Effects of effective microorganisms on the physiological status, intestinal microbiome, and serum metabolites of Eriocheir sinensis / Efectos de microorganismos eficaces sobre el estado fisiológico, el microbioma intestinal y los metabolitos séricos de Eriocheir sinensis

The compound known as effective microorganisms (EMs) is widely used in aquaculture to improve water quality, but how they affect the health of Chinese mitten crab (Eriocheir sinensis) is unclear, especially in terms of intestinal microbiota and serum metabolites. In this study, we fed juvenile crabs with an EM-containing diet to explore the effects of EM on the physiological status, intestinal microbiome, and metabolites of E. sinensis. The activities of alanine aminotransferase and alkaline phosphatase were significantly enhanced by EM, indicating that EM supplementation effectively enhanced the antioxidant capacity of E. sinensis. Proteobacteria, Tenericutes, Firmicutes, Bacteroidetes, and Actinobacteria were the main intestinal microbes in both the control and EM groups. Linear discriminant effect size analysis showed that Fusobacteriaceae, Desulfovibrio, and Morganella were biomarkers in the control group, and Exiguobacterium and Rhodobacteraceae were biomarkers in the EM group. Metabolomics analysis revealed that EM supplementation increased cellular energy sources and decreased protein consumption, and oxidative stress. Together, these results indicate that EM can optimize the intestinal microbiome and serum metabolites, thereby benefiting the health of E. sinensis.(AU)

Effects of effective microorganisms on the physiological status, intestinal microbiome, and serum metabolites of Eriocheir sinensis.

The compound known as effective microorganisms (EMs) is widely used in aquaculture to improve water quality, but how they affect the health of Chinese mitten crab (Eriocheir sinensis) is unclear, especially in terms of intestinal microbiota and serum metabolites. In this study, we fed juvenile crabs with an EM-containing diet to explore the effects of EM on the physiological status, intestinal microbiome, and metabolites of E. sinensis. The activities of alanine aminotransferase and alkaline phosphatase were significantly enhanced by EM, indicating that EM supplementation effectively enhanced the antioxidant capacity of E. sinensis. Proteobacteria, Tenericutes, Firmicutes, Bacteroidetes, and Actinobacteria were the main intestinal microbes in both the control and EM groups. Linear discriminant effect size analysis showed that Fusobacteriaceae, Desulfovibrio, and Morganella were biomarkers in the control group, and Exiguobacterium and Rhodobacteraceae were biomarkers in the EM group. Metabolomics analysis revealed that EM supplementation increased cellular energy sources and decreased protein consumption, and oxidative stress. Together, these results indicate that EM can optimize the intestinal microbiome and serum metabolites, thereby benefiting the health of E. sinensis.

Involvement of reactive oxygen species (ROS) in the hepatopancreatic cytotoxicity, oxidative stress, and apoptosis induced by microcystin-LR in Eriocheir sinensis.

There is limited knowledge about the toxicity of Microcystin-LR (MC-LR) in crustaceans, despite its high toxicity to aquatic organisms. This research aimed to explore the effects of MC-LR on cytotoxicity, oxidative stress, and apoptosis in the hepatopancreas of Eriocheir sinensis, as well as elucidate the involvement of reactive oxygen species (ROS) and potential mechanisms of toxicity. In vivo and in vitro exposures of crabs to MC-LR and N-acetylcysteine (NAC) were performed, followed by assessments of cell morphology, viability, tissue pathology, biochemical indicators, gene expression, and hepatopancreatic transcriptome. Results revealed that MC-LR facilitated the entry of the MC-LR transporter oatp3a into hepatopancreatic cells, leading to upregulated expression of phase I detoxification enzyme genes (cyp4c, cyp2e1, and cyp3) and downregulated the phase II enzyme genes (gst1, gpx, gsr2, gclc, and nqo1), resulting in increased ROS levels and cytotoxic effects. MC-LR exhibited cytotoxicity, reducing cell viability and inducing abnormal nuclear morphology with a 48 h-IC50 value of approximately 120 µm. MC-LR exposure caused biochemical changes indicative of oxidative stress damage and evident hepatopancreatic lesions. Additionally, MC-LR exposure regulated the levels of bax and bcl-2 expression, activating caspase 3 and 6 to induce cell apoptosis. Intervention with NAC attenuated MC-LR-induced ROS production and associated toxic effects. Transcriptome analysis revealed enrichment of differentially expressed genes in pathways related to cytochrome P450-mediated xenobiotic metabolism and the FoxO signaling pathway. These findings shed light on the potential mechanisms underlying MC-LR toxicity and provide valuable references for further research and conservation efforts regarding the health of aquatic animals.

