Toxicogenomics of Eohaustorius estuarius following pyrethroid and ammonia exposure
ABSTRACT: A variety of contaminants find their way to the marine sediments from different sources, and these contaminants can pose serious risks to the natural marine flora and fauna. For example, pyrethroids, which are a potent pesticide family, are often used in agriculture fields worldwide, and these find their way into the marine environment through run off. Further, pyrethroids are used in farmed Atlantic salmon cages in Chile, Great Britain and Norway. Ammonia is another contaminant that is used in agriculture in form of ammonia-rich fertilizer and can be carried during run-offs to localized rivers and streams. Ammonia is also detectable after emission of effluents from sewage treatment plants and industrial plants like oil refineries and meat processing plants. Contaminants may have short and long term effects on non-target organisms living in the water column or in the marine sediment. Importantly, the sediment ecosystem houses a variety of plants, animals and crustaceans, including the American lobster Homarus americanus. Lobster is the most fished crustacean in New Brunswick and Quebec and its resale and exportation produced over $1.6 billion in 2011. Due to its economic and environmental importance, it is essential to study the effects of contaminants present in its ecosystem. Sediment samples are often used as pollution markers during toxicity testing due to their tendency to accumulate hydrophobic contaminants. To better understand the possible effects of contaminants in sediment, a total gene expression study was developed using the marine amphipod Eohaustorius estuarius. A 10 day spike-in exposure was performed using ammonia and two pyrethroids, namely cypermethrin and deltamethrin. As pyrethroids and ammonia are known to have vastly different mechanisms of action in living organisms, we compared global gene expression patterns following exposure to ammonia against the patterns observed following exposure to pyrethroids. Total gene expression was measured by oligonucleotide microarray. The expression of five genes of interest involved in different biological processes such as metabolism, transcription, translation, immunity and stress, which were found to be differently expressed by microarray, was validated by RT-qPCR. A set of genes was identified that showed differential expression levels in a treatment-dependent manner, thus further highlighting the different mechanisms of action of ammonia and pyrethroids in the marine sediment. This study provides a proof of concept for the use of DNA microarrays with model crustaceans for the study of marine sediment contaminants. This specific study is aimed at evaluating the effect of ammonia and pyrethroid exposure on E.estuarius and to identify possible biomarkers of these exposures.
Project description:Chemical analysis of the compounds present in sediment, although informative, often is not indicative of the downstream biological effects that these contaminants exert on resident aquatic organisms. More direct molecular methods are needed to determine if marine life is affected by exposure to sediments. In this study, we used an aquatic multispecies microarray and q-PCR to investigate the effects on gene expression in juvenile sea bream (Sparus aurata) of two contaminated sediments defined as sediment 1 and 2 respectively, from marine areas in Northern Italy.
Project description:The processing of seafood for human or animal consumption creates vast amounts of by-product, which is often considered waste. Although some of these by-products are used as fishmeal, bone-meal or fertilizer, as a whole, it remains under utilized. Significant amounts of proteins, lipid fractions, vitamins, and other bioactive molecules are present in these by-products, all with potential beneficial properties that could be used as alternatives to fishmeal or as supplements for aquaculture species. In an attempt to investigate their potential benefit in Atlantic salmon fish nutrition, nine experimental diets were formulated using by-products originating from various seafood processing plants. A control basal diet (no by-product added) was also formulated. Juvenile Atlantic salmons were fed one of the nine experimental diets (30% marine by-product, 70% basal diet) or the basal diet and hepatic gene expression profiling was done on fish fed each diet after 14 and 56 days. Analysis of hepatic gene expression revealed a significant amount of differentially expressed genes for each diet with roles in various pathways and biological processes. By comparing differences in hepatic gene expression levels with the nutritional composition of the various feeds, we were able to identify a number of nutritional elements that affect specific gene families. This information will be very useful for the formulation of novel fish feeds, which may be designed with specific aims, such as rapid growth, increased immunity or better general health This specific study is aimed at evaluating the hepatic transcriptional responses in juvenile Atlantic salmon fed with fish feed formulation supplemented with one of nine marine by-products.
