Transcription of NOD1 and NOD2 and their interaction with CARD9 and RIPK2 in IFN signaling in a perciform fish, the Chinese perch, Siniperca chuatsi.

NOD1 and NOD2 as two representative members of nucleotide-binding oligomerization domain (NOD)-like receptor (NLR) family play important roles in antimicrobial immunity. However, transcription mechanism of nod1 and nod2 and their signal circle are less understood in teleost fish. In this study, with the cloning of card9 and ripk2 in Chinese perch, the interaction between NOD1, NOD2, and CARD9 and RIPK2 were revealed through coimmunoprecipitation and immunofluorescence assays. The overexpression of NOD1, NOD2, RIPK2 and CARD9 induced significantly the promoter activity of NF-κB, IFNh and IFNc. Furthermore, it was found that nod1 and nod2 were induced by poly(I:C), type I IFNs, RLR and even NOD1/NOD2 themselves through the ISRE site of their proximal promoters. It is thus indicated that nod1 and nod2 can be classified also as ISGs due to the presence of ISRE in their proximal promoter, and their expression can be mechanistically controlled through PRR pathway as well as through IFN signaling in antiviral immune response.

Informing the Exposure Landscape: The Fate of Microplastics in a Large Pelagic In-Lake Mesocosm Experiment.

Understanding microplastic exposure and effects is critical to understanding risk. Here, we used large, in-lake closed-bottom mesocosms to investigate exposure and effects on pelagic freshwater ecosystems. This article provides details about the experimental design and results on the transport of microplastics and exposure to pelagic organisms. Our experiment included three polymers of microplastics (PE, PS, and PET) ranging in density and size. Nominal concentrations ranged from 0 to 29,240 microplastics per liter on a log scale. Mesocosms enclosed natural microbial, phytoplankton, and zooplankton communities and yellow perch (Perca flavescens). We quantified and characterized microplastics in the water column and in components of the food web (biofilm on the walls, zooplankton, and fish). The microplastics in the water stratified vertically according to size and density. After 10 weeks, about 1% of the microplastics added were in the water column, 0.4% attached to biofilm on the walls, 0.01% within zooplankton, and 0.0001% in fish. Visual observations suggest the remaining >98% were in a surface slick and on the bottom. Our study suggests organisms that feed at the surface and in the benthos are likely most at risk, and demonstrates the value of measuring exposure and transport to inform experimental designs and achieve target concentrations in different matrices within toxicity tests.

Preparing for the future offspring: European perch (Perca fluviatilis) biosynthesis of physiologically required fatty acids for the gonads happens already in the autumn.

Long-chain polyunsaturated fatty acids (PUFA) are critical for reproduction and thermal adaptation. Year-round variability in the expression of fads2 (fatty acid desaturase 2) in the liver of European perch (Perca fluviatilis) in a boreal lake was tested in relation to individual variation in size, sex, and maturity, together with stable isotopes values as well as fatty acids (FA) content in different tissues and prey items. ARA and DHA primary production was restricted to the summer months, however, perch required larger amounts of these PUFA during winter, as their ARA and DHA muscle content was higher compared to summer. The expression of fads2 in perch liver increased during winter and was higher in mature females. Mature females stored DHA in their gonads already in late summer and autumn, long before the upcoming spring spawning period in May. Lower δ13CDHA values in the gonads in September suggest that these females actively synthesized DHA as part of this reproductive investment. Lower δ13CARA values in the liver of all individuals during winter suggest that perch were synthesizing essential FA to help cope with over-wintering conditions. Perch seem able to modulate its biosynthesis of physiologically required PUFA in situations of stress (fasting or cold temperatures) or in situations of high energetic demand (gonadal development). Biosynthesis of physiologically required PUFA may be an important part of survival and reproduction in aquatic food webs with long cold periods.

Molecular Analysis and Sex-specific Response of the Hepcidin Gene in Yellow Perch (Perca Flavescens) Following Lipopolysaccharide Challenge.

