Unexposed Fish Research Articles

Mucosal organs exhibit distinct response signatures to hydrogen sulphide in Atlantic salmon (Salmo salar)

Hydrogen sulphide (H2S) is considered an immunotoxicant, and its presence in the water can influence the mucosal barrier functions of fish. However, there is a significant knowledge gap on how fish mucosa responds to low environmental H2S levels. The present study investigated the consequences of prolonged exposure to sub-lethal levels of H2S on the mucosal defences of Atlantic salmon (Salmo salar). Fish were continuously exposed to two levels of H2S (low: 0.05 µM; and high: 0.12 µM) for 12 days. Unexposed fish served as control. Molecular and histological profiling focused on the changes in the skin, gills and olfactory rosette. In addition, metabolomics and proteomics were performed on the skin and gill mucus. The gene expression profile indicated that the gills and olfactory rosette were more sensitive to H2S than the skin. The olfactory rosette showed a dose-dependent response, but not the gills. Genes related to stress responses were triggered at mucosal sites by H2S. Moreover, H2S elicited strong inflammatory responses, particularly in the gills. All mucosal organs demonstrated the key molecular repertoire for sulphide detoxification, but their temporal and spatial expression was not substantially affected by sub-lethal H2S levels. Mucosal barrier integrity was not considerably affected by H2S. Mucus metabolomes of the skin and gills were unaffected, but a matrix-dependent response was identified. Comparing the high-concentration group's skin and gills mucus metabolomes identified altered amino acid biosynthesis and metabolism pathways. The skin and gill mucus exhibited distinct proteomic profiles. Enrichment analysis revealed that proteins related to immunity and metabolism were affected in both mucus matrices. The present study expands our knowledge of the defence mechanisms against H2S at mucosal sites in Atlantic salmon. The findings offer insights into the health and welfare consequences of sub-lethal H2S, which can be incorporated into the risk assessment protocols in salmon land-based farms.

Multigenerational effects of alcohol: A behavioral study in three zebrafish populations

Fetal alcohol exposure can result in fetal alcohol spectrum disorder (FASD), which encompasses a range of cognitive and behavioral impairments. Although zebrafish have been used as a reliable model to study FASD, little is known about the ontogeny of this disorder and population differences in subsequent generations not directly exposed to alcohol. In this study, we evaluated the behavioral outcomes of zebrafish populations AB, Outbred (OB), and Tubingen (TU), offspring of parents exposed to alcohol during embryonic development. The offspring of adult fish with FASD (exposed to 1 % alcohol at the embryonic stage) was compared to the offspring of unexposed parental fish (0 % alcohol at the embryo phase). The behavioral profile of the offspring was assessed at 6 days post-fertilization (dpf) and 45 dpf. At 6dpf, the AB FASD offspring exhibited hyperactivity and increased time at the edge of the tank, while the TU and OB FASD offspring showed hypoactivity. At 45dpf, TU fish maintained the larval locomotor pattern, characterized by decreased average speed and total distance traveled and increased immobility. However, AB and OB fish did not show alterations in locomotor activity and anxiety-related responses at 45dpf. Our results demonstrate, for the first time, that FASD zebrafish offspring display behavioral differences, which were most evident during the early ontogenetic phase (6dpf) but may vary throughout animal ontogeny. TU fish exhibited the most consistent behavioral pattern across different developmental stages. These findings provide insights into the multigenerational and persistent behavioral consequences of embryonic alcohol exposure in zebrafish. Further research should focus on other features that can be inherited and the development of treatments for the offspring affected by it.

Impacts of caffeine on fathead minnow behaviour and physiology

Pollution from regularly used substances such as pharmaceuticals, cleaning agents, and even food and beverages is an increasing problem in the environment. Caffeine, a commonly ingested stimulant, is one such contaminant that has been detected in aquatic environments worldwide. Yet, little is known about how ecologically relevant concentrations of caffeine influence the morphology, behaviour, and physiology of exposed organisms. To address this knowledge gap, we exposed fathead minnow (Pimephales promelas) to three caffeine treatments: a freshwater control (nominal: 0 ng/L), a low (nominal: 1,000 ng/L) and high environmentally relevant dose (nominal: 10,000 ng/L), for 35 days. We tested the learning abilities, anxiety, metabolic rates, and morphological features of exposed vs. control fish. Caffeine exposure did not affect the ability of fish to learn but did influence anxiety levels. Over the course of repeated anxiety testing, unexposed control fish visited a black square more often while fish exposed to low levels of caffeine did not, potentially indicating that these fish remained in a more anxious state. While caffeine did not impact metabolism, fish growth, or body size, it was associated with lower liver investment—although this response was only observed in our low caffeine treatment. Overall, our results suggest that even relatively low concentrations of caffeine may impact the liver size and anxiety of exposed fish, but further research is needed to assess how extended exposure to caffeine impacts fitness. Given the increase in anthropogenic contaminants in aquatic environments, it is important that we continue to investigate their effects on the organisms exposed to them.