Prometryn exposure disrupts the intestinal health of Eriocheir sinensis: Physiological responses and underlying mechanism.

Most toxicity studies of prometryn in non-target aquatic animals have focused on hepatotoxicity, cardiotoxicity, embryonic developmental and growth toxicity, while studies on the molecular mechanisms of intestinal toxicity of prometryn are still unknown. In the current study, the intestinal tissues of the Chinese mitten crab (Eriocheir sinensis) were used to uncover the underlying molecular mechanisms of stress by 96-h acute in vivo exposure to prometryn. The results showed that prometryn activated the Nrf2-Keap1 pathway and up-regulated the expression of downstream antioxidant genes. Prometryn induced the expression of genes associated with non-specific immunity and autophagy, and induced apoptosis through the MAPK pathway. Interestingly, the significant up-or down-regulation of the above genes mainly occurred at 12 h- 24 h after exposure. Intestinal flora sequencing revealed that prometryn disrupted the intestinal normal barrier function mainly by reducing beneficial bacteria abundance, which further weakened the intestinal resistance to exogenous toxicants and caused an inflammatory response. Correlation analyses found that differential flora at the genus level had potential associations with gut stress-related genes. In conclusion, our study contributes to understanding the molecular mechanisms behind the intestinal stress caused by herbicides on aquatic crustaceans.

Microcystin-LR-induced autophagy via miR-282-5p/PIK3R1 pathway in Eriocheir sinensis hepatopancreas.

With the intensifying climate warming, blue-green algae blooms have become more frequent and severe, releasing environmental hazards such as microcystin that pose potential threats to human and animal health. Autophagy has been shown to play a crucial role in regulating immune responses induced by environmental hazards, enabling cells to adapt to stress and protect against damage. Although microcystin-LR (MC-LR) has been identified to affect autophagy in mammalian, its impact on aquatic animals has been poorly studied. To investigate the toxicological effects of MC-LR in aquatic ecosystems, we constructed a microRNA profile of acute MC-LR stress in the hepatopancreas of the Chinese mitten crab. Interestingly, we found the MC-LR exposure activated autophagy in the hepatopancreas based on the following evidence. Specifically, mRNA expression level of ATG7, Beclin1 and Gabarap was significantly up-regulated, autophagy regulatory pathways were significantly enriched, and numerous autolysosomes and autophagosomes were observed. Additionally, we found that miR-282-5p and its target gene PIK3R1 played important regulatory roles in autophagy by in vivo and in vitro experiments. Overexpression of miR-282-5p mimicked MC-LR-induced autophagy by inhibiting PIK3R1 expression, while miR-282-5p silencing inhibited autophagy by promoting PIK3R1 expression. Altogether, our findings suggest that MC-LR increases miR-282-5p, which then targets inhibition of PIK3R1 to stimulate autophagy. This study focused on the stress response regulatory mechanisms of juvenile crabs to toxic pollutants in water, offering a potential target for alleviating the toxicity of MC-LR. These findings lay a foundation for reducing the toxicity of MC-LR and environmental hazards in organisms.

Effects of Prometryn Exposure on Hepatopancreas Oxidative Stress and Intestinal Flora in Eriocheir sinensis (Crustacea: Decapoda).