Hepcidin antimicrobial peptide (hamp) is active in teleosts against invading pathogens and plays important roles in the stress and immune responses of finfish. The response of hamp gene was studied in yellow perch (yp) (Perca flavescens) challenged with lipopolysaccharides to understand if this immunity response is sex-specifically different. The cloned hamp gene consists of an open-reading frame of 273 bp and encodes a deduced protein of 90 amino acids (a.a.), which includes a signal peptide of 24 a.a., a pro-domain of 40 a.a. and a mature peptide of 26 a.a. Yp hamp involves 8 cysteine residues with 4 disulfide bonds, and a protein with an internal alpha helix flanked with C- and N-terminal random coils was modeling predicted. RT-qPCR was used to analyze the relative abundances (RAs) of hamp mRNA in the livers of juvenile female and male yellow perch challenged with lipopolysaccharide. The expression levels of hamp were significantly elevated by 3 h (RA = 7.3) and then peaked by 6 h (RA = 29.4) post-treatment in females but the peak was delayed to 12 h (RA = 65.4) post-treatment in males. The peak mRNA level of challenged males was shown 7.6-fold higher than females. The post-treatment responses in both genders decreased to their lowest levels by 24 h and 48 h. Overall, female perch had an earlier but less-sensitive response to the lipopolysaccharide challenge than male.

Significance of chemical affinity on metal subcellular distribution in yellow perch (Perca flavescens) livers from Lake Saint-Pierre (QUEBEC, Canada).

The subcellular partitioning approach provides useful information on the location of metals within cells and is often used on organisms with high levels of bioaccumulation to establish relationships between the internal concentration and the potential toxicity of metals. Relatively little is known about the subcellular partitioning of metals in wild fish with low bioaccumulation levels in comparison with those from higher contaminated areas. This study aims to examine the subcellular partitioning of various metals considering their chemical affinity and essentiality at relatively low contamination levels. Class A (Y, Sr), class B (Cu, Cd, MeHg), and borderline (Fe, Mn) metal concentrations were measured in livers and subcellular fractions of yellow perch (n = 21) collected in Lake Saint-Pierre, QC, Canada. The results showed that all metals, apart from MeHg, were distributed among subcellular fractions according to their chemical affinity. More than 60% of Y, Sr, Fe, and Mn were found in the metal-sensitive fractions. Cd and Cu were largely associated with the metallothionein-like proteins and peptides (60% and 67% respectively) whereas MeHg was found mainly in the metal-sensitive fractions (86%). In addition, the difference between the subcellular distribution of Cu and other essential metals like Fe and Mn denotes that, although the essentiality of some metals is a determinant of their subcellular distribution, the chemical affinity of metals is also a key driver. The similarity of the subcellular partitioning results with previous studies on yellow perch and other fish species from higher contaminated areas supports the idea that metals are distributed in the cellular environment according to their chemical properties regardless of the bioaccumulation gradient.

Pesticides Inhibit Retinoic Acid Catabolism in PLHC-1 and ZFL Fish Hepatic Cell Lines.