Segmentation of Renal Thyroid Follicle Colloid in Common Carp: Insights into Perfluorooctanoic Acid-Induced Morphometric Alterations.

Perfluorooctanoic acid (PFOA) is a globally prevalent contaminant of concern recognised for its persistence and detrimental effects on both wildlife and humans. While PFOA has been established as a disruptor of thyroid function, limited data exist regarding its impact on thyroid morphology. The kidney of the common carp (Cyprinus carpio) harbours numerous thyroid follicles, rendering it a valuable biomarker organ for investigating PFOA-induced thyroid alterations. Renal tissue slides, stained with the Alcian blue/PAS method, were examined from carp in three experimental groups: unexposed, exposed to 200 ng L-1, and exposed to 2 mg L-1 of PFOA over 56 days. Thyroid follicle colloids were segmented, and related morphometric parameters, including perimeter, area, and shape descriptors, were obtained. Statistical analyses revealed significant reductions in thyroid follicle colloid perimeter and area in the 200 ng L-1 PFOA group compared to the unexposed and 2 mg L-1 PFOA groups. Additionally, the fish exposed to PFOA exhibited a significantly higher follicle count compared to the unexposed fish. These findings collectively suggest that PFOA induces thyroid folliculogenesis, emphasising its impact on thyroid morphology even at an environmentally relevant concentration (200 ng L-1).

Effects of water quality on fish parasite biodiversity and physiological responses in the host fish Clarias gariepinus from a eutrophic lake subjected to acid mine drainage in South Africa.

Elevated concentrations of contaminants have negative impacts on aquatic organisms and their parasites. Changes in parasite infections have been proposed as a technique for monitoring the health of aquatic ecosystems. Furthermore, alterations in physiological responses (biomarkers) of organisms have also been used to delineate ecosystem quality. Lake Heritage is situated along the Crocodile River in Muldersdrift, Gauteng, South Africa, and is subject to contamination by acid mine drainage. Clarias gariepinus is a well-studied bioindicator species and host to numerous endoparasites and ectoparasites. The aims of this study were to delineate the health status of Lake Heritage through a multifaceted approach by comparing the water quality, biomarker responses, and parasite biodiversity of C. gariepinus, compared to unexposed laboratory-reared fish. Physical and chemical water quality parameters were determined using a hand-held probe, test kits, and element analysis with inductively coupled plasma-mass spectrometry. Biomarker responses in the gill, liver, and muscle tissues from C. gariepinus were assessed for total protein, metallothioneins, superoxide dismutase (SOD), and reduced glutathione (GSH) concentrations and activities of acetylcholinesterase and catalase. Results for water quality variables showed higher pH, nitrate, hardness, and copper levels compared with the South African Target Water Quality Guidelines. Catalase activity and concentrations of SOD and reduced GSH showed a response in C. gariepinus to the water quality. Ectoparasites had lower prevalence and mean intensity than endoparasites. However, there were no differences in the physiological responses between infected and uninfected hosts. The study shows that the eutrophic conditions in Lake Heritage cause biomarker responses in the host when compared to host fish in laboratory conditions. Integr Environ Assess Manag 2024;20:1539-1553. © 2023 The Authors. Integrated Environmental Assessment and Management published by Wiley Periodicals LLC on behalf of Society of Environmental Toxicology & Chemistry (SETAC).

Coinfection of channel catfish (Ictalurus punctatus) with virulent Aeromonas hydrophila and Flavobacterium covae exacerbates mortality.

