Assessing the impact of Benzo[a]pyrene on Marine Mussels: Application of a novel targeted low density microarray complementing classical biomarker responses

Abstract

Despite the increasing use of mussels in environmental monitoring and ecotoxicological studies, their genomes and gene functions have not been thoroughly explored. Several cDNA microarrays were recently proposed for Mytilus spp., but putatively identified partial transcripts have rendered the generation of robust transcriptional responses difficult in terms of pathway identification. We developed a new low density oligonucleotide microarray with 465 probes covering the same number of genes. Target genes were selected to cover most of the well-known biological processes in the stress response documented over the last decade in bivalve species at the cellular and tissue levels. Our new ‘STressREsponse Microarray’ (STREM) platform consists of eight sub-arrays with three replicates for each target in each sub-array. To assess the potential use of the new array, we tested the effect of the ubiquitous environmental pollutant benzo[a]pyrene (B[a]P) at 5, 50, and 100 μg/L on two target tissues, the gills and digestive gland, of Mytilus galloprovincialis exposed invivo for three days. Bioaccumulation of B[a]P was also determined demonstrating exposure in both tissues. In addition to the well-known effects of B[a]P on DNA metabolism and oxidative stress, the new array data provided clues about the implication of other biological processes, such as cytoskeleton, immune response, adhesion to substrate, and mitochondrial activities. Transcriptional data were confirmed using qRT-PCR. We further investigated cellular functions and possible alterations related to biological processes highlighted by the microarray data using oxidative stress biomarkers (Lipofuscin content) and the assessment of genotoxicity. DNA damage, as measured by the alkaline comet assay, increased as a function of dose.DNA adducts measurements using 32P-postlabeling method also showed the presence of bulky DNA adducts (i.e. dG-N2-BPDE). Lipofiscin content increased significantly in B[a]P exposed mussels. Immunohistochemical analysis of tubulin and actin showed changes in cytoskeleton organisation. Our results adopting an integrated approach confirmed that the combination of newly developed transcriptomic approcah, classical biomarkers along with chemical analysis of water and tissue samples should be considered for environmental bioimonitoring and ecotoxicological studies to obtain holistic information to assess the impact of contaminants on the biota.

Role of mTOR in autophagic and lysosomal reactions to environmental stressors in molluscs

“…Adult Mytilus galloprovincialis Lam. (50.7 ± 2.8 mm) were collected from the intertidal zone at Trebarwith Strand,Cornwall,UK (50° 38' 40" N,4° 45' 44" W) in October 2014 (Banni et al, 2017). The site is relatively free of disease and is remotely located (Bignell et al, 2011).…”

“…Good seawater oxygenation was provided by a bubbling system. Seawater quality was monitored in each of the beakers by measuring salinity (35.4 ± 0.09‰), pH (7.9 ± 0.06), % dissolved oxygen (97.9 ± 3.22%) and temperature (15.3 ± 0.68 • C) (Banni et al, 2017). Groups of mussels were exposed to four treatments i.e.…”

“…The B[a]P concentrations used in these experiments were selected taking into account that the levels of PAHs in the sea water of contaminated environment range from 0.26 μg/L (Manodori et al, 2006), 18.34 μg/L (Sinaei and Mashinchian, 2014) to 46 μg/L (Nasher et al, 2013). After 3 d of exposure, the amount of chemical in the tissues of exposed animals was similar to that detected in the tissues of mussels sampled in field contaminated coastal waters (Banni et al, 2017;Widdows et al, 2002).…”

“…8). System complexity (connectance %) was strongly correlated with the concentration of B[a]P (inverse), lysosomal stability (direct), first principal component (direct) and DNA damage (inverse; COMET assay data from Banni et al, 2017; see also Canova et al, 1998) as shown in Fig. 9.…”

Relative comparison of tissue specific bioaccumulation and radiation dose estimation in marine and freshwater bivalve molluscs following exposure to phosphorus-32

“…These two species exhibit anatomical similarities, prevalence within respective water bodies and have known ability to concentrate contaminants within tissues. They are widely distributed and extensively used for ecotoxicological studies (Bayne 1976;Chatel et al 2012;Dallas et al 2012Dallas et al , 2013Binelli et al 2015;Jaeschke et al 2015;Banni et al 2017). They serve as important monitoring systems in coastal and inland water systems (Viarengo et al 2007;Bourgeault et al 2010;Binelli et al 2015;Sforzini et al 2018), and are important components in marine and freshwater food chains (Bayne 1976;Prejs et al 1990).…”