The population of yellow perch (Perca flavescens) in lake Saint-Pierre (QC, Canada) has been dramatically declining since 1995 without any sign of recovery. Previous studies have shown disrupted retinoid (vitamin A) metabolic pathways in these fish, possibly due to the influence of pesticides. Our study aimed to evaluate the impact of some herbicides and neonicotinoids on retinoic acid catabolism in the fish hepatic cell lines PLHC-1 and ZFL. We hypothesized that pesticides accelerate the catabolism of retinoic acid through oxidative stress that exacerbates the oxidation of retinoic acid. Results obtained with talarozole, a specific CYP26A1 inhibitor, and ketoconazole, a generalist inhibitor of cytochrome-P450 enzymes, revealed that CYP26A1 is mainly responsible for retinoic acid catabolism in ZFL but not PLHC-1 cells. The impacts of pesticides on retinoic acid catabolism were evaluated by incubating the cells with all-trans-retinoic acid and two herbicides, atrazine and glyphosate, or three neonicotinoids, clothianidin, imidacloprid, and thiamethoxam. Intracellular thiols and lipid peroxidation were measured following pesticide exposure. The possible causal relation between oxidative stress and the perturbation of retinoic acid catabolism was investigated using the antioxidant N-acetylcysteine. The data revealed that pesticides inhibit retinoic acid catabolism, with the involvement of oxidative stress in the case of atrazine, imidacloprid, and thiamethoxam but not with clothianidin and glyphosate. Pesticides also affected the isomerization of all-trans-retinoic acid over time, leading to an increased proportion of active isomers. These results hint at a possible perturbation of retinoic acid catabolism in fish living in pesticide-contaminated waters, as suggested by several in vivo studies. Such a disruption of retinoid metabolism is worrying, given the numerous physiological pathways driven by retinoids.

Impacts on antioxidative enzymes and transcripts in darter (Etheostoma spp.) brains in the Grand River exposed to wastewater effluent.

The Grand River watershed is the largest in southern Ontario and assimilates thirty wastewater treatment plants (WWTP) with varied degrees of treatment. Many WWTPs are unable to effectively eliminate several contaminants of emerging concern (CECs) from final effluent, leading to measurable concentrations in surface waters. Exposures to CECs have reported impacts on oxidative stress measured through antioxidative enzymes (SOD, CAT, GPX). This study focuses on the effects of WWTP effluent on four Etheostoma (Darter) species endemic to the Grand River, by investigating if increased antioxidative response markers are present in darter brains downstream from the effluent outfall compared to an upstream reference site relative to the Waterloo, ON WWTP across two separate years (Oct 2020 and Oct 2021). This was assessed using transcriptional and enzyme analysis of antioxidant enzymes and an enzyme involved in serotonin synthesis, tryptophan hydroxylase (tph). In fall 2020, significant differences in transcript markers were found between sites and sexes in GSD with SOD and CAT showing increased expression downstream, in JD with both sexes showing increased SOD downstream, and an interactive effect for tph in RBD. Changes in transcripts aligned with enzyme activity where interactive effects with sex-related differences were observed in fish collected fall 2020. In contrast, transcripts measured in fall 2021 were increased upstream compared to downstream species in RBD and GSD. This study additionally displayed yearly, species and sex differences in antioxidant responses. Continued investigation on the impacts of CECs in effluent in non-target species is required to better understand WWTP effluent impacts.

Effect of starvation and refeeding on reactive oxygen species, autophagy and oxidative stress in Chinese perch (Siniperca chuatsi) muscle growth.

In skeletal muscle, autophagy regulates the development and growth of muscle fibres and maintains the normal muscle metabolism. Under starvation and refeeding conditions, the effect of reactive oxygen species (ROS) levels on skeletal muscle autophagy is still unclear, although the excessive accumulation of ROS has been shown to increase autophagy in cells. The purpose of this study was to explore the effects of starvation and diet after starvation on the autophagy of adult Chinese perch muscle, and to determine the level of ROS in the muscle. We performed zero (Normal control), three and seven starvation treatments on adult Chinese perch, and returned to normal feeding for 3 days after starvation for 7 days. In the muscles of the adult Chinese perch muscle after 3 days of starvation, the autophagy marker protein LC3 and the number of autophagosomes remained basically the same as in the normal feeding situation. However, on starvation for 7 days, the mitochondrial autophagy was sensitive and the number of autophagosomes increased, but the antioxidant-related molecules (malondialdehyde, catalase, glutathione S-transferase, glutathione and anti-superoxide anion) decreased and the accumulation of ROS was obvious. In addition, the extended starvation time also increased the level of LC3 protein. However, by refeeding after starvation this nutritional stress resulted in a decrease in ROS levels and a partial restoration of antioxidant enzyme activity. Our data show that in the adult Chinese perch muscle, starvation could reduce the antioxidant activity through the accumulation of ROS, and that the number of autophagosomes continues to increase. Refeeding after starvation could effectively compensate for the level of ROS, and restore the mRNA abundance of antioxidant genes and the activity of antioxidant enzymes to reduce autophagy and improve feed efficiency. Further research should optimize starvation conditions to reduce autophagy in muscles and maintain normal muscle metabolism.