Flavobacterium covae and virulent Aeromonas hydrophila are prevalent bacterial pathogens within the US catfish industry that can cause high mortality in production ponds. An assessment of invivo bacterial coinfection with virulent A. hydrophila (ML09-119) and F. covae (ALG-00-530) was conducted in juvenile channel catfish (Ictalurus punctatus). Catfish were divided into seven treatments: (1) mock control; (2) and (3) high and low doses of virulent A. hydrophila; (4) and (5) high and low doses of F. covae; (6) and (7) simultaneous challenge with high and low doses of virulent A. hydrophila and F. covae. In addition to the mortality assessment, anterior kidney and spleen were collected to evaluate immune gene expression, as well as quantify bacterial load by qPCR. At 96 h post-challenge (hpc), the high dose of virulent A. hydrophila infection (immersed in 2.3 × 107 CFU mL-1 ) resulted in cumulative percent mortality (CPM) of 28.3 ± 9.5%, while the high dose of F. covae (immersed in 5.2 × 106 CFU mL-1 ) yielded CPM of 23.3 ± 12.9%. When these pathogens were delivered in combination, CPM significantly increased for both the high- (98.3 ± 1.36%) and low-dose combinations (76.7 ± 17.05%) (p < .001). Lysozyme activity was found to be different at 24 and 48 hpc, with the high-dose vAh group demonstrating greater levels than unexposed control fish at each time point. Three proinflammatory cytokines (tnfα, il8, il1b) demonstrated increased expression levels at 48 hpc. These results demonstrate the additive effects on mortality when these two pathogens are combined. The synthesis of these mortality and health metrics advances our understanding of coinfections of these two important catfish pathogens and will aid fish health diagnosticians and channel catfish producers in developing therapeutants and prevention methods to control bacterial coinfections.

Experimental arena size alters larval zebrafish photolocomotor behaviors and influences bioactivity responses to a model neurostimulant

Zebrafish behavior is increasingly common in biomedical and environmental studies of chemical bioactivity. Multiple experimental arena sizes have been used to measure photolocomotion in zebrafish depending on age, endpoints observed, and instrumentation, among other factors. However, the extent to which methodological parameters may influence naïve behavioral performance and detection of behavioral changes is poorly understood. Here we measured photolocomotion and behavioral profiles of naïve larval zebrafish across arena sizes. We then performed concentration response studies with the model neurostimulant caffeine, again across various arena dimensions. We found total swimming distance of unexposed fish to increase logarithmically with arena size, which as related to circumference, area, and volume. Photomotor response during light/dark transitions also increased with arena size. Following caffeine exposure, total distance travelled was significantly (p < 0.001) affected by well size, caffeine treatment (p < 0.001), and the interaction of these two experimental factors (p < 0.001). In addition, behavioral response profiles showed differences between 96 well plates and larger well sizes. Biphasic response, with stimulation at lower concentrations and refraction at the highest concentration, was observed in dark conditions for the 96 well size only, though almost no effects were identified in the light. However, swimming behavior was significantly (p < 0.1) altered in the highest studied caffeine treatment level in larger well sizes during both light and dark periods. Our results indicate zebrafish swim more in larger arenas and arena size influences behavioral response profiles to caffeine, though differences were mostly observed between very small and large arenas. Further, careful consideration should be given when choosing arena size, because small wells may lead to restriction, while larger wells may differentially reflect biologically relevant effects. These findings can improve comparability among experimental designs and demonstrates the importance of understanding confounding methodological variables.

Temperature impacts Atlantic salmon's (Salmo salar) immunological response to infectious salmon anemia virus (ISAv).

Ocean temperatures continue to rise annually due to the ever-growing consequences of global climate change. These temperature changes can have an impact on the immunological robustness of cultured fish, especially cold-water species such as Atlantic salmon. The salmon farming industry already loses hundreds of millions of dollars each year to infectious and non-infectious diseases. One particularly important and WOAH reportable disease is infectious salmon anemia caused by the orthomyxovirus ISAv. Considering the changing environment, it is necessary to find ways to mitigate the effect of diseases on the industry. For this study, 20 Atlantic salmon families were housed in each of 38 different tanks at the AVC, with half of the fish being kept at 10 °C and half being kept at 20 °C. Donor Atlantic salmon IP- injected with a highly virulent ISAv isolate (HPR4; TCID50 of 1 × 105/mL) were added to each tank as the source of co-habitation infection. Both temperatures were sampled at onset of mortality in co-habited fish and at resolution of mortality. Family background and temperature significantly impacted ISAv load, as assessed by qPCR, time to mortality and overall mortality. Mortality was more acute at 20 °C, but overall mortality was higher at 10 °C. Based on percent mortality calculated over the course of the study, different families demonstrated different levels of survival. The three families that demonstrated the highest percent mortality, and the three families with the lowest percent mortality were then assessed for their antiviral responses using relative gene expression. Genes significantly upregulated between the unexposed fish and ISAv exposed fish included mx1, il4/13a, il12rb2, and trim25, and these were further impacted by temperature. Understanding how ISAv resistance is impacted by temperature can help identify seasonal risks of ISAv outbreaks as well as ideal responses to be targeted through immunopotentiation.

