Use of Stress-Inducible Transgenic Nematodes as Biomarkers of Heavy Metal Pollution in Water Samples from an English River System

Abstract

Transgenic strains of the nematode Caenorhabditiselegans, which carry stress-inducible lacZ reporter genes, aremeasurably stressed by exposure to heavy metals in aqueous solution. Thisstress response can be quantified, using enzymatic assays for the reportergene-product (Escherichia coli beta-galactosidase), or estimatedapproximately by in situ staining for beta-galactosidase in exposedworms. Stress responses to heavy metals have been demonstrated both inlaboratory tests using Cd2+ or Hg2+, and also in watersamples taken from a metal-polluted river system in southwest England. TheRiver Carnon flows through an area with an ancient mining history,principally for Sn, but also for Cu and other metals; As, Cd, Al, Mn, and Zn,as well as large amounts of Fe, are all present in these ore bodies. Foursites in the Carnon river basin were compared with respect to theirmacroinvertebrate diversity, physical and chemical characteristics (includingthe concentrations of As, Cd, Al, Cu, Mn, Zn, and Fe). Transgenic worms wereexposed to water samples from these four sites, and also to a 0.33%(v/v) dilution of metal-laden minewater from the principal local mine (WhealJane). Transgene expression was induced in all five cases, though markedlyless so for the least polluted of the sites (which also supported a richermacroinvertebrate fauna). Two different transgenic strains were tested inthis study; strain PC72 (using a homologous hsp16 promoter) isslightly more sensitive to most metal-containing water samples than strainCB4027 (using a heterologous Drosophila hsp70 promoter). Bothtransgenic strains and two different assay methods gave essentially similarresults. These findings demonstrate that transgenic nematodes could provide arapid and simple assessment of aquatic pollution, in that the transgeneresponse is inducible by mixtures of dissolved metals at concentrationsactually encountered in metal-polluted watercourses.

The toxicity of dithiocarbamate fungicides to soil nematodes, assessed using a stress-inducible transgenic strain ofCaenorhabditis elegans

“…Thus b-galactosidase expression is inducible under stress conditions that activate the endogenous hsp16 genes [29]. When tested in parallel against metal-contaminated field water samples, strain PC72 shows slightly greater sensitivity than CB4027 [30] and is also advantageous in that optimal stress-inducibility is seen at 24-25ЊC rather than the 31-32ЊC needed for CB4027. In both strains, optimal stressor sensitivity is observed at temperatures 3-5ЊC below those required for heat induction of the hsp promoter used (28ЊC for hsp16, 34ЊC for hsp70 in C. elegans [31]).…”

“…Strain PC72 has been used as a biomonitor for quantitative assessment of stress responses to a wide range of chemical and physical stressors [32], including heavy metals [29], contaminated field samples [30], captan fungicides [33], immunological attack against the worm surface [34], and radiofrequency radiation as used in mobile telephones [35]. Much of this work has been done in aqueous media [29][30][31][32][33][34], but procedures for recovering worms from soil [17] allow this approach to be adapted for assessments of soil toxicity [36] (R. Power and D. de Pomerai, unpublished). Apart from a previous study involving captan fungicides [33], there are few reports of stress responses induced by widely used commercial pesticides, despite widespread public concern over possible health hazards associated with long-term chronic exposure.…”

Use of transgenic GFP reporter strains of the nematode Caenorhabditis elegans to investigate the patterns of stress responses induced by pesticides and by organic extracts from agricultural soils

“…The nematode Caenorahbditis elegans has been utilised widely for toxicity testing of field samples (Mutwakil et al 1997;Traunspurger et al 1997;Power and de Pomerai , 2001), as well as single and mixed toxicants (Leung et al 2008), including pesticides ranging from the fungicide captan (Candido and Jones, 1996) to a series of organophosphates (Rajini et al 2008). Several different types of assay are available, including whole-organism endpoints such as growth, reproduction and motility (Dhawan et al 1999; Thompson fully characterised somatic cell lineage (Sulston et al 1983), complete genome sequence (C. elegans Sequencing Consortium 1998), and the possibility of genome-wide RNA interference by feeding (Kamath et al 2003).…”

The Stress-Response Network in Animals: Proposals to Develop a Predictive Mathematical Model

“…We are currently testing a series of single toxicants, but will later progress to simple mixtures of 2 or 3 such toxicants. This approach was originally pioneered using hsp-16 and hsp-70 reporters [3][4][5] as biomarkers for both aquatic [34] and soil [35] contamination. Our list of test toxicants will include several metals as well as a variety of widely used pesticides acting on different targets.…”