Tracing explosives in soil with transcriptional regulators of Pseudomonas putida evolved for responding to nitrotoluenes.
Bottom Line: When strains bearing transcriptional fusions to reporters with an optical output (luxAB, GFP) were spread on soil spotted with nitrotoluenes, the signal triggered by promoter activation allowed localization of the target compounds on the soil surface.Our data provide a proof of concept that non-natural transcription factors evolved to respond to nitroaromatics can be engineered in soil bacteria and inoculated on a target site to pinpoint the presence of explosives.This approach thus opens new ways to tackle this gigantic humanitarian problem.
Affiliation: Centro Nacional de Biotecnología-CSIC, Campus de Cantoblanco, Madrid 28049, Spain.Show MeSH
Related in: MedlinePlus
Mentions: In order to assess further the ability of XylR3 and XylR5 for detection of explosive‐related compounds, we next compared the responses of P. putida cells bearing plasmids encoding each of these proteins in liquid versus agar media. Water limitation and matric stress, a frequent condition of bacteria living in soil, is known to dramatically alter both overall physiology and the structure and activity of specific proteins (Cytryn et al., 2007). In order to compare responses in excess water (liquid medium) with those in non‐saturating water conditions (growth on 1.5% agar plates) we streaked out strain P. putida Po→luxAB (Table 1), bearing plasmids pCON916 (xylR+), pCON922 (xylR3+) and pCON924 (xylR5+) on agar plates amended with 2,4 DNT. In this case, the cells were endowed with a luminiscent luxAB reporter fusion that can be visualized non‐disruptively (Experimental procedures). Following growth, plates were exposed to traces of n‐decanal and placed on X‐ray film. The results of such a procedure for the xylR5‐bearing strain are shown in Fig. 3A (nearly identical results were obtained with the xylR3 counterpart, not shown). The signal brought about by luxAB as reporter basically reproduced the β‐galactosidase data of liquid culture of Fig. 2B. These results indicated that the induction properties of the XylR mutants revealed in liquid cultures were kept in a medium not saturated with water. Furthermore, the data of Fig. 3A suggested that luxAB was a suitable visual reporter for revealing the presence of the nitroaromatics under study.
Affiliation: Centro Nacional de Biotecnología-CSIC, Campus de Cantoblanco, Madrid 28049, Spain.