Widespread psychoactive pollutant augments daytime restfulness and disrupts diurnal activity rhythms in fish

Pharmaceutical pollution is a major driver of global change, with the capacity to alter key behavioural and physiological traits in exposed animals. Antidepressants are among the most commonly detected pharmaceuticals in the environment. Despite well-documented pharmacological effects of antidepressants on sleep in humans and other vertebrates, very little is known about their ecologically relevant impacts as pollutants on non-target wildlife. Accordingly, we investigated the effects of acute 3-day exposure of eastern mosquitofish (Gambusia holbrooki) to field-realistic levels (nominal concentrations: 30 and 300 ng/L) of the widespread psychoactive pollutant, fluoxetine, on diurnal activity patterns and restfulness, as indicators of disruptions to sleep. We show that exposure to fluoxetine disrupted diel activity patterns, which was driven by augmentation of daytime inactivity. Specifically, unexposed control fish were markedly diurnal, swimming farther during the day and exhibiting longer periods and more bouts of inactivity at night. However, in fluoxetine-exposed fish, this natural diel rhythm was eroded, with no differences in activity or restfulness observed between the day and night. As a misalignment in the circadian rhythm has been shown to adversely affect fecundity and lifespan in animals, our findings reveal a potentially serious threat to the survival and reproductive success of pollutant-exposed wildlife.

Slow-Release Lanthanum Effectively Reduces Phosphate in Eutrophic Ponds without Accumulating in Fish

Nutrient runoff is a major water quality issue affecting water resources. Excess nutrients such as nitrate (NO3−) and phosphate (PO43−) entering surface waters promote eutrophication. Recent research showed that floating treatment wetlands combined with slow-release lanthanum composites deployed through airlift pumps can reduce NO3− and PO43− concentrations, minimize algae and weeds, and increase dissolved oxygen concentrations. While water quality improves following this biological and chemical approach, questions remain about the toxicity and potential accumulation of lanthanum in lentic organisms. We addressed this concern by analyzing flesh and liver of fish exposed to the slow-release lanthanum following two years of treatment and compared results to fish harvested from a control, untreated pond. We also conducted an aquarium fish study that used higher lanthanum concentrations than those observed in the field. The field study confirmed that under the concentrations of lanthanum released to treat eutrophic ponds (109 µg L−1), no adverse effects were observed in harvested fish. We also observed no significant differences between lanthanum-exposed and -unexposed fish (α = 0.05) in our controlled tank study. Given the laboratory tank lanthanum concentrations were approximately nine times higher (916 µg L−1) than the observed field concentrations, we conclude the slow-release lanthanum composites used to treat eutrophic ponds are effective in improving water quality and do not lead to significant lanthanum accumulation in fish.

Parasite infection but not chronic microplastic exposure reduces the feeding rate in a freshwater fish

Microplastics (plastics <5 mm) are an environmental contaminant that can negatively impact the behaviour and physiology of aquatic biota. Although parasite infection can also alter the behaviour and physiology of their hosts, few studies have investigated how microplastic and parasite exposure interact to affect hosts. Accordingly, an interaction experiment tested how exposure to environmentally relevant microplastic concentrations and the trophically transmitted parasite Pomphorhynchus tereticollis affected the parasite load, condition metrics and feeding rate of the freshwater fish final host chub Squalius cephalus. Microplastic exposure was predicted to increase infection susceptibility, resulting in increased parasite loads, whereas parasite and microplastic exposure were expected to synergistically and negatively impact condition indices and feeding rates. Following chronic (≈170 day) dietary microplastic exposure, fish were exposed to a given number of gammarids (4/8/12/16/20), with half of the fish presented with parasite infected individuals, before a comparative functional response experiment tested differences in feeding rates on different live prey densities. Contrary to predictions, dietary microplastic exposure did not affect parasite abundance at different levels of parasite exposure, specific growth rate was the only condition index that was lower for exposed but unexposed fish, with no single or interactive effects of microplastic exposure detected. However, parasite infected fish had significantly lower feeding rates than unexposed fish in the functional response experiment, with exposed but unexposed fish also showing an intermediate decrease in feeding rates. Thus, the effects of parasitism on individuals were considerably stronger than microplastic exposure, with no evidence of interactive effects. Impacts of environmentally relevant microplastic levels might thus be relatively minor versus other stressors, with their interactive effects difficult to predict based on their single effects.

