Chronic cypermethrin induced toxicity and molecular fate assessment within common carp (Cyprinus carpio) using multiple biomarkers approach and its novel therapeutic detoxification.

Cypermethrin (CYP) is a chemical of emerging concern which has persistent and bioaccumulating impacts as it can be found extensively in freshwater ecosystem and agricultural products. It has exposure risk and toxic effects over human edible fish, as common carp. Four groups were designed for toxicity assessment and detoxification approach: control group (CL), CYP exposure group (CYP), CYP + 10% M. oleifera leaves and 10% M. oleifera seeds (CMO group), 10% M. oleifera leaves and 10% M. oleifera seeds (MO group). Trial period was forty days during which cohort of 240 fish in CYP and CMO group was exposed to 1/5 of 96h LC50 of CYP (0.1612 µg/L). CYP-exposed carp exhibited lower growth parameters, but carp fed with 10% M. oleifera seeds and leaves showed significant improvement in growth rate (SGR, RGR) and weight gain (WG) as compared to the control group. CYP exposure negatively affected haemato-biochemical parameters. Moreover, CYP exposure also led to oxidative stress, damaged immunological parameters, genotoxicity and histopathological damage in liver and intestinal cells. Whereas, M. oleifera supplementation has ameliorated these conditions. Thereby, supplementation with M. oleifera is potential and novel therapeutic detoxication approach for common carp and human health against persistent and bioaccumulating emerging chemicals.

Dietary sodium acetate and sodium butyrate improve high-carbohydrate diet utilization by regulating gut microbiota, liver lipid metabolism, oxidative stress, and inflammation in largemouth bass (Micropterus salmoides).

BACKGROUND: Adequate level of carbohydrates in aquafeeds help to conserve protein and reduce cost. However, studies have indicated that high-carbohydrate (HC) diet disrupt the homeostasis of the gut-liver axis in largemouth bass, resulting in decreased intestinal acetate and butyrate level. METHOD: Herein, we had concepted a set of feeding experiment to assess the effects of dietary sodium acetate (SA) and sodium butyrate (SB) on liver health and the intestinal microbiota in largemouth bass fed an HC diet. The experimental design comprised 5 isonitrogenous and isolipidic diets, including LC (9% starch), HC (18% starch), HCSA (18% starch; 2 g/kg SA), HCSB (18% starch; 2 g/kg SB), and HCSASB (18% starch; 1 g/kg SA + 1 g/kg SB). Juvenile largemouth bass with an initial body weight of 7.00 ± 0.20 g were fed on these diets for 56 d. RESULTS: We found that dietary SA and SB reduced hepatic triglyceride accumulation by activating autophagy (ATG101, LC3B and TFEB), promoting lipolysis (CPT1α, HSL and AMPKα), and inhibiting adipogenesis (FAS, ACCA, SCD1 and PPARγ). In addition, SA and SB decreased oxidative stress in the liver (CAT, GPX1α and SOD1) by activating the Keap1-Nrf2 pathway. Meanwhile, SA and SB alleviated HC-induced inflammation by downregulating the expression of pro-inflammatory factors (IL-1ß, COX2 and Hepcidin1) through the NF-κB pathway. Importantly, SA and SB increased the abundance of bacteria that produced acetic acid and butyrate (Clostridium_sensu_stricto_1). Combined with the KEGG analysis, the results showed that SA and SB enriched carbohydrate metabolism and amino acid metabolism pathways, thereby improving the utilization of carbohydrates. Pearson correlation analysis indicated that growth performance was closely related to hepatic lipid deposition, autophagy, antioxidant capacity, inflammation, and intestinal microbial composition. CONCLUSIONS: In conclusion, dietary SA and SB can reduce hepatic lipid deposition; and alleviate oxidative stress and inflammation in largemouth bass fed on HC diet. These beneficial effects may be due to the altered composition of the gut microbiota caused by SA and SB. The improvement effects of SB were stronger than those associated with SA.

Dietary sodium acetate and sodium butyrate attenuate intestinal damage and improve lipid metabolism in juvenile largemouth bass (Micropterus salmoides) fed a high carbohydrate diet by reducing endoplasmic reticulum stress.

