Comparative Biochemistry and Physiology Part C: Toxicology & Pharmacology
Are biochemical biomarker responses related to physiological performance of juvenile sea bass (Dicentrarchus labrax) and turbot (Scophthalmus maximus) caged in a polluted harbour?
Introduction
A multitude of xenobiotics contaminate the marine environment, and although chemical analyses are able to measure many of these compounds qualitatively and quantitatively, complex mixtures of these chemical pollutants cannot be fully assessed. Furthermore, chemical analyses alone do not reveal the impact of chemical pollution on the aquatic environment because of potential synergistic/antagonistic effects of complex mixtures of chemical pollutants. In this context, utilisation of biomarkers as an early warning of pollution or degradation in ecosystems has increased over the past 20 years (Adams 2002).
Identifying a suitable biomarker of toxicity represents a major challenge in current ecotoxicological research (Cheung et al. 2007). Many parameters have been investigated to assess disturbances of various physiological functions linked to chemical exposure (Van der Oost et al. 2003). Peakall (1994) defined biomarkers as “biological responses that can be related to an exposure to, or toxic effect of, an environmental chemical or chemicals”. The subdivision of biomarkers in literature is rather diffuse since the impact of toxic xenobiotics on fish has been analysed with various types of exposure and effect biomarkers, ranging from molecular, through cellular and physiological responses, to behavioural changes. In order to avoid confusion, in this paper, biomarkers acting at the subcellular level as biotransformation process or oxidative stress protection will be referred to as “biochemical biomarkers” and biomarkers reflecting fish growth or physiological condition will be referred to as “physiological biomarkers”.
Because many toxic effects initially occur at the subcellular level, there has been an increasing use of biochemical biomarkers that help to determine causative agents responsible for altered cellular function (Schlenk et al. 1996). Among the biochemical biomarkers described in relating literature, phase I and phase II biotransformation parameters such as EROD (ethoxyresorufin-O-deethylase) and GST (glutathione-S-transferase) activities are currently used in environmental risk assessment (Sanchez et al. 2008). Biotransformation of chemicals is a requisite for detoxification and excretion (Gravato and Santos 2003). The first step is usually catalysed by cytochrome P450-dependent monooxygenases (phase I) and their products, or several other organic pollutants, are subsequently coupled to endogenous metabolites (phase II) (Buhler and Williams, 1988, Landis and Yu, 1995). Antioxidant enzymes are also commonly used to understand the associated toxic-mechanisms of xenobiotics (Sanchez et al., 2005, Oliveira et al., 2008). Many pollutants exert their effects through redoxcycling, resulting in the production of reactive oxygen species (ROS). The role of antioxidant systems is to protect the cells from this oxidative stress. Thus, measurement of components of the antioxidant defence system may be helpful to determine organism exposure to pollutant (Bilbao et al. 2010).
Attempting to relate biomarker responses of individual organisms to increasing pollutant exposure and stress, both in the laboratory and in situ, offers considerable potential for improving the ecological relevance of ecotoxicological test procedures (Depledge et al. 1995). However, although the role of biochemical biomarkers as early warning tools is recognised, it is difficult to understand their significance at higher levels of biological organisation. Indeed, in spite of their rapid responsiveness and sensitivity to contaminant exposure, biochemical biomarkers have questionable ecological relevance, as a result of being endpoints at a low level of biological organisation (Castro et al., 2004). On the contrary, changes in physiology and fitness seem to be a common response in marine organism exposed to stressful pollutants (Alquezar et al., 2006, Faucher et al., 2008). Pollutants can induce various biological responses in fish, affecting the organisms from the biochemical to the population-community levels (Adams 2002). For example, many investigations on biological responses of fish populations to contamination indicated a general decrease of the relative fecundity, the growth rate and condition factor in contaminated estuaries (Laroche et al., 2002, Marchand et al., 2003, Amara et al., 2007). It is likely that changes in individual health manifest themselves at higher levels of ecological organisation, leading to reduced fish recruitment, abundance and production.