Mucosal immune and stress responses of Neoparamoeba perurans-infected Atlantic salmon (Salmo salar) treated with peracetic acid shed light on the host-parasite-oxidant interactions.

Treatment development for parasitic infestation is often limited to disease resolution as an endpoint response, and physiological and immunological consequences are not thoroughly considered. Here, we report the impact of exposing Atlantic salmon affected with amoebic gill disease (AGD) to peracetic acid (PAA), an oxidative chemotherapeutic. AGD-affected fish were treated with PAA either by exposing them to 5 ppm for 30 min or 10 ppm for 15 min. Unexposed fish from both infected and uninfected groups were also included. Samples for molecular, biochemical, and histological evaluations were collected at 24 h, 2 weeks, and 4 weeks post-treatment. Behavioral changes were observed during PAA exposure, and post-treatment mortality was higher in the infected and PAA treated groups, especially in 10 ppm for 15 min. Plasma indicators showed that liver health was affected by AGD, though PAA treatment did not exacerbate the infection-related changes. Transcriptome profiling in the gills showed significant changes, triggered by AGD and PAA treatments, and the effects of PAA were more notable 24 h after treatment. Genes related to immune pathways of B- and T- cells and protein synthesis and metabolism were downregulated, where the magnitude was more remarkable in 10 ppm for 15 min group. Even though treatment did not fully resolve the pathologies associated with AGD, 5 ppm for 30 min group showed lower parasite load at 4 weeks post-treatment. Mucous cell parameters (i.e., size and density) increased within 24 h post-treatment and were significantly higher at termination, especially in AGD-affected fish, with some treatment effects influenced by the dose of PAA. Infection and treatments resulted in oxidative stress—in the early phase in the gill mucosa, while systemic reactive oxygen species (ROS) dysregulation was evident at the later stage. Infected fish responded to elevated circulating ROS by increasing antioxidant production. Exposing the fish to a crowding stress revealed the interference in the post-stress responses. Lower cortisol response was displayed by AGD-affected groups. Collectively, the study established that PAA, within the evaluated treatment protocols, could not provide a convincing treatment resolution and, thus, requires further optimization. Nonetheless, PAA treatment altered the mucosal immune and stress responses of AGD-affected Atlantic salmon, shedding light on the host-parasite-treatment interactions.

Nrf2a dependent and independent effects of early life exposure to 3,3'-dichlorobiphenyl (PCB-11) in zebrafish (Danio rerio).

The environmental pollutant 3,3'-dichlorobiphenyl (PCB-11) is a lower-chlorinated polychlorinated biphenyl (PCB) congener present in air and water samples. Both PCB-11 and its metabolite, 4-PCB-11-Sulfate, are detected in humans, including in pregnant women. Previous research in zebrafish (Danio rerio) has shown that 0.2μM exposures to 4-PCB-11-Sulfate starting at 1 day post fertilization (dpf) increase hepatic neutral lipid accumulation in larvae at 15 dpf. Here, we explored whether nuclear factor erythroid 2-related factor 2 (Nrf2), known as the master-regulator of the adaptive response to oxidative stress, contributes to metabolic impacts of 4-PCB-11-Sulfate. For this work, embryos were collected from homozygous wildtype or Nrf2a mutant adult zebrafish that also express GFP in pancreatic β-cells, rendering Tg(ins:GFP;nrf2afh318+/+) and Tg(ins:GFP;nrf2afh318-/-) lines. Exposures were conducted from 1-15 dpf to either 0.05% DMSO or DMSO-matched 0.2µM 4-PCB-11-Sulfate, and at 15 dpf subsets of larvae were imaged for overall morphology, primary pancreatic islet area, and collected for fatty acid profiling and RNAseq. At 15 dpf, independent of genotype, fish exposed to 4-PCB-11-Sulfate survived significantly more at 80-85% compared to 65-73% survival for unexposed fish, and had primary pancreatic islets 8% larger compared to unexposed fish. Fish growth at 15 dpf was dependent on genotype, with Nrf2a mutant fish a significant 3-5% shorter than wildtype fish, and an interaction effect was observed where Nrf2a mutant fish exposed to 4-PCB-11-Sulfate experienced a significant 29% decrease in the omega-3 fatty acid DHA compared to unexposed mutant fish. RNAseq revealed 308 differentially expressed genes, most of which were dependent on genotype. These findings suggest that Nrf2a plays an important role in growth as well as for DHA production in the presence of 4-PCB-11-Sulfate. Further research would be beneficial to understand the importance of Nrf2a throughout the lifecourse, especially in the context of toxicant exposures.