There is growing evidence that long-term exposure to prometryn (a widely used herbicide) can induce toxicity in bony fish and shrimp. Our previous study demonstrated its 96 h acute toxicity on the crab Eriocheir sinensis. However, studies on whether longer exposure to prometryn with a lower dose induces toxicity in E. sinensis are scarce. Therefore, we conducted a 20 d exposure experiment to investigate its effects on the hepatopancreas and intestine of E. sinensi. Prometryn reduce the activities of antioxidant enzymes, increase the level of lipid peroxidation and cause oxidative stress. Moreover, long-term exposure resulted in immune and detoxification fatigue, while short-term exposure to prometryn could upregulate the expression of genes related to immunity, inflammation and detoxification. Prometryn altered the morphological structure of the hepatopancreas (swollen lumen) and intestine (shorter intestinal villi, thinner muscle layer and thicker peritrophic membrane). In addition, prometryn changed the species composition of the intestinal flora. In particular, Bacteroidota and Proteobacteria showed a dose-dependent decrease accompanied by a dose-dependent increase in Firmicutes at the phylum level. At the genus level, all exposure groups significantly increased the abundance of Zoogloea and a Firmicutes bacterium ZOR0006, but decreased Shewanella abundance. Interestingly, Pearson correlation analysis indicated a potential association between differential flora and hepatopancreatic disorder. Phenotypic abundance analysis indicated that changes in the gut flora decreased the intestinal organ's resistance to stress and increased the potential for opportunistic infection. In summary, our research provides new insights into the prevention and defense strategies in response to external adverse environments and contributes to the sustainable development of E. sinensis culture.

Integrated Analysis of Transcriptomics and Metabolomics Unveil the Novel Insight of One-Year-Old Precocious Mechanism in the Chinese Mitten Crab, Eriocheir sinensis.

Eriocheir sinensis is traditionally a native high-value crab that is widely distributed in eastern Asia, and the precocity is considered the bottleneck problem affecting the development of the industry. The precocious E. sinensis is defined as a crab that reaches complete sexual maturation during the first year of its lifespan rather than as normally in the second year. However, the exact regulatory mechanisms underlying the precocity are still unclear to date. This study is the first to explore the mechanism of precocity with transcriptome-metabolome association analysis between the precocious and normal sexually mature E. sinensis. Our results indicated that the phenylalanine metabolism (map00360) and neuroactive ligand-receptor interaction (map04080) pathways play an important role in the precocity in the ovary of E. sinensis. In map00360, the predicted aromatic-L-amino-acid decarboxylase and 4-hydroxyphenylpyruvate dioxygenase isoform X1 genes and the phenethylamine, phenylethyl alcohol, trans-2-hydroxycinnamate, and L-tyrosine metabolites were all down-regulated in the ovary of the precocious E. sinensis. The map04080 was the common KEGG pathway in the ovary and hepatopancreas between the precocious and normal crab. In the ovary, the predicted growth hormone secretagogue receptor type 1 gene was up-regulated, and the L-glutamate metabolite was down-regulated in the precocious E. sinensis. In the hepatopancreas, the predicted forkhead box protein I2 gene and taurine metabolite were up-regulated and the the L-glutamate metabolite was down-regulated in the precocious crab. There was no common pathway in the testis. Numerous common pathways in the hepatopancreas between male precocious and normal crab were identified. The specific amino acids, fatty acids and flavorful nucleotide (inosine monophosphate (MP), cytidine MP, adenosine MP, uridine MP, and guanosine MP) contents in the hepatopancreas and gonads further confirmed the above omics results. Our results suggest that the phenylalanine metabolism may affect the ovarian development by changing the contents of the neurotransmitter and tyrosine. The neuroactive ligand-receptor interaction pathway may affect the growth by changing the expressions of related genes and affect the umami taste of the gonads and hepatopancreas through the differences of L-glutamate metabolite in the precocious E. sinensis. The results provided valuable and novel insights on the precocious mechanism and may have a significant impact on the development of the E. sinensis aquaculture industry.

Effects of prometryn on oxidative stress, immune response and apoptosis in the hepatopancreas of Eriocheir sinensis (Crustacea: Decapoda).

