PFAS-induced metabolic stress with greater sensitivity than conventional growth assays and highlights their potential as rapid tools for ecotoxicological screening of fluorinated contaminants.