Evaluating Phenotypic and Transcriptomic Responses Induced by Low-Level VOCs in Zebrafish: Benzene as an Example.

Urban environments are plagued by complex mixtures of anthropogenic volatile organic compounds (VOCs), such as mixtures of benzene, toluene, ethylene, and xylene (BTEX). Sources of BTEX that drive human exposure include vehicle exhaust, industrial emissions, off-gassing of building material, as well as oil spillage and leakage. Among the BTEX mixture, benzene is the most volatile compound and has been linked to numerous adverse health outcomes. However, few studies have focused on the effects of low-level benzene on exposure during early development, which is a susceptible window when hematological, immune, metabolic, and detoxification systems are immature. In this study, we used zebrafish to conduct a VOC exposure model and evaluated phenotypic and transcriptomic responses following 0.1 and 1 ppm benzene exposure during the first five days of embryogenesis (n = 740 per treatment). The benzene body burden was 2 mg/kg in 1 ppm-exposed larval zebrafish pools and under the detection limit in 0.1 ppm-exposed fish. No observable phenotypic changes were found in both larvae except for significant skeletal deformities in 0.1 ppm-exposed fish (p = 0.01) compared with unexposed fish. Based on transcriptomic responses, 1 ppm benzene dysregulated genes that were implicated with the development of hematological system, and the regulation of oxidative stress response, fatty acid metabolism, immune system, and inflammatory response, including apob, nfkbiaa, serpinf1, foxa1, cyp2k6, and cyp2n13 from the cytochrome P450 gene family. Key genes including pik3c2b, pltp, and chia.2 were differentially expressed in both 1 and 0.1 ppm exposures. However, fewer transcriptomic changes were induced by 0.1 ppm compared with 1 ppm. Future studies are needed to determine if these transcriptomic responses during embryogenesis have long-term consequences at levels equal to or lower than 1 ppm.

Desorption of polycyclic aromatic hydrocarbons from polyethylene microplastics in two morphologically different digestive tracts of marine teleosts: Gastric red seabream (Pagrus major) and agastric mummichog (Fundulus heteroclitus)

In this study, we elucidated the desorption potency of polycyclic aromatic hydrocarbons (PAHs) sorbed on microplastics (MP; polyethylene) in the digestive tract of two fish species: gastric red seabream and agastric mummichog. In our in vitro assay system using the real gut sample of unexposed fish, the digestive tract was firstly removed from the fish and divided into three parts. Then, MP that had previously been sorbed with 16 PAHs were incubated with extracts of the gut contents or tissue with buffer or only a buffer. The desorption potency of PAHs was individually assessed for gut contents and tissue, which revealed that PAH desorption from MP was elevated in extracts of the gut contents compared with that in the buffer alone for both fish species. PAH desorption potency was the highest in the midgut for gastric red seabream and in the foregut for agastric mummichog, which indicates that PAH desorption from MP varies among different parts of the digestive tract and among fish with distinct gut morphology. In the midgut contents of red seabream and foregut contents of mummichog, the desorption fraction was 5.6% and 8.1% of the total PAHs sorbed on MP, respectively. The desorption fraction enhancement achieved by adding gut contents extracts tended to be greater with an increase in the n-octanol/water partition ratio, suggesting that enhancement of the desorption fraction in the digestive tract depends on the physicochemical properties of PAHs. Thus, morphological differences in digestive tracts and PAH properties should be considered when evaluating the effect of MP vector on pollutant exposure in fish.