Mandarin Fish (Siniperca chuatsi) p53 Regulates Glutaminolysis Induced by Virus via the p53/miR145-5p/c-Myc Pathway in Chinese Perch Brain Cells.

p53, as an important tumor suppressor protein, has recently been implicated in host antiviral defense. The present study found that the expression of mandarin fish (Siniperca chuatsi) p53 (Sc-p53) was negatively associated with infectious spleen and kidney necrosis virus (ISKNV) and Siniperca chuatsi rhabdovirus (SCRV) proliferation as well as the expression of glutaminase 1 (GLS1) and glutaminolysis pathway-related enzymes glutamate dehydrogenase (GDH) and isocitrate dehydrogenase 2 (IDH2). This indicated that Sc-p53 inhibited the replication and proliferation of ISKNV and SCRV by negatively regulating the glutaminolysis pathway. Moreover, it was confirmed that miR145-5p could inhibit c-Myc expression by targeting the 3' untranslated region (UTR). Sc-p53 could bind to the miR145-5p promoter region to promote its expression and to further inhibit the expression of c-Myc. The expression of c-Myc was proved to be positively correlated with the expression of GLS1 as well. All these suggested a negative relationship between the Sc-p53/miR145-5p/c-Myc pathway and GLS1 expression and glutaminolysis. However, it was found that after ISKNV and SCRV infection, the expressions of Sc-p53, miR145-5p, c-Myc, and GLS1 were all significantly upregulated, which did not match the pattern in normal cells. Based on the results, it was suggested that ISKNV and SCRV infection altered the Sc-p53/miR145-5p/c-Myc pathway. All of above results will provide potential targets for the development of new therapeutic strategies against ISKNV and SCRV. IMPORTANCE Infectious spleen and kidney necrosis virus (ISKNV) and Siniperca chuatsi rhabdovirus (SCRV) as major causative agents have caused a serious threat to the mandarin fish farming industry (J.-J. Tao, J.-F. Gui, and Q.-Y. Zhang, Aquaculture 262:1-9, 2007, https://doi.org/10.1016/j.aquaculture.2006.09.030). Viruses have evolved the strategy to shape host-cell metabolism for their replication (S. K. Thaker, J. Ch'ng, and H. R. Christofk, BMC Biol 17:59, 2019, https://doi.org/10.1186/s12915-019-0678-9). Our previous studies showed that ISKNV replication induced glutamine metabolism reprogramming and that glutaminolysis was required for efficient replication of ISKNV and SCRV. In the present study, the mechanistic link between the p53/miR145-5p/c-Myc pathway and glutaminolysis in the Chinese perch brain (CPB) cells was provided, which will provide novel insights into ISKNV and SCRV pathogenesis and antiviral treatment strategies.

The mTOR/PGC-1α/SIRT3 Pathway Drives Reductive Glutamine Metabolism to Reduce Oxidative Stress Caused by ISKNV in CPB Cells.

