Ref. ECF project 11/2003

Project Title : Development of a Rapid and Sensitive Bioassay of Heavy Metal Toxicity by Coupling a Transgenic Reporter Gene in a Mutant Nematode Strain

Applicant : Hong Kong University of Science and Technology

Total Approved Grant : $149,920

Duration : 15 March 2004 - 14 March 2005

Project Status/Remarks : Completed

Scope : The project aims to establish a hypersensitive nematode reporter strain to monitor heavy metal toxicity in environmental samples. This is achieved by selecting genetic mutants with impaired signaling pathway conferring heavy metal and stress resistance. A stress-inducible reporter established in our laboratory will be introduced into these mutant strains to generate new reporter strains of superior sensitivity to heavy metal stress. With the induced fluorescence detection, this strain will reveal the sub-lethal dose of heavy metal in micro-molar level within hours at a level of sensitivity comparable to that offered by a physico-chemical monitoring scheme.

Summary of findings :

Toxic substances in polluted water have adverse effects on human health and the ecosystem, whereas their bioavailability can only be assessed by bioassays. Caenorhabditis elegans with unique biological features, serves as an excellent indicator model organism. In this project, we have screened through over twenty different mutants and have identified nine mutant strains more sensitive by 1.5- to five-fold to heavy metals toxicity using cadmium as the testing pollutant. Among these more sensitive mutants are ones affecting genes required for environmental cue monitoring, chemosensory function and, surprisingly, a gene required for neural differentiation. Based on these results, five different tester and control double mutants were generated to verify the sensitivity of the mutant combination. Significant synergistic sensitization of response to heavy metals was observed in one double mutant, daf-16 unc-75. We have also tested two new stress-response reporter transgenes. Combining the more sensitive reporters with the double mutant, a supersensitive strain was created that is ten-fold more sensitive than the wild type animals. Combined with the shortening of the assay time, over one-hundred-fold of enhancement of sensitivity in monitoring heavy metal stress could be achieved. While additional double mutants with reporter transgenes are still being created near the end of this project, we have clearly demonstrated in principle that a combined approach of genetics and transgenesis can be used to create sensitive reporter indicator animals for environment monitoring bioassays.