Saponin and chitosan-based oral vaccine against viral haemorrhagic septicaemia virus (VHSV) provides protective immunity in olive flounder (Paralichthys olivaceus)

Production losses of olive flounder (Paralichthys olivaceus) have increased owing to viral haemorrhagic septicaemia virus (VHSV) infection. In this study, we determined safe concentrations of orally administered saponin and chitosan by analysing serum enzyme (AST/ALT) levels as biochemical markers of hepatic injury. Furthermore, we demonstrated the efficacy, duration of protection, and safety of saponin and chitosan-based vaccines with inactivated VHSV (IV). Oral administration of saponin, chitosan, and their combination did not induce fish mortality at all tested concentrations (0.29, 1.45, and 2.9 mg/g of fish body weight/day) 10 days after administration. However, AST level was high at a dose >0.29 mg/g of fish body weight/day. Both saponin and chitosan were found to be safe and acceptable for vaccination studies at a dose of 0.29 mg/g of fish body weight/day. Administration of IV alone did not induce protection at 2 and 4 weeks post vaccination (wpv). Olive flounders administered saponin + IV and chitosan + IV vaccines had higher immunity against VHSV with relative percentage survival (RPS) of 12.5–7.5% and 0–20.1%, respectively; however, additional immunisation with combination of saponin + chitosan + IV clearly enhanced the protection with RPS values of 10–15%, 26.7%, 42.9%, and 37.5% at 4, 8, 12, and 20 wpv, respectively. Although the RPS value of oral immunisation was not comparable to that of injectable vaccines, the manufacturing process is simple and oral administration causes less stress to juvenile fish. To investigate the development of a protective immune response, olive flounder were re-challenged with VHSV (107.8 TCID50/fish) at 70 days postinfection; 100% of the previously unexposed fish died, whereas 80–100% of the previously immunised fish survived. Our results showed the possibility of developing preventive measures against VHSV using saponin and chitosan-based oral vaccines with inactivated virus.

Evaluation of UV-B protective properties of leaves and seeds of Achyranthes aspera in Asian catfish Clarias batrachus (Linn.).

The ultraviolet-B (UV-B) radiation is harmful to the aquatic organisms. The UV-B protective properties of leaves and seeds of herb Achyranthes aspera were evaluated in Clarias batrachus. Fish were fed with four diets-EFL1, EFL2 containing 0.25 and 0.5% leaves, EFS containing 0.5% seeds and control, CF. After 83days of feeding, fish were exposed to UV-B (157µW/cm2) for 7days at the rate of 15min/day. One batch of fish in each treatment was kept unexposed. Significantly higher final weight was found in EFS followed by EFL2 and EFL1 treatments. It was higher in unexposed fish compared to the exposed ones. Among exposed fish, significantly higher lysozyme was found in EFS and myeloperoxidase in EFS and EFL2 compared to others. Nitric oxide synthase and superoxide dismutase levels were significantly higher in liver and head kidney of EFS diet fed fish compared to others. Thiobarbituric acid reactive substances (TBARS) and carbonyl protein levels were minimum in EFS followed by EFL2. The independent sample t-test showed that nitric oxide synthase was significantly higher and myeloperoxidase and TBARS levels were significantly lower in unexposed group compared to the exposed fish in respective treatment. There were up-regulations of TNF-α, iNOS, NF-kB, BAX, Cytochrome c, SOD-c, Caspase 3, Caspase 9, BCL2 in liver and head kidney of leaves and seeds incorporated diets fed fish compared to control. Supplementation of A. aspera seeds and leaves at 0.5% level in diets gave UV-B protection to the fish.

Effects of ethyl acetate extract of Moringa oleifera leaf and vitamin c co-exposures on acute cadmium chloride-induced toxicopathological changes in Clarias gariepinus

Cadmium (Cd2+) is reportedly one of the most toxic metals, which is increasingly polluting our aquatic environments. The need to modify Cd2+ toxicity led to the investigation of the possible effects of ethyl acetate extract of Moringa oleifera (M. oleifera) leaf (EAEMOL) and vitamin C on cadmium chloride (CdCl2) co-exposures of adult Clarias gariepinus over a 96-hour period. The exposure involved 105 fish in five groups of seven fish each in triplicates. Group I contained the control unexposed fish, Group II contained Tween 80, and Groups III-V contained CdCl2, EAEMOL co-exposure, and the vitamin C co-exposure, respectively. A total of 52.38%, 28.57%, and 47.62% mortalities occurred in Group III, IV, and V exposed fish, respectively, with marked deterioration in fish water quality over the exposure period. Although CdCl2 exposure caused considerable lipid peroxidation, the EAEMOL and vitamin C co-exposures insignificantly (p>0.05) aggravated it rather than ameliorate it. Similarly, the co-exposures slightly elevated the recorded drop in liver metallothionein (MT) level induced by the CdCl2 exposure. Although CdCl2 exposure caused severe gill damage, only moderate gill damages occurred in both co-exposures based on the degree of tissue change protocol. Therefore, the EAEMOL and vitamin C co-exposures, as administered, ameliorated the CdCl2 toxicity in the exposed fish based on the recorded mortality and gills cellular damage as the induced biochemical changes were insignificant (p >0.05). However, the rationale behind the recorded slightly elevated serum lipid peroxidation and liver MT levels by the co-exposures need further elucidation.

