Whole-cell biosensors for detection of heavy metal ions in environmental samples based on metallothionein promoters from Tetrahymena thermophila.
Bottom Line: In the present paper, we report results with two gene constructs using the Tetrahymena thermophila MTT1 and MTT5 metallothionein promoters linked with the eukaryotic luciferase gene as a reporter.Validation of these whole-cell biosensors was carried out using both artificial and natural samples, including methods for detecting false positives and negatives.Comparison with other published cell biosensors indicates that the Tetrahymena metallothionein promoter-based biosensors appear to be the most sensitive eukaryotic metal biosensors and compare favourably with some prokaryotic biosensors as well.
Affiliation: Departamento de Microbiología-III, Facultad de Biología, C/. José Antonio Novais 2, Universidad Complutense, Madrid, Spain.Show MeSH
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Mentions: To test the responses of the potential WCBs MTT1Luc and MTT5Luc, we exposed these cell cultures to a variety of individual heavy metals (Cd2+, Cu2+, Zn2+, Pb2+, As5+ and Hg2+), and combinations of metals. In addition, stress conditions unrelated to heavy metals were also tested, as described further below. Results with individual metals show that lucFF induction, when under the control of either the MTT1 or MTT5 promoters, is sensitive to both the type of metal and its concentration (Fig. 2). The bioluminescence increases with metal concentration to a maximum level, after which the level decreases because of the metal toxicity (Fig. 2). The majority of metals induce bioluminescence in both strains, including at very low metal concentrations. In general, Cd2+ induced the highest bioluminescence output for both recombinant strains (Fig. 2). However, the two strains showed differences in their level of induction. The induced bioluminescence from MTT5Luc (Fig. 2B) ranged between 10‐ to 60‐fold higher than the basal level, while for MTT1Luc the equivalent increase was about 2‐ to 18‐fold (Fig. 2A). Zn2+ and Hg2+ produced the lowest bioluminescence increases (< 10 times the basal levels) in the MTT5Luc strain (Fig. 2B). On the other hand, MTT1Luc showed small bioluminescent increases in response to Pb2+ or As5+, but responded strongly to Cd2+, Zn2+, Cu2+ or Hg2+ (Fig. 2A). Therefore, the two strains show differential bioluminescence output that is sensitive to both metal type and concentration.
Affiliation: Departamento de Microbiología-III, Facultad de Biología, C/. José Antonio Novais 2, Universidad Complutense, Madrid, Spain.