High-carbohydrate (HC) diets decrease the intestinal levels of sodium acetate (SA) and sodium butyrate (SB) and impair the gut health of largemouth bass; however, SA and SB have been shown to enhance immunity and improve intestinal health in farmed animals. Thus, the present study was to investigate the effects of dietary SA and SB on HC diet-induced intestinal injury and the potential mechanisms in juvenile largemouth bass. The experiment set five isonitrogenous and isolipidic diets, including a low-carbohydrate diet (9% starch) (LC), a high carbohydrate diet (18% starch) (HC), and the HC diet supplemented with 2 g/kg SA (HCSA), 2 g/kg SB (HCSB) or a combination of 1 g/kg SA and 1 g/kg SB (HCSASB). The feeding experiment was conducted for 8 weeks. A total of 525 juvenile largemouth bass with an initial body weight of 7.00 ± 0.20 g were used. The results showed that dietary SA and SB improved the weight gain rate and specific growth rate (P < 0.05) and ameliorated serum parameters (alkaline phosphatase, acid phosphatase, glutamate transaminase, and glutamic oxaloacetic transaminase) (P < 0.05). And, importantly, dietary SA and SB repaired the intestinal barrier by increasing the expression levels of zonula occludens-1, occludin, and claudin-7 (P < 0.05), reduced HC-induced intestinal damage, and alleviated intestinal inflammation and cell apoptosis by attenuating HC-induced intestinal endoplasmic reticulum stress (P < 0.05). Further results revealed that dietary SA and SB reduced HC-induced intestinal fat deposition by inhibiting adipogenesis and promoting lipolysis (P < 0.05). In summary, this study demonstrated that dietary SA and SB attenuated HC-induced intestinal damage and reduced excessive intestinal fat deposition in largemouth bass.

Exposure to Intermittent Environmental Hypoxia Promotes Vascular Remodeling through Angiogenesis in the Liver of Largemouth Bass (Micropterus salmoides).

In this study, we explored the effects of 4 weeks of intermittent hypoxic exposure (IHE) on liver angiogenesis and related regulatory mechanisms in largemouth bass (Micropterus salmoides). The results indicated that the O2 tension for loss of equilibrium (LOE) decreased from 1.17 to 0.66 mg/L after 4 weeks of IHE. Meanwhile, the red blood cell (RBC) and hemoglobin concentrations significantly increased during IHE. Our investigation also found that the observed increase in angiogenesis was correlated with a high expression of related regulators, such as Jagged, phosphoinositide-3-kinase (PI3K), and mitogen-activated protein kinase (MAPK). After 4 weeks of IHE, the overexpression of factors related to angiogenesis processes mediated by HIF-independent pathways (such as nuclear factor kappa-B (NF-κB), NADPH oxidase 1 (NOX1), and interleukin 8 (IL8)) was correlated with the accumulation of lactic acid (LA) in the liver. The addition of cabozantinib, a specific inhibitor of VEGFR2, blocked the phosphorylation of VEGFR2 and downregulated the expression of downstream angiogenesis regulators in largemouth bass hepatocytes exposed to hypoxia for 4 h. These results suggested that IHE promoted liver vascular remodeling by the regulation of angiogenesis factors, presenting a potential mechanism for the improvement of hypoxia tolerance in largemouth bass.

Chronic hypoxia and Cu2+ exposure induce gill remodeling of largemouth bass through endoplasmic reticulum stress, mitochondrial damage and apoptosis.

Hypoxia and Cu2+ pollution often occur simultaneously in aquatic ecosystems and jointly affect physiology of fish. As the respiratory and ion exchange tissue of fish, how gill responds to the stress induced by these two abiotic environmental factors is still unclear. We have conducted a study by exposing largemouth bass (Micropterus salmoides) to hypoxia (2.0 mg·L-1) and/or Cu2+ (0.5 mg·L-1) for 28 days to answer this question. We subsequently studied respiratory rate, Cu2+ transport, endoplasmic reticulum (ER) stress, mitochondrial damage, and morphology in gill tissue on day 7, 14, 21 and 28. We found that hypoxia exposure increased the respiratory rate of largemouth bass, reflecting the response of largemouth bass to cope with hypoxia. Of note, Cu2+ entered gill by specifically binding to CTR1 and its accumulation dramatically in gill disrupted the response of largemouth bass to hypoxia. Hypoxia and/or Cu2+ exposure led to ER stress and mitochondrial damage in gills of largemouth bass. ER stress and mitochondrial damage induced apoptosis by activating caspase-8 and caspase-9 signaling pathways, respectively. Apoptosis induced by hypoxia and Cu2+ exposure had a positive and synergistic effect on gill remodeling by reducing interlamellar cell masses. In addition, Cu2+ exposure induced hypoxia-like remodeling to gill morphology through mechanisms similar to hypoxia exposure. Most of gene expression changed mainly within 21 days and recovered to the control level on day 28, reflecting the acclimation of largemouth bass to hypoxia and/or Cu2+ exposure at gene expression level. Overall, our research suggests that chronic hypoxia and Cu2+ exposure could induce gill remodeling of largemouth bass through ER stress, mitochondrial damage and apoptosis. The outcomes could provide an insight for fish environmental adaptation and environmental toxicology.

