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Summary

Burkholderia species are highly resistant to heavy metals (HMs), yet their resistance mechanisms are largely unknown. In this study we screened 5000 mini-Tn5 transposon insertion mutants of Burkholderia cenocepacia H111 for loss of cadmium tolerance. Of the four genes identified three affected outer membrane biogenesis and integrity or DNA repair. The fourth gene, BCAE0587, encoded a P1-type ATPase belonging to the CadA family of HM exporters. CadA-deficient strains lost the ability to grow in the presence of cadmium, zinc and lead, whereas resistance to nickel, copper and cobalt was not affected. Expression studies using a transcriptional fusion of the cadA promoter to gfp confirmed this specificity, as induction was only observed in presence of cadmium, zinc and lead. The promoter activity was found to be highest at neutral pH with an activation threshold of 30 nM cadmium. Inoculation of the HM-hyperaccumulating plant Arabidopsis halleri with a RFP-marked derivative of B. cenocepacia H111 containing the PcadAgfp fusion demonstrated the applicability of this biosensor for monitoring cadmium at the single cell level in a natural environment.