Project description:Aquatic organisms are exposed to many toxic chemicals and interpreting the cause and effect relationships between occurrence and impairment is difficult. Toxicity Identification Evaluation (TIE) provides a systematic approach for identifying responsible toxicants. TIE relies on relatively uninformative and potentially insensitive toxicological endpoints. Gene expression analysis may provide needed sensitivity and specificity aiding in the identification of primary toxicants. The current work aims to determine the added benefit of integrating gene expression endpoints into the TIE process. A cDNA library and a custom microarray were constructed for the marine amphipod Ampelisca abdita. Phase 1 TIEs were conducted using 10% and 40% dilutions of acutely toxic sediment. Gene expression was monitored in survivors and controls. An expression-based classifier was developed and evaluated against control organisms, organisms exposed to low or medium toxicity diluted sediment, and chemically selective manipulations of highly toxic sediment. The expression-based classifier correctly identified organisms exposed to toxic sediment even when little mortality was observed, suggesting enhanced sensitivity of the TIE process. The ability of the expression-based endpoint to correctly identify toxic sediment was lost concomitantly with acute toxicity when organic contaminants were removed. Taken together, this suggests that gene expression enhances the performance of the TIE process. Wild-collected Ampelisca abdita were exposed to either control (from sites in Long Island Sound, labeled LIS) sediment, toxic (from site on Elizabeth River, labeled ER) sediment, a series of mixtures of LIS and ER sediment, sediments manipulated to alter toxin bioavailability, or toxicant amended sediments. Lethality was scored, and survivors were subjected to mRNA expression analysis via oligo microarray.
Project description:The zebrafish embryo has repeatedly proved to be a useful model for the analysis of effects by environmental toxicants. This study was performed to investigate if an approach combining mechanism-specific bioassays with microarray techniques can obtain more in-depth insights into the ecotoxicity of complex pollutant mixtures as present, e.g., in freeze-dried whole sediment samples and their corresponding organic extracts in parallel. To this end, altered gene expression was compared to data from established bioassays as well as to results from chemical analysis. Microarray analysis revealed several classes of significantly regulated genes which could to a considerable extent be related to the hazard potential. Results indicate that potential classes of contaminants can be assigned to sediment extracts by both classical biomarker genes and corresponding expression profile analyses of known substances. However, it is difficult to distinguish between specific responses and more universal detoxification of the organism. Additionally, different gene expression was shown to be less influenced by the sampling site than by the method of exposure, which could be attributed to differential bioavailability of contaminants. Microarray analyses were performed with early life stages of zebrafish exposed to sediment extracts or freeze-dried sediment from three sampling sites (Ehingen, Lauchert, Sigmaringen) along the Upper part of the Danube River, Germany. The expression profiles were compared within the sampling sites, between the exposure scheme and to the expression pattern of model toxicants, such as 4-chloroaniline, Cadmium, DDT, TCDD, and Valproic acid (Gene Expression Omnibus Series GSE9357). Additionally, mechanism-specific bioassays and chemical analysis of the sediments have been combined and compared to the present gene expression data.
Project description:The experiment intended to identify biomarker genes for arsenite exposure in rice roots. These biomarker genes were tested as alternative endpoints in an sediment-contact test with rice (Brinke et al. Environ Sci Pollut R 22.16 (2015): 12664-12675)
Project description:The abundance of bacterial (AOB) and archaeal (AOA) ammonia oxidisers, assessed using quantitative PCR measurements of their respective a-subunit of the ammonia monooxygenase (amoA) genes, and ammonia oxidation rates were measured in four contrasting coastal sediments in the Western English Channel. Sediment was sampled bimonthly from July 2008 to May 2011, and measurements of ammonia oxidiser abundance and activity compared to a range of environmental variables including salinity, temperature, water column nutrients and sediment carbon and nitrogen content. Despite a higher abundance of AOA amoA genes within all sediments, and at all time-points, rates of ammonia oxidation correlated with AOB and not AOA amoA gene abundance. Other than ammonia oxidation rate, sediment particle size was the only variable that correlated with the spatial and temporal patterns of AOB amoA gene abundance, implying a preference of the AOB for larger sediment particles. This is possibly due to deeper oxygen penetration into the sandier sediments, increasing the area available for ammonia oxidation to occur, higher concentrations of inhibitory sulphide with pore waters of muddier sediments or a combination of both oxygen and sulphide concentrations. Similar to many other temporal studies of nitrification within estuarine and coastal sediments, decreases in AOB amoA gene abundance were evident during summer and autumn, with maximum abundance and ammonia oxidation rates occurring in winter and early spring. The lack of correlation between AOA amoA gene abundance and ammonium oxidation rate suggests an alternative role for amoA-carrying AOA within these sediments. Two color array (Cy3 and Cy5): the universal standard 20-mer oligo is printed to the slide with a 70-mer oligo (an archetype). Environmental DNA sequences (fluoresced with Cy3) within 15% of the 70-mer conjugated to a 20-mer oligo (fluoresced with Cy5) complementary to the universal standard will bind to the oligo probes on the array. Signal is the ratio of Cy3 to Cy5. Three replicate probes were printed for each archetype. Two replicate arrays were run on duplicate targets.