Prometryn, a triazine pesticide product used to control weed growth, poses a high risk to aquatic organisms in the environment. Several toxicological evaluations have been performed on bony fish and shrimp exposed to prometryn. However, there have been no reports conducted on the toxic mechanism of prometryn with regard to Eriocheir sinensis. In this study, our research evaluated the toxic effects of prometryn via in vitro and in vivo toxicity tests on E. sinensis. Firstly, we estimated the exposure toxicity of prometryn to E. sinensis, and then we constructed a 6 h transcriptional profile and conducted an enrichment analysis. To further reveal the toxicity of prometryn, the hepatopancreas (hepatopancreatic cells) was analyzed for antioxidant, immune and lipid-metabolism-related enzymes, antioxidant- and apoptosis-related gene expression, histopathology and TUNEL. From the results, we determined that the 96 h-LD50 was 70.059 mg/kg, and using RNA-seq, we identified 933 differentially expressed genes (DEGs), which were mainly enriched in the amino and fatty acid metabolism and the cell-fate-determination-related signaling pathway. The results of the biochemical assays showed that prometryn could significantly decrease the activities/levels of CAT, SOD, GSH, AKP and ACP, reduce the levels of T-AOC, TG, TCH, C3 and C4, and increase the MDA content. In addition, the expression levels of Nrf2, GSTs and HO-1 were first upregulated and then downregulated with increasing time. Histopathology showed that prometryn damaged the structure of the hepatopancreas cells and induced apoptosis, suggesting that the PI3K-Akt signaling pathway may be involved in the damage process of hepatopancreas cells (PI3K, PDK and Akt were downregulated whereas Bax was upregulated), leading to their apoptosis. The above results indicated that prometryn could cause injury of the hepatopancreas through oxidative stress, induce cell apoptosis, disrupt the lipid metabolism and cause immune damage. This study provided useful data for understanding and evaluating the toxicity of prometryn to aquatic crustacea.

Salvianolic Acid B Regulates Oxidative Stress, Autophagy and Apoptosis against Cyclophosphamide-Induced Hepatic Injury in Nile Tilapia (Oreochromis niloticus).

Salvianolic acid B (Sal B), as one of the main water-soluble components of Salvia miltiorrhizae, has significant pharmacological activities, including antioxidant, free radical elimination and biofilm protection actions. However, the protective effect of Sal B on Nile tilapia and the underlying mechanism are rarely reported. Therefore, the aim of this study was to evaluate the effects of Sal B on antioxidant stress, apoptosis and autophagy in Nile tilapia liver. In this experiment, Nile tilapia were fed diets containing sal B (0.25, 0.50 and 0.75 g·kg-1) for 60 days, and then the oxidative hepatic injury of the tilapia was induced via intrapleural injection of 50 g·kg-1 cyclophosphamide (CTX) three times. After the final exposure to CTX, the Nile tilapia were weighed and blood and liver samples were collected for the detection of growth and biochemical indicators, pathological observations and TUNEL detection, as well as the determination of mRNA expression levels. The results showed that after the CTX treatment, the liver was severely damaged, the antioxidant capacity of the Nile tilapia was significantly decreased and the hepatocyte autophagy and apoptosis levels were significantly increased. Meanwhile, dietary Sal B can not only significantly improve the growth performance of tilapia and effectively reduce CTX-induced liver morphological lesions, but can also alleviate CTX-induced hepatocyte autophagy and apoptosis. In addition, Sal B also significantly regulated the expression of genes related to antioxidative stress, autophagy and apoptosis pathways. This suggested that the hepatoprotective effect of Sal B may be achieved through various pathways, including scavenging free radicals and inhibiting hepatocyte apoptosis and autophagy.

Oxidative stress and immune response of hepatopancreas in Chinese mitten crab Eriocheir sinensis under lipopolysaccharide challenge.

Chinese mitten crab (Eriocheir sinensis; H. Milne Edwards, 1853) is one of the important farmed crustaceans in China. Lipopolysaccharide (LPS), as a harmful factor, is prone to occur during the farming process of crabs. Aiming to test the hypothesis that damage degrees of the hepatopancreas in E. sinensis is correlated to LPS concentrations, in this study, E. sinensis were injected with LPS (50 µg/kg, and 500 µg/kg) and analyzed for the activity of antioxidant and immune-related enzymes, immune-related gene expression, and histopathological of hepatopancreas. As result, the hepatopancreas of E. sinensis immune-related genes, i.e., Dorsal, HSP90, Toll2, TLRs, Tube, and proPO, were significantly affected by LPS challenge. Among immune-related genes, Dorsal and proPO might play key roles in combating the LPS challenge. The activity of CAT gradually decreased with the increase of time, and the total antioxidant capacity was decreased after LPS challenge, indicating the inhibition of LPS on the antioxidant system. Interestingly, the decreasing trend of AKP and ACP activity suggested the immune system of crabs was affected by LPS challenge. The hepatopancreas section showed that the damage degree of hepatopancreas was different under the challenge of LPS with different concentrations, and the damage degree was proportional to the concentration. Our findings provide useful information for understanding the mechanism of hepatopancreas injury of E. sinensis induced by LPS infection.