Under oxidative stress, viruses prefer glycolysis as an ATP source, and glutamine is required as an anaplerotic substrate to replenish the TCA cycle. Infectious spleen and kidney necrosis virus (ISKNV) induces reductive glutamine metabolism in the host cells. Here we report that ISKNV infection the increased NAD+/NADH ratio and the gene expression of glutaminase 1 (GLS1), glutamate dehydrogenase (GDH), and isocitrate dehydrogenase (IDH2) resulted in the phosphorylation and activation of mammalian target of rapamycin (mTOR) in CPB cells. Inhibition of mTOR signaling attenuates ISKNV-induced the upregulation of GLS1, GDH, and IDH2 genes expression, and exhibits significant antiviral activity. Moreover, the expression of silent information regulation 2 homolog 3 (SIRT3) in mRNA level is increased to enhance the reductive glutamine metabolism in ISKNV-infected cells. And those were verified by the expression levels of metabolic genes and the activities of metabolic enzymes in SIRT3-overexpressed or SIRT3-knocked down cells. Remarkably, activation of mTOR signaling upregulates the expression of the peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α) gene, leading to increased expression of SIRT3 and metabolic genes. These results indicate that mTOR signaling manipulates reductive glutamine metabolism in ISKNV-infected cells through PGC-1α-dependent regulation of SIRT3. Our findings reveal new insights on ISKNV-host interactions and will contribute new cellular targets to antiviral therapy. IMPORTANCE Infectious spleen and kidney necrosis virus (ISKNV) is the causative agent of farmed fish disease that has caused huge economic losses in fresh and marine fish aquaculture. The redox state of cells is shaped by virus into a favorable microenvironment for virus replication and proliferation. Our previous study demonstrated that ISKNV replication induced glutamine metabolism reprogramming, and it is necessary for the ISKNV multiplication. In this study, the mechanistic link between the mTOR/PGC-1α/SIRT3 pathway and reductive glutamine metabolism in the ISKNV-infected cells was provided, which will contribute new insights into the pathogenesis of ISKNV and antiviral treatment strategies.

Effects of dietary carbohydrate to lipid ratios on growth, biochemical indicators, lipid metabolism, and appetite in Chinese perch (Siniperca chuatsi).

An 8-week feeding trial was conducted to evaluate the effects of dietary carbohydrate to lipid (CHO:L) ratios on growth performance, body composition, serum biochemical indexes, lipid metabolism, and gene expression of central appetite regulating factors in Chinese perch (Siniperca chuatsi) (mean initial weight: 12.86 ± 0.10 g). Five isonitrogenous and isoenergetic diets (fish meal, casein as main protein sources) were formulated to contain different graded CHO:L ratio diets ranging from 0.12, 0.86, 1.71, 3.29, and 7.19. Each diet was assigned to triplicate groups of 18 experimental fish. Our results revealed that final body weight (FBW), weight gain rate (WGR), specific growth rate (SGR), and protein efficiency ratio (PER) increased with dietary CHO:L ratio from 0.12 to 1.71 and then decreased with further increases in dietary CHO:L ratio. A two-slope broken-line regression analysis based on WGR showed that the optimal dietary CHO:L level for maximum growth performance of fish was 1.60. Crude lipid and crude protein content in the liver and glycogen concentration in the muscle and liver were significantly influenced by the dietary CHO:L ratios (P < 0.05). The lowest crude lipid content in the liver was observed in fish fed the diet with a CHO:L ratio of 1.71(P < 0.05). Dietary CHO:L ratios significantly induced the glucose concentration of serum (P < 0.05). The relative expression levels of genes involved in lipid metabolism, such as srebp1 and fas in the liver, showed a trend of first decreased and then increased with the increase of dietary CHO:L ratio levels. Appropriate CHO:L ratio in the diet can effectively reduce the accumulation of liver fat. We observed in fish fed the 1.71 CHO:L ratio diet showed higher feed intake, up-regulated mRNA expression of neuropeptide Y (npy) and agouti gene-related protein (agrp), and down-regulated mRNA expression of cocaine- and amphetamine-regulated transcript (cart) and pro-opiomelanocorticoid (pomc) significantly as compared to control group. Thus, these results provide the theoretical basis for feed formulation to determine the appropriate CHO:L ratio requirement of Chinese perch.

Integrated transcriptomic and proteomic analysis of the physiological changes of the liver in domesticated Eurasian perch, Perca fluviatilis.

