Measuring the effects of selection on the genome imposed by human-altered environment is currently a major goal in ecological genomics. Given the polygenic basis of most phenotypic traits, quantitative genetic theory predicts that selection is expected to cause subtle allelic changes among covarying loci rather than pronounced changes at few loci of large effects. The goal of this study was to test for the occurrence of polygenic selection in both North Atlantic eels (European Eel, Anguilla anguilla and American Eel, A. rostrata), using a method that searches for covariation among loci that would discriminate eels from 'control' vs. 'polluted' environments and be associated with specific contaminants acting as putative selective agents. RAD-seq libraries resulted in 23 659 and 14 755 filtered loci for the European and American Eels, respectively. A total of 142 and 141 covarying markers discriminating European and American Eels from 'control' vs. 'polluted' sampling localities were obtained using the Random Forest algorithm. Distance-based redundancy analyses (db-RDAs) were used to assess the relationships between these covarying markers and concentration of 34 contaminants measured for each individual eel. PCB153, 4'4'DDE and selenium were associated with covarying markers for both species, thus pointing to these contaminants as major selective agents in contaminated sites. Gene enrichment analyses suggested that sterol regulation plays an important role in the differential survival of eels in 'polluted' environment. This study illustrates the power of combining methods for detecting signals of polygenic selection and for associating variation of markers with putative selective agents in studies aiming at documenting the dynamics of selection at the genomic level and particularly so in human-altered environments.

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The European eel (Anguilla anguilla L.) is severely threatened with extinction. Surprisingly, even though their unusual life cycle makes them particularly vulnerable to pollution, the possible contribution of contamination remains especially poorly known. Here we have investigated the possible effect of cadmium (Cd), a widespread nonessential metal, on eel reproductive capacities. Both control and Cd precontaminated female silver eels were experimentally matured and forced to swim in metal-free conditions to mimic their reproductive migration. Cd pre-exposure was found to strongly stimulate the pituitary-gonad-liver axis of maturing female silver eels leading to early and enhanced vitellogenesis. This was followed by a strong phenomenon of oocyte atresia and eel mortality. These phenomena occurred before oocytes could reach full maturation and were associated with a large entry of both vitellogenin and Cd into the ovaries. Indeed, a redistribution of previously stored cadmium, even from the low Cd levels of control eels, was observed during sexual maturation. Atresia and mortality phenomena were also associated with an overexpression of the pituitary gene encoding the growth hormone, a marker of physiological stress and energy reserves exhaustion. Significantly, these devastating effects of Cd were observed in organisms that presented liver and kidney Cd concentrations still below those observed in eels from Cd contaminated hydrosystems. Our research shows how common levels of cadmium contamination could disrupt endocrine pathways implicated in gonad maturation and subsequently impair reproductive capacity of eel future genitors.

There is increasing evidence that epigenetics can play a key role in the etiology of diseases engendered by chronic pollutant exposure. Although epigenetics has received significant attention in the field of biomedicine during the last years, epigenetics research is surprisingly very limited in ecotoxicology. The aim of the present study was to investigate the possible effects of low-dose cadmium exposure on the DNA methylation profile in a critically endangered fish species, the European eel. Eels were exposed to environmentally realistic concentrations of cadmium (0.4 and 4 μg·L(-1)) during 45 days. The global CpG methylation status of eel liver was determined by means of a homemade ELISA assay. We then used a methylation-sensitive arbitrarily primed PCR method to identify genes that are differentially methylated between control and Cd-exposed eels. Our results show that cadmium exposure is associated with DNA hypermethylation and with a decrease in total RNA synthesis. Among hypermethylated sequences identified, several fragments presented high homologies with genes encoding for proteins involved in intracellular trafficking, lipid biosynthesis, and phosphatidic acid signaling pathway. In addition, few fragments presented high homologies with retrotransposon-like sequences. Our study illustrates how DNA methylation can be involved in the chronic stress response to Cd in fish.

To investigate the mechanisms involved in metal stress in wild fish, yellow perch (Perca flavescens) were collected in eight lakes of the Rouyn-Noranda and Sudbury regions (Canada). Due to mining and smelting activities, these two regions indeed present a broad contamination gradient in metal concentrations (Cd, Cu, Zn and Ni; water, sediment and prey) and offer a unique research opportunity to investigate relationships between metal bioaccumulation and resulting deleterious effects in indigenous biota chronically exposed to metal mixtures. The expression level of genes encoding for proteins involved in metal detoxification (metallothioneins, mts), protein protection (heat shock protein-70, hsp-70), growth (insulin-like growth factor-1, igf-1), aerobic energy metabolism (cytochrome c oxydase, cco-1) and protection against oxidative stress (Cu/Zn superoxide dismutase, sod-1) were assessed in fish liver and muscle in association with protein and enzymatic assays for cytochrome c oxidase (CCO). Bioaccumulation of both Cd and Cu increased in response to higher ambient metal concentrations, but the two metals clearly have different modes of action. For Cd, changes in gene expression levels were more marked in the liver than in the dorsal muscle, whereas for Cu the opposite trend was observed. Hepatic Cd accumulation was linked to decreased cco-1 and sod-1 gene expression, whereas Cu accumulation was associated with a decrease in CCO enzymatic activity and an increase in total protein concentration and in cco-1, mts and hsp-70 gene expression levels. For Ni, no significant correlations were observed at the transcriptional level, but increasing hepatic Ni concentrations were significantly and positively correlated with protein concentrations and CCO activity. By coupling gene expression to biochemical and physiological endpoints, this work provides new insights into the mechanisms involved in metal stress and the adaptive response of fish chronically exposed to metal mixtures.

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Given the inherent variability of aquatic systems, predicting the in situ effects of contaminants on such ecosystems still represents a major challenge for ecotoxicology. In this context, transcriptomic tools can help identify and investigate the mechanisms of toxicity beyond the traditional morphometric, physiological and population-level endpoints. In this study, we used the 454 sequencing technology to examine the in situ effects of chronic metal (Cd, Cu) exposure on the yellow perch (Perca flavescens) transcriptome. Total hepatic mRNA from fish sampled along a polymetallic gradient was extracted, reverse transcribed, labeled with unique barcode sequences and sequenced. This approach allowed us to identify correlations between the transcription level of single genes and the hepatic concentrations of individual metals; 71% of the correlations established were negative. Chronic metal exposure was thus associated with a decrease in the transcription levels of numerous genes involved in protein biosynthesis, in the immune system, and in lipid and energy metabolism. Our results suggest that this marked decrease could result from an impairment of bile acid metabolism by Cd and energy restriction but also from the recruitment of several genes involved in epigenetic modifications of histones and DNA that lead to gene silencing.