Project description:Nitrogen assimilation in plants is a tightly regulated process that integrates developmental and environmental signals. The legume-rhizobial symbiosis results in the formation of a specialized organ called root nodule, where the rhizobia fixes atmospheric nitrogen into ammonia. Ammonia is assimilated by the plant enzyme glutamine synthetase, which is specifically inhibited by PPT. The expression of key genes related to the regulation of root nodule metabolism will likely be affected by glutamine synthetase inhibition. We used microarrays to detail the global programme of gene expression in response to Glutamine synthetase inhibition in root nodules and identified genes differentially expressed over a time course. Medicago truncatula nodulated plants (20 days post inoculation) were treated with 0.25 mM of PPT. Root nodules were harvested at 4, 8 and 24 hours after PPT application. As a control, root nodules collected just before PPT application were used (PPT 0h). Three biological replicates consisting of pools of root nodules harvested from five distinct plants were used for RNA extraction and hybridization on Affymetrix GeneChips.
Project description:To further explore the biotoxicity mechanisms of zinc oxide nanoparticles (ZnO NPs) and the recovery strategies of the accordingly impaired Nitrosomonas europaea (N. europaea, ATCC 19718) cells, a genome-sequenced model ammonia-oxidizing bacterium (AOB) commonly detected in the activated sludge of biological wastewater treatment plants, whole-genome microarray analysis was applied to retrieve the induced transcriptional responses, after their physiological and metabolic activities were revealed. The whole-genome expressions were measured after exposure to 50 ppm ZnO NPs and 12-hrs recovery incubation when the ammonia removal rate (ARA) declined by 10% in the chemostat bioreactor. Three independent experiments were performed for each experiment.
Project description:To further explore the biotoxicity mechanisms of CeO2 nanoparticles (NPs) and the recovery strategies of the according impaired Nitrosomonas europaea (N. europaea, ATCC 19718) cells, a genome-sequenced model ammonia oxidizing bacterium (AOB) commonly detected in the activated sludge of biological wastewater treatment plants, the whole-genome microarray analysis was applied to retrieve the induced transcriptional responses, after their physiological and metabolic activities were evealed. Overall design: The whole-genome expressions were measured after exposure to 50 ppm CeO2 NPs and 12-hr recovery incubation when the ammonia removal rate (ARA) declined by 10% in the chemostat bioreactor .Three biological replicates were included for exposure and recovery experiment.
Project description:Salmon infected with an ectoparasitic marine copepod, the salmon louse Lepeophtheirus salmonis, incur a wide variety of consequences depending upon host sensitivity. Juvenile pink salmon (Oncorhynchus gorbuscha) migrate from natal freshwater systems to the ocean at a young age relative to other Pacific salmon, and require rapid development of appropriate defences against marine pathogens. We analyzed the early transcriptomic responses of naïve juvenile pink salmon of sizes 0.3g (no scales), 0.7g (mid-scale development) and 2.4g (scales fully developed) to a low-level laboratory exposure with early moult stage L. salmonis. All infected size groups exhibited unique transcriptional profiles. Inflammation and inhibition of cell proliferation was identified in the smallest size class (0.3g), while increased glucose absorption and retention was identified in the middle size class (0.7g). Tissue-remodelling genes were also up-regulated in both the 0.3g and 0.7g size groups. Profiles of the 2.4g size class indicated cell-mediated immunity and possibly parasite-induced growth augmentation. Understanding a size-based threshold of resistance to L. salmonis is important for fisheries management. This work characterizes molecular responses reflecting the gradual development of innate immunity to L. salmonis between the susceptible (0.3g) and refractory (2.4g) pink salmon size classes. Six-condition experiment, 3 size groups each infected and uninfected, duplicate tanks. Biological replicates: 6 control, 6 infected for each size group. Sampled 6 days post exposure. cDNA samples with reference design (aRNA), two-color array.