Artificial domestication during aquaculture practice has strongly shaped the physiological characteristics of Perca fluviatilis. Thus, revealing the genetic changes in domesticated P. fluviatilis will improve aquaculture and selective breeding. In this study, comparative analysis of the liver transcriptome, proteome, and physiological and biochemical indices of domesticated and wild P. fluviatilis was conducted. Our results indicated that the activity of lipase and the content of glucose were higher; however, the total antioxidant capacity and superoxide dismutase were lower in domesticated P. fluviatilis. Integrated analysis of "omics" data identified 174 and 127 genes and proteins that showed consistent upregulation and downregulation in domesticated P. fluviatilis, respectively. GO and KEGG enrichment of differentially expressed genes and proteins and the protein-protein interaction network indicated that energy metabolism (lipid and carbohydrate metabolism) was enhanced, and that signal transduction and the stress response were reduced in domesticated P. fluviatilis. This study revealed that artificial domestication may significantly shape the physiological changes in energy metabolism and stress resistance in domesticated P. fluviatilis, which makes them more adaptable to the artificial aquaculture environment, thereby promoting growth and development.

Comprehensive Characterization of Multitissue Expression Landscape, Co-Expression Networks and Positive Selection in Pikeperch.

Promising efforts are ongoing to extend genomics resources for pikeperch (Sander lucioperca), a species of high interest for the sustainable European aquaculture sector. Although previous work, including reference genome assembly, transcriptome sequence, and single-nucleotide polymorphism genotyping, added a great wealth of genomic tools, a comprehensive characterization of gene expression across major tissues in pikeperch still remains an unmet research need. Here, we used deep RNA-Sequencing of ten vital tissues collected in eight animals to build a high-confident and annotated trancriptome atlas, to detect the tissue-specificity of gene expression and co-expression network modules, and to investigate genome-wide selective signatures in the Percidae fish family. Pathway enrichment and protein-protein interaction network analyses were performed to characterize the unique biological functions of tissue-specific genes and co-expression modules. We detected strong functional correlations and similarities of tissues with respect to their expression patterns-but also significant differences in the complexity and composition of their transcriptomes. Moreover, functional analyses revealed that tissue-specific genes essentially play key roles in the specific physiological functions of the respective tissues. Identified network modules were also functionally coherent with tissues' main physiological functions. Although tissue specificity was not associated with positive selection, several genes under selection were found to be involved in hypoxia, immunity, and gene regulation processes, that are crucial for fish adaption and welfare. Overall, these new resources and insights will not only enhance the understanding of mechanisms of organ biology in pikeperch, but also complement the amount of genomic resources for this commercial species.

Twelve new microsatellite loci of Eurasian perch Perca fluviatilis Linnaeus, 1758.

The Eurasian perch (Perca fluviatilis Linnaeus, 1758) is native to almost entire Eurasia. For over the last two decades, this species became an important candidate for intensive freshwater aquaculture due to its high consumer's acceptance and overall market value. Hence, the intensive production of Eurasian perch has increased considerably allowing effective domestication; there is still a need for the development of effective selective breeding programmes allowing its further expansion. This process, in turn, can be significantly facilitated by molecular genetics. The genetic information of Eurasian perch and its populations is limited. Up to date information of regarding genetic diversity of many populations is still missing, including microsatellites for Eurasian perch, which could be useful during the selective breeding programmes allowing parental assignment and/or to follow heritability of desired traits. In this study, we have developed and characterized new polymorphic microsatellites. Subsequently, those 12 markers have been used further to compare two Hungarian and one Polish Eurasian perch populations. The Hungarian stocks had high genetic similarity (with low diversity), as we assumed, while the Polish population differed significantly. All populations deviated significantly from the Hardy-Weinberg equilibrium, and heterozygote deficiency was detected in all, showing the presence of an anthropogenic effect.

Proteomic profile and morphological characteristics of skeletal muscle from the fast- and slow-growing yellow perch (Perca flavescens).

