F. Brulle

Transcriptomic underpinning of toxicant-mediated physiological function alterations in three terrestrial invertebrate taxa: A review.

Diverse anthropogenic activities often lead to the accumulation of inorganic and organic residues in topsoils. Biota living in close contact with contaminated soils may experience stress at different levels of biological organisation throughout the continuum from the molecular-genetic to ecological and community levels. To date, the relationship between changes at the molecular (mRNA expression) and biochemical/physiological levels evoked by exposures to chemical compounds has been partially established in a limited number of terrestrial invertebrate species. Recently, the advent of a family of transcriptomic tools (e.g. Real-time PCR, Subtractive Suppressive Hybridization, Expressed Sequence Tag sequencing, pyro-sequencing technologies, Microarray chips), together with supporting informatic and statistical procedures, have permitted the robust analyses of global gene expression changes within an ecotoxicological context. This review focuses on how transcriptomics is enlightening our understanding of the molecular-genetic responses of three contrasting terrestrial macroinvertebrate taxa (nematodes, earthworms, and springtails) to inorganics, organics, and agrochemicals.

Metallic trace element body burdens and gene expression analysis of biomarker candidates in Eisenia fetida, using an "exposure/depuration" experimental scheme with field soils.

Smelting plant activities lead to the accumulation of Metal Trace Elements (MTEs) in soils. The presence of high concentrations of MTEs can generate an environmental stress likely to affect macroinvertebrates living in close soil contact such as the Annelida Oligochaeta. Eisenia fetida, an ecotoxicologically important test species, was successively exposed to two field soils: (1) a highly contaminated agricultural topsoil collected near the former smelter Metaleurop Nord (Noyelles-Godault, France) which contaminated surrounding soils by its atmospheric emissions [exposure phase], and then (2) a slightly contaminated topsoil from an urban garden located in the conurbation of Lille (Wambrechies) [depuration phase]. Two analyses were performed during each phase. Firstly, the gene expression levels of four biomarker candidates identified in previous studies were analyzed in E. fetida coelomocytes. These candidates are Cd-metallothionein, phytochelatin synthase, coactosin-like protein and lysenin. Secondly, the body burdens of the following elements Cd, Pb, Zn, Cu, Fe, Ca, and P were measured. Moreover, both analyses were also performed in Lumbricus rubellus, an Annelid species collected from the two tested soil-originating sites. Analysis of gene expression and MTE body burdens in both species are discussed to: (1) evaluate expression biomarkers; (2) gain insight the detoxification processes and the long-term response to a metallic stress and (3) compare the responses observed in a test species (E. fetida) with the responses of a field species (L. rubellus).

Gene expression analysis of 4 biomarker candidates in Eisenia fetida exposed to an environmental metallic trace elements gradient: A microcosm study

Past activities of 2 smelters (Metaleurop Nord and Nyrstar) led to the accumulation of high amounts of Metal Trace Elements (TEs) in top soils of the Noyelles-Godault/Auby area, Northern France. Earthworms were exposed to polluted soils collected in this area to study and better understand the physiological changes, the mechanisms of acclimation, and detoxification resulting from TE exposure. Previously we have cloned and transcriptionally characterized potential biomarkers from immune cells of the ecotoxicologically important earthworm species Eisenia fetida exposed in vivo to TE-spiked standard soils. In the present study, analysis of expression kinetics of four candidate indicator genes (Cadmium-metallothionein, coactosin like protein, phytochelatin synthase and lysenin) was performed in E. fetida after microcosm exposures to natural soils exhibiting an environmental cadmium (Cd) gradient in a kinetic manner. TE body burdens were also measured. This microcosm study provided insights into: (1) the ability of the 4 tested genes to serve as expression biomarkers, (2) detoxification processes through the expression analysis of selected genes, and (3) influence of land uses on the response of potential biomarkers (gene expression or TE uptake).

Identification of suitable qPCR reference genes in leaves of Brassica oleracea under abiotic stresses

Real-time quantitative PCR is nowadays a standard method to study gene expression variations in various samples and experimental conditions. However, to interpret results accurately, data normalization with appropriate reference genes appears to be crucial. The present study describes the identification and the validation of suitable reference genes in Brassica oleracea leaves. Expression stability of eight candidates was tested following drought and cold abiotic stresses by using three different softwares (BestKeeper, NormFinder and geNorm). Four genes (BolC.TUB6, BolC.SAND1, BolC.UBQ2 and BolC.TBP1) emerged as the most stable across the tested conditions. Further gene expression analysis of a drought- and a cold-responsive gene (BolC.DREB2A and BolC.ELIP, respectively), confirmed the stability and the reliability of the identified reference genes when used for normalization in the leaves of B. oleracea. These four genes were finally tested upon a benzene exposure and all appeared to be useful reference genes along this toxicological condition. These results provide a good starting point for future studies involving gene expression measurement on leaves of B. oleracea exposed to environmental modifications.

Biomarker measurements in Trifolium repens and Eisenia fetida to assess the toxicity of soil contaminated with landfill leachate: a microcosm study.

To assess the toxicity of a soil contaminated with landfill leachate, biomarker measurements in two species living in close contact with the soil, i.e. a plant species Trifolium repens and an animal species Eisenia fetida, were conducted. Briefly, both species were studied after simultaneous exposure conducted in microcosms. The organisms were exposed to soil supplemented with pure leachate, leachate diluted to 50%; leachate diluted to 25% and without leachate. After a 10 weeks exposure period, we observed an increase in the Olive Trail Moment in T. repens, compared to the reference, for 50% and pure leachate. The response observed appears to be dose-dependent and linear in our experimental conditions. Addition of the leachate to the reference soil induced an increase in Cd-Metallothionein-coding mRNA quantity in E. fetida. In addition, expression level of another gene implied in detoxification and coding Phytochelatin synthase was significantly induced in worms exposed to the reference soil spiked with the leachate, regardless presence of T. repens. Thus, T. repens and E. fetida can be used in a complementary manner to assess soil quality. Sensitivities of the test species yield sensitive bioassays as both species responded at low doses despite the buffering effect of the soil.