Acetochlor Affects Bighead Carp (Aristichthys Nobilis) by Producing Oxidative Stress, Lowering Tissue Proteins, and Inducing Genotoxicity.

Acetochlor is persistently used in the agroproduction sector to control broadleaf weeds. Due to frequent and continuous applications, this herbicide can reach nearby water bodies and may induce deleterious changes in aquatic life. Therefore, investigation of harmful impacts of different environmental pollutants, including herbicides, is vital to knowing the mechanisms of toxicity and devising control strategies. The current experiment included bighead carp (n = 80) to estimate adverse impacts. Fish were randomly placed in 4 different experimental groups (T0-T3) and were treated for 36 days with acetochlor at 0, 300, 400, and 500 μg/L. Fresh blood without any anticoagulant was obtained and processed for nuclear and morphological changes in erythrocytes. At the same time, various visceral organs, including the gills, liver, brain, and kidneys, were removed and processed on days 12, 24, and 36 to determine oxidative stress and various antioxidant biomarkers. Comet assays revealed significantly increased DNA damage in isolated cells of the liver, kidneys, brain, and gills of treated fish. We recorded increased morphological and nuclear changes (P ≤ 0.05) in the erythrocyte of treated fish. The results on oxidative stress showed a higher quantity of oxidative biomarkers and a significantly (P ≤ 0.05) low concentration of cellular proteins in the gills, liver, brain, and kidneys of treated fish compared to unexposed fish. Our research findings concluded that acetochlor renders oxidative stress in bighead carp.

Multiomics Provide Insights into the Key Molecules and Pathways Involved in the Physiological Adaptation of Atlantic Salmon (Salmo salar) to Chemotherapeutic-Induced Oxidative Stress

Although chemotherapeutics are used to treat infections in farmed fish, knowledge on how they alter host physiology is limited. Here, we elucidated the physiological consequences of repeated exposure to the potent oxidative chemotherapeutic peracetic acid (PAA) in Atlantic salmon (Salmo salar) smolts. Fish were exposed to the oxidant for 15 (short exposure) or 30 (long exposure) minutes every 15 days over 45 days. Unexposed fish served as the control. Thereafter, the ability of the remaining fish to handle a secondary stressor was investigated. Periodic chemotherapeutic exposure did not affect production performance, though survival was lower in the PAA-treated groups than in the control. Increased ventilation, erratic swimming, and a loss of balance were common behavioural manifestations during the oxidant exposure. The plasma reactive oxygen species levels increased in the PAA-treated groups, particularly after the third exposure, suggesting an alteration in the systemic oxidative stress status. Plasma indicators for internal organ health were affected to a certain degree, with the changes mainly observed after the second and third exposures. Metabolomics disclosed that the oxidant altered several circulating metabolites. Inosine and guanosine were the two metabolites significantly affected by the oxidative stressor, regardless of exposure time. A microarray analysis revealed that the gills and liver were more responsive to the oxidant than the skin, with the gills being the most sensitive. Moreover, the magnitude of the transcriptomic modifications depended on the exposure duration. A functional analysis showed that genes involved in immunity and ribosomal functions were significantly affected in the gills. In contrast, genes crucial for the oxidation-reduction process were mainly targeted in the liver. Skin mucus proteomics uncovered that the changes in the mucosal proteome were dependent on exposure duration and that the oxidant interfered with ribosome-related processes. Mucosal mapping revealed gill mucous cell hypertrophy after the second and third exposures, although the skin morphological parameters remained unaltered. Lastly, repeated oxidant exposures did not impede the ability of the fish to mount a response to a secondary stressor. This study provides insights into how a chemical oxidative stressor alters salmon physiology at both the systemic and mucosal levels. This knowledge will be pivotal in developing an evidence-driven approach to the use of oxidative therapeutics in fish, with some of the molecules and pathways identified as potential biomarkers and targets for assessing the physiological cost of these treatments.