The objective of the present study was to compare skeletal muscle proteomic profiles, histochemical characteristics, and expression levels of myogenic regulatory factors (MRFs) between fast- versus slow-growing yellow perch Perca flavescens and identify the proteins/peptides that might play a crucial role in the muscle growth dynamic. Yellow perch were nursed in ponds for 6 weeks from larval stage and cultured in two meter diameter tanks thereafter. The fingerlings were graded to select the top 10% and bottom 10% fish which represented fast- and slow-growing groups (31 yellow perch per each group). Our statistical analyses showed 18 proteins that had different staining intensities between fast- and slow-growing yellow perch. From those proteins 10 showed higher expression in slow-growers, and 8 demonstrated higher expression in fast-growers. Fast-growing yellow perch with a greater body weight was influenced by both the muscle fiber hypertrophy and mosaic hyperplasia compared to slow-growing fish. These hyperplastic and hypertrophic growth in fast-grower were associated with not only metabolic enzymes, including creatine kinase, glycogen phosphorylase, and aldolase, but also myoD and myogenin as MRFs. Overall, the results of the present study contribute to the identification of different expression patterns of gene products in fast- and slow-growing fish associated with their muscle growth.

Exposure via biotransformation: Oxazepam reaches predicted pharmacological effect levels in European perch after exposure to temazepam.

It is generally expected that biotransformation and excretion of pharmaceuticals occurs similarly in fish and mammals, despite significant physiological differences. Here, we exposed European perch (Perca fluviatilis) to the benzodiazepine drug temazepam at a nominal concentration of 2 µg L-1 for 10 days. We collected samples of blood plasma, muscle, and brain in a time-dependent manner to assess its bioconcentration, biotransformation, and elimination over another 10 days of depuration in clean water. We observed rapid pharmacokinetics of temazepam during both the exposure and depuration periods. The steady state was reached within 24 h of exposure in most individuals, as was complete elimination of temazepam from tissues during depuration. Further, the biologically active metabolite oxazepam was produced via fish biotransformation, and accumulated significantly throughout the exposure period. In contrast to human patients, where a negligible amount of oxazepam is created by temazepam biotransformation, we observed a continuous increase of oxazepam concentrations in all fish tissues throughout exposure. Indeed, oxazepam accumulated more than its parent compound, did not reach a steady state during the exposure period, and was not completely eliminated even after 10 days of depuration, highlighting the importance of considering environmental hazards posed by pharmaceutical metabolites.

Isolation and characterization of chitosan from Ugandan edible mushrooms, Nile perch scales and banana weevils for biomedical applications.

Of recent, immense attention has been given to chitosan in the biomedical field due to its valuable biochemical and physiological properties. Traditionally, the chief source of chitosan is chitin from crab and shrimp shells. Chitin is also an important component of fish scales, insects and fungal cell walls. Thus, the aim of this study was to isolate and characterize chitosan from locally available material for potential use in the biomedical field. Chitosan ash and nitrogen contents ranged from 1.55 to 3.5% and 6.6 to 7.0% respectively. Molecular weight varied from 291 to 348KDa. FTIR spectra revealed high degree of similarity between locally isolated chitosan and commercial chitosan with DD ranging from 77.8 to 79.1%. XRD patterns exhibited peaks at 2θ values of 19.5° for both mushroom and banana weevil chitosan while Nile perch scales chitosan registered 3 peaks at 2θ angles of 12.3°, 20.1° and 21.3° comparable to the established commercial chitosan XRD pattern. Locally isolated chitosan exhibited antimicrobial activity at a very high concentration. Ash content, moisture content, DD, FTIR spectra and XRD patterns revealed that chitosan isolated from locally available materials has physiochemical properties comparable to conventional chitosan and therefore it can be used in the biomedical field.

Thermal plasticity of the cardiorespiratory system provides cross-tolerance protection to fish exposed to elevated nitrate.