It has long been suggested that biochemical biomarkers should be used in conjunction with measurements of fitness (Anderson et al., 1994, Depledge et al., 1995). However, few studies have demonstrated correlative relationships between biochemical biomarkers responses and reduced fitness of aquatic organisms exposed to toxicants (Depledge et al., 1995, Lesser et al., 2001, Fonseca et al., 2009). Biological impairment (e.g. embryonic malformations, growth and condition depletion, fecundity and pollution tolerance) may directly affect the survivorship of organisms. Therefore, this type of multifaceted approach is important because it will improve our ability to use molecular biomarker responses of organisms to predict higher-level consequences of toxicant exposure (Rose et al. 2006).
Although biochemical or physiological biomarkers are intended to be useful tools for environmental assessment in the field, most of them have been developed under laboratory controlled toxic conditions. These studies often fail to recreate the mixed exposure situations occurring in nature. However, in field situations, the migration of many fish species for feeding and breeding creates uncertainty about how individuals sampled in some habitat truly reflects the water quality around the site of capture. In comparison, a caging strategy may, in specific situations, give more realistic results in studies of bioavailability, bioaccumulation and biological effects of contaminants in fish. The technique of caging offers advantages over organism chemical exposure (Oikari 2006) with a control of the precise location and duration of exposure while environmental field conditions are preserved. The use of cage-held animals from a common source (e.g. hatchery) also removes the potential for genotype adaptation, which is a distinct possibility in feral fish populations exposed to contaminants (Winter et al. 2005). The development of caging field experiments would provide an indication of the impact of contamination on marine fish and information on the applicability of this experimental strategy for assessing habitat quality.
The present study was designed to analyse the responses of two biotransformation parameters (EROD and GST activities) and an antioxidant enzyme (catalase) on the basis of a caging field experiment of two marine fish species in a polluted area (Boulogne sur Mer). In a second part, the responses of these enzymatic biomarkers were compared to different growth rates (in length and weight) and condition indices, measured on the same individual fish and analysed in a previous study (Kerambrun et al. submitted for publication). We used three condition indices: the Fulton's K condition factor; the RNA:DNA ratio which is used in numerous studies as indices for nutritional condition and growth assessment in larvae and juvenile fish (Buckley, 1984, Gwack and Tanaka, 2001, Amara et al., 2009); and a lipid storage index based on the ratio of the quantity of triacylglycerols (TAG; reserve lipids) to the quantity of sterols (ST; structural lipids) in fish (Amara et al. 2007).
Section snippets
Material and methods
This experiment was conducted in accordance with the Commission recommendation 2007/526/EC on revised guidelines for the accommodation and care of animals used for experimental and other scientific purposes. The University of Littoral Côte d'Opale is authorised to conduct experimentation on animals in its capacity as a certified establishment; according to the administrative order N° B62-160-2.
Environmental parameters
At the beginning of the experiment, bottom temperature (station A: 14.6 °C; station B: 14.9 °C), salinity (station A: 34.1 PSU; station B: 33.5 PSU) and oxygen contents (station A: 7.86 mg.L− 1; station B: 7.04 mg.L− 1) were similar in the two caging stations. The studied area temperature had increased during June and July; the temperature was 18 °C in both stations at the end of the caging exposure, which was also the highest temperature during the caging experiment.
The results of metal
Biochemical biomarkers responses
Fish, display close physiological relationships to their environment as ectothermic organisms, and as such, are sensitive to environmental disruptions and in particular to chemical stress. As a consequence many authors have begun measuring fish cellular detoxification or defence mechanisms that occur in response to exposure select environmental xenobiotics (Van der Oost et al., 2003). The first aim of this study was to analyse the response of three biochemical biomarkers in two marine fish
Conclusions
This multibiomarker approach allowed us to assess different levels of fish responses to chemical contamination. This study was a first approach of simultaneous comparison of both biochemical and physiological biomarkers in a caging study. For individual sea bass, biochemical biomarkers, growth and condition indices such as the Fulton's K condition index, the RNA:DNA ratio and the lipid storage index, showed numerous significant correlations. Conversely, there were only a few significant
Acknowledgments
This work was supported by post Grenelle programme 190, DEVIL of the French ministry for ecology and the Franco-British INTERREG IVA European project, DIESE. We would like to thank Michel LAREAL for making the cages, and Gregory Germain and François Gevaert for their help during their placement at sea. We thank Peter Magee for revision of the English grammar and syntax (www.anglais.webs.com).
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