Enhance energy supply of largemouth bass (Micropterus salmoides) in gills during acute hypoxia exposure.

Gills are the location of gas exchange and also the first target organ of fish response for environmental stress. As a multifunctional organ, its energy supply, when faced with insufficient dissolved oxygen in the water, remains unclear. In this study, largemouth bass was subjected to hypoxia stress (1.2 mg/L) for 24 h and 12 h reoxygenation (R12) to evaluate energy supply strategy of gills. Under hypoxia exposure, the respiratory rate of largemouth bass increased by an average of 20 breaths per minute. A total of 2026, 1744, 1003, 579, 485, and 265 differentially expressed genes (DGEs) were identified at 0 h, 4 h, 8 h, 12 h, 24 h, and R12h in gills after hypoxia exposure. KEGG functional analysis of DEGs revealed that the glycolysis/gluconeogenesis pathway was enriched across all the sampling points (0, 4, 8, 12, 24 h, R12). The gene expression and enzyme activity of three rate-limiting enzymes (hexokinase, phosphofructokinase-6, pyruvate kinase) in glycolysis pathway were significantly increased. Increased levels of glycolysis products pyruvate and lactic acid, as well as the number of mitochondria (1.8-fold), suggesting an enhancement of aerobic and anaerobic metabolism of glucose in gills. These results suggest that the gill of largemouth bass enhanced the energy supply during acute exposure to hypoxia stress.

Metabolic response provides insights into the mechanism of adaption to hypoxia in largemouth bass (Micropterus salmoides) under intermittent hypoxic conditions.

In metabolism, molecular oxygen is a necessary substrate. Oxygen imbalances are linked to a variety of circumstances in the organism's homeostasis. Recently, the positive effects of hypoxia treatment in improving exercise ability and hypoxia tolerance have become a research focus. We explored the effects of intermittent hypoxia exposure (IHE, for one hour or three hours per day) on the hypoxia tolerance of largemouth bass in this study. The results showed that (1) IHE significantly reduced the LOEcrit (the critical O2 tension for loss of equilibrium) value of largemouth bass, indicating that its hypoxia tolerance was enhanced. (2) The level of oxidative stress in the liver decreased in the HH3 group (exposed to a hypoxic condition for 3 h per day) compared to HH1 group (exposed to a hypoxic condition for 1 h per day). (3) IHE reduced the content of lactic acid and enhanced the process of gluconeogenesis in the liver. (4) Importantly, lipid mobilization and fatty acid oxidation in the liver of largemouth bass were significantly enhanced during IHE. In short, the results of this study indicate that IHE can improve hypoxia tolerance by regulating the energy metabolism of largemouth bass.

Chromosome-level genome assembly of largemouth bass (Micropterus salmoides) using PacBio and Hi-C technologies.

The largemouth bass (Micropterus salmoides) has become a cosmopolitan species due to its widespread introduction as game or domesticated fish. Here a high-quality chromosome-level reference genome of M. salmoides was produced by combining Illumina paired-end sequencing, PacBio single molecule sequencing technique (SMRT) and High-through chromosome conformation capture (Hi-C) technologies. Ultimately, the genome was assembled into 844.88 Mb with a contig N50 of 15.68 Mb and scaffold N50 length of 35.77 Mb. About 99.9% assembly genome sequences (844.00 Mb) could be anchored to 23 chromosomes, and 98.03% assembly genome sequences could be ordered and directed. The genome contained 38.19% repeat sequences and 2693 noncoding RNAs. A total of 26,370 protein-coding genes from 3415 gene families were predicted, of which 97.69% were functionally annotated. The high-quality genome assembly will be a fundamental resource to study and understand how M. salmoides adapt to novel and changing environments around the world, and also be expected to contribute to the genetic breeding and other research.