Exposure to nitrate is toxic to aquatic animals due to the formation of methaemoglobin and a subsequent loss of blood-oxygen carrying capacity. Yet, nitrate toxicity can be modulated by other stressors in the environment, such as elevated temperatures. Acclimation to elevated temperatures has been shown to offset the negative effects of nitrate on whole animal performance in fish, but the mechanisms underlying this cross-tolerance interaction remain unclear. In this study, juvenile silver perch (Bidyanus bidyanus) were exposed to a factorial combination of temperature (28 °C or 32 °C) and nitrate concentrations (0, 50 or 100 mg NO3- L-1) treatments to test the hypothesis that thermal acclimation offsets the effects of nitrate via compensatory changes to the cardiorespiratory system (gills, ventricle and blood oxygen carrying capacity). Following 21 weeks of thermal acclimation, we found that fish acclimated to 32 °C experienced an expansion of gill surface area and an increase in ventricular thickness regardless of nitrate exposure concentration. Exposure to nitrate (both 50 and 100 mg NO3- L-1) reduced the blood oxygen carrying capacity of silver perch due to increases in methaemoglobin concentration and a right shift in oxygen-haemoglobin binding curves in fish from both thermal acclimation treatments. These results indicate that plasticity of the gills and ventricle of warm acclimated fish are potential mechanisms which may provide cross-tolerance protection to elevated nitrate concentrations despite nitrate induced reductions to oxygen transport.

Multi-Level Responses of Yellow Perch (Perca flavescens) to a Whole-Lake Nanosilver Addition Study.

Silver nanoparticles (AgNP) are widely used as antibacterial agents in both commercial products and for industrial applications. As such, AgNP has a high potential for release into freshwater environments. As part of a whole-lake ecosystem experiment to examine the impacts of AgNP exposure at low µg/L concentrations over multiple years, we evaluated biological responses in Yellow Perch (Perca flavescens) before, during, and after AgNP additions to a freshwater lake. Yellow Perch were monitored for responses to in situ AgNP additions at the cellular (suite of biomarkers), individual (growth, prey consumption, and metabolism), and population (abundance and gross prey consumption) scales. At the cellular level, several biomarkers of oxidative stress in liver tissues revealed down-regulation, including decreased mRNA levels of catalase and glutathione peroxidase in Yellow Perch collected during AgNP exposure, and elevated ratios of reduced to oxidized glutathione. At the individual level, Yellow Perch bioenergetic models revealed that prey consumption and total metabolism significantly declined during AgNP additions and remained depressed one year after AgNP addition. At the population level, Yellow Perch densities and gross prey consumption declined after AgNP was added to the lake. Together, these results reveal a holistic assessment of the negative impacts of chronic exposure to environmentally relevant AgNP concentrations (i.e., µg/L) on Yellow Perch at cellular, individual, and population levels.

Most abundant metabolites in tissues of freshwater fish pike-perch (Sander lucioperca).

Quantitative metabolomic analysis was performed for eleven tissues of freshwater fish pike-perch (Sander lucioperca), including gill, heart, liver, kidney, spleen, muscle, brain, milt, lens, aqueous (AH) and vitreous (VH) humors with the use of NMR spectroscopy. The absolute values of concentrations were determined for more than 65 most abundant metabolites in every tissue. It was found that from the metabolomic viewpoint, kidney and gill are the most similar tissues, while the metabolomic compositions of ocular tissues-lens, AH, and VH significantly differ from that of other tissues. The combinations of intracellular osmolytes and antioxidants are specific for every tissue. In particular, the concentration of antioxidant ovothiol A in the lens is much higher than in any other tissue, while the brain enjoys the elevated level of ascorbate. The most abundant osmolyte in the fish spleen, muscle, and heart is taurine, and in the brain, gill, and lens-myo-inositol. Other important osmolytes specific for particular tissues are N-acetyl-histidine, N-acetyl-aspartate, betaine, threonine-phosphoethanolamine, and serine-phosphoethanolamine. The quantitative data obtained in the present work can be used as the baseline metabolite concentrations in the fish tissues to evaluate the influence of seasonal, ecological and other factors on the fish metabolism.