Alleviative effects of astragaloside IV on cyclophosphamide-induced oxidative damage and immunosuppression in tilapia (Oreochromis niloticus).

Astragaloside IV (ASIV) has effects of antioxidation and immunologic enhancement. However, there are few reports on the application and potential mechanism of ASIV in aquaculture. In this study, we investigated the effect of ASIV on growth, antioxidation, and immune function of tilapia. Tilapia were fed a diet containing 0.1, 0.2, and 0.5 g·kg-1 ASIV for 60 days, followed by an intrapleural injection of 50 mg·kg-1 cyclophosphamide (CTX) to induce oxidative damage and immunosuppression. Then tilapia were weighed and blood, liver, spleen, kidney, and intestinal were collected. The results showed ASIV increased the final weight, relative weight rate, and specific growth rate of tilapia, reduce conversion ratio, and reduced the morphological lesions of tissues. Meanwhile, ASIV alleviated CTX-induced oxidative damage by improving antioxidant activity in serum and tissues and inhibiting lipid peroxidation. Additionally, ASIV attenuated the immunosuppression of tilapia caused by CTX, regulated immunochemical indexes in serum, increased the viability of peripheral blood leukocytes and head kidney macrophages, and restored respiratory burst activity (O2-) in head kidney macrophages and splenocytes. Furthermore, qPCR data showed ASIV up-regulated antioxidant-related gene expression of nrf2, ho-1, gpx3, and cat and immune-related gene expression including C3 and igm. In conclusion, ASIV as a feed additive can not only improve the growth performance but also enhance the antioxidant capacity and immune function of tilapia, which may be associated with the ability of ASIV to scavenge free radicals, reduce lipid peroxidation levels, and stabilize numbers of immune cells.

Protective Effects of Glycyrrhiza Total Flavones on Liver Injury Induced by Streptococcus agalactiae in Tilapia (Oreochromis niloticus).

Clinical studies have confirmed that Glycyrrhiza total flavones (GTFs) have good anti-hepatic injury, but whether they have a good protective effect on anti-hepatic injury activity induced by Streptococcus agalactiae in tilapia (Oreochromis niloticus) is unknown. The aims of this study were to investigate the protective effects of Glycyrrhiza total flavones on liver injury induced by S. agalactiae (SA) and its underlying mechanism in fish. A total of 150 tilapia were randomly divided into five groups, each with three replicates containing 10 fish: normal control group, S. agalactiae infection group, and three Glycyrrhiza total flavone treatment groups (addition of 0.1, 0.5, or 1.0 g of GTF to 1 kg of feed). The normal control group was only fed with basic diet, after 60 d of feeding, and intraperitoneal injection of the same volume of normal saline (0.05 mL/10 g body weight); the S. agalactiae infection group was fed with basic diet, and the S. agalactiae solution was intraperitoneally injected after 60 d of feeding (0.05 mL/10 g body weight); the three GTF treatment groups were fed with a diet containing 0.1, 0.5, or 1.0 g/kg of GTF, and the S. agalactiae solution was intraperitoneally injected after 60 d of feeding (0.05 mL/10 g body weight). After 48 h injection, blood and liver tissues were collected to measure biochemical parameters and mRNA levels to evaluate the liver protection of GTFs. Compared with the control group, the serum levels of glutamic oxaloacetic transaminase (GOT), glutamic pyruvic transaminase (GPT), alkaline phosphatase (AKP) and glucose (GLU) in the streptococcal infection group increased significantly, while the levels of total antioxidant capacity (T-AOC), superoxide dismutase (SOD), catalase (CAT) and reduced glutathione (GSH) decreased significantly; observations of pathological sections showed obvious damage to the liver tissue structure in response to streptococcal infection. S. agalactiae can also cause fatty liver injury, affecting the function of fatty acid ß-oxidation and biosynthesis in the liver of tilapia, and also causing damage to function of the immune system. The addition of GTFs to the diet could improve oxidative stress injury caused by S. agalactiae in tilapia liver tissue to different degrees, promote the ß-oxidation of fatty acids in the liver, accelerate the lipid metabolism in the liver, and repair the damaged liver tissue. GTFs have a good protective effect on liver injury caused by streptococcus.

Exploration of the regulatory mechanisms of regeneration, anti-oxidation, anti-aging and the immune response at the post-molt stage of Eriocheir sinensis.

Eriocheir sinensis is widely appreciated by the surrounding population due to its culinary delicacy and rich nutrients. The E. sinensis breeding industry is very prosperous and molting is one of the important growth characteristics. Research on the regulation of molting in E. sinensis is still in the initial stages. There is currently no relevant information on the regulatory mechanisms of heart development following molting. Comparative transcriptome analysis was used to study developmental regulation mechanisms in the heart of E. sinensis at the post-molt and inter-molt stages. The results indicated that many regulatory pathways and genes involved in regeneration, anti-oxidation, anti-aging and the immune response were significantly upregulated after molting in E. sinensis. Aside from cardiac development, the differentially expressed genes (DEGs) were relevant to myocardial movement and neuronal signal transduction. DEGs were also related to the regulation of glutathione homeostasis and biological rhythms in regard to anti-oxidation and anti-aging, and to the regulation of immune cell development and the immune response. This study provides a theoretical framework for understanding the regulation of molting in E. sinensis and in other economically important crustaceans.

Multi-kingdom microbiota and functions changes associated with culture mode in genetically improved farmed tilapia (Oreochromis niloticus).

Genetically improved farmed tilapia (GIFT, Oreochromis niloticus) are intensively farmed in China, where most of the yield derives from the pond culture system (PCS). The in-pond raceway system (IPRS) is a new type of highly efficient aquaculture mode, and has been recommended as a novel system for GIFT farming. To determine the effects of these culture modes on the gut microbiome of GIFT, we conducted a 90-days experiment in IPRS and PCS units. A 16S rRNA gene profile analysis showed that the composition of gut microbiota in GIFT under IPRS and PCS conditions gradually separated as rearing progressed, with divergent responses by the midgut and hindgut bacteria. The α-diversity in hindgut decreased significantly by day 90, as compared with on day 7 (p < 0.05), with a significantly greater decrease in PCS-reared fish than in IPRS fish (p < 0.05). The α-diversity of microbiota in midgut remained stable (p > 0.05). The overall dominant gut bacteria were Bacteroidetes, Proteobacteria, and Firmicutes. Rearing mode affected the taxonomic profile of the gut bacteria; in midgut, IPRS samples had more Firmicutes and Fusobacteria compared with PCS samples, but less Proteobacteria, Verrucomicrobia, and Actinobacteria. Firmicutes was enriched in IPRS hindgut, and Fusobacteria was enriched in PCS hindgut. Using random-forest models and LEfSe, we also screened core taxa that could discriminate between the gut microbial communities under IPRS and PCS conditions. The genus Cetobacterium (of family Fusobacteriaceae) was significantly enriched in midgut in IPRS fish, and enriched in hindgut in PCS fish. The genus Clostridium sensu stricto (of family Clostridiaceae 1) was significantly enriched in both IPRS midgut and hindgut. Analysis with PICRUSt2 software revealed that the culture modes were similar in their effects on the gut microbial metabolic functions. The predicted pathways were significantly enriched in the metabolism class (level 1). Further, the relative abundance of functions related to amino acid metabolic, carbohydrate metabolic, energy metabolic, and metabolic of cofactors and vitamins were high at hierarchy level 2, as the metabolic activity of intestinal bacteria is especially active. Overall, this study enhances our understanding of the characteristics of gut microbiota in GIFT under IPRS and PCS culture modes. Moreover, our findings provide insights into the microecological balance in IPRS units, and a theoretical reference for further development of this culture system.

Effects of Submerged Macrophytes on the Growth, Morphology, Nutritional Value, and Flavor of Cultured Largemouth Bass (Micropterus salmoides).

Aquaculture environment plays important roles in regulating the growth, morphology, nutrition, and flavor of aquatic products. The present study investigated growth, morphology, nutrition, and flavor formation in largemouth bass (Micropterus salmoides) cultured in the ponds with (EM group) and without (M group) the submerged macrophytes (Elodea nuttallii). Fish in the EM group showed a significantly greater body length, higher growth rate, and lower hepatosomatic index than those in the M group (p< 0.05). Moreover, compared with fish in the M group, those in the EM group showed improved muscle quality with significantly elevated levels of crude protein, total free and hydrolysable amino acids, and polyunsaturated fatty acids (p < 0.05). Specifically, certain amino acids related to flavor (Glu, Asp, Ala, and Arg) and valuable fatty acids (C18:2, C18:3n3, C20:3n3, and C22:6) were more abundant in the EM group (p < 0.05). In addition, the levels of 19 volatile (p < 0.05) were significantly higher in the EM group than in the M group. Therefore, E. nuttallii significantly improved growth, morphological traits, nutritional components, and characteristic flavor in largemouth bass, indicating the superior nutritional value and palatability of fish cultured with submerged macrophytes.

Effective mechanisms of water purification for nitrogen-modified attapulgite, volcanic rock, and combined exogenous microorganisms.

The study tested the water purification mechanism of the combination of microorganisms and purification materials via characteristic, enzymatic, and metagenomics methods. At 48 h, the removal rates of total nitrogen, total phosphorous, and Mn chemical oxygen demand in the combination group were 46.91, 50.93, and 65.08%, respectively. The alkaline phosphatase (AKP) activity increased during all times tested in the volcanic rock, Al@TCAP, and exogenous microorganism groups, while the organophosphorus hydrolase (OPH), dehydrogenase (DHO), and microbial nitrite reductase (NAR) activities increased at 36-48, 6-24, and 36-48 h, respectively. However, the tested activities only increased in the combination groups at 48 h. Al@TCAP exhibits a weak microbial loading capacity, and the Al@TCAP removal is primarily attributed to adsorption. The volcanic rock has a sufficient ability to load microorganisms, and the organisms primarily perform the removal for improved water quality. The predominant genera Pirellulaceae and Polynucleobacter served as the sensitive biomarkers for the treatment at 24, 36-48 h. Al@TCAP increased the expression of Planctomycetes and Actinobacteria, while volcanic rock increased and decreased the expression of Planctomycetes and Proteobacteria. The growth of Planctomycetes and the denitrification reaction were promoted by Al@TCAP and the exogenous microorganisms. The purification material addition group decreased the expression of Hyaloraphidium, Chytridiomycetes (especially Hyaloraphidium), and Monoblepharidomycetes and increased at 36-48 h, respectively. Ascomycota, Basidiomycota, and Kickxellomycota increased in group E, which enhanced the nitrogen cycle through microbial enzyme activities, and the growth of the genus Aspergillus enhanced the phosphorous purification effect.

Effects of resveratrol on lipid metabolism in liver of red tilapia Oreochromis niloticus.

Resveratrol (RES), as a polyphenol natural plant extract, mainly accumulates in the root of Polygonum cuspidatum, which can alleviate liver injury in mammals. Our study aims to explore the effects and potential mechanism of RES on lipid metabolism of red tilapia, and the effects of RES on liver structure, fat synthesis and metabolism of red tilapia were determined. The present study designed four groups named as 8 % fat (8%CK), 10 % fat (10 % HF), 10 % HF + RES and 10 % HF + RES + EX527 (selisistat). The liver tissues of red tilapia were collected at 3 (3 W), 6 (6 W) and 9 (9 W) weeks for parameter determination. Compared to the normal diet group, the hepatocyte of tilapia showed nuclear shift and vacuoles of different sizes when fed a high-fat diet. Meanwhile, the high-fat diet increased the contents of LDL, TC and TG significantly at 6 W, and significantly decreased the content of NAD+ at 9 W. Compared to the high-fat group, the nuclei of tilapia fed with RES were increased and visible, the degree of steatosis and the number of vacuoles were both reduced. At 3/6/9 W, RES significantly decreased the contents of LDL, TG and TMAO, and significantly increased the content of NAD+. A total of 1416 genes were up-regulated and 1928 genes were down-regulated in the group with added RES when compared to the 10 % HF group. The pathways related to lipid metabolism including PPAR signaling pathway have been enriched. Interestingly, the expressions of sirt1, pparα, fabp7 and cpt1b genes were up-regulated in RES diet group, while the expressions of pparγ, me1, scd and lpl genes were down-regulated. After the addition of an inhibitor (EX527), the above indexes showed an opposite trend when compared to the group with added RES. The overall results showed that the high-fat diet could cause fatty liver lesions in the liver of red tilapia, and RES could activate the sirt1 gene, regulate the PPARα/γ pathway and related genes, and thus regulate liver fat synthesis and metabolism leading to the alleviation of damage to liver tissue.

Microcystin-LR induces apoptosis in Juvenile Eriocheir sinensis via the mitochondrial pathway.

Microcystin-LR (MC-LR), the toxic substance of cyanobacteria secondary metabolism, widely exists in water environments and poses great risks to living organisms. Some toxicological assessments of MC-LR have performed at physiological and biochemical levels. However, plenty of blanks about the potential mechanism in aquatic crustacean remains. In this study, we firstly assessed the exposure toxicity of MC-LR to juvenile E. sinensis and clarified that the 96 h LD50 of MC-LR was 73.23 µg/kg. Then, hepatopancreas transcriptome profiles of MC-LR stressed crabs were constructed at 6 h post-injection and 37 differential expressed genes (DEGs) were identified. These DEGs were enriched in cytoskeleton, peroxisome and apoptosis pathways. To further reveal the toxicity of MC-LR, oxidative stress parameters (SOD, CAT, GSH-px and MDA), apoptosis genes (caspase 3, bcl-2 and bax) and apoptotic cells were detected. Significant accumulated MDA and rise-fall enzyme activities verified the oxidative stress caused by MC-LR. It is noteworthy that quantitative real-time PCR and TUNEL assay indicated that MC-LR stress-induced apoptosis via the mitochondrial pathway. Interestingly, activator protein-1 may play a crucial role in mediating the hepatotoxicity of MC-LR by regulating apoptosis and oxidative stress. Taken together, our study investigated the toxic effects and the potential molecular mechanisms of MC-LR on juvenile E. sinensis. It provided useful data for exploring the toxicity of MC-LR to aquatic crustaceans at molecular levels.

Effects of submerged macrophytes (Elodea nuttallii) on water quality and microbial communities of largemouth bass (Micropterus salmoides) ponds.

Traditional aquaculture ponds are one of the most vulnerable ecosystems; thus, ecological aquaculture is increasingly valued for its beneficial ecological properties and ecosystem services. However, little is known about ecological aquaculture of largemouth bass with submerged vegetation. Here, we designed three ecological ponds of cultured largemouth bass with submerged macrophytes (the EM group) and three ponds with traditional aquaculture (the M group) to reveal the response of water quality, and phytoplankton and bacterial communities, to submerged macrophyte bioremediation during a 90-day culture period. We observed that Cyanobacterial outbreak occurred in the M group ponds from day 7 to the end of the experiment; however, there were no Cyanobacterial blooms in the EM group ponds throughout the culture period. Compared with the M group ponds, the EM group ponds, which had submerged hydrophytes, had significantly decreased concentrations of TP, TN, and CODMn, but significantly increased DO concentrations throughout the experimental period. Moreover, ecological aquaculture with submerged macrophytes showed strong effects on the phytoplankton and bacterial community compositions. In particular, the M group ponds had higher phytoplankton density and mainly included Cyanobacteria, whereas the EM group had lower phytoplankton density and mainly included Chlorophyta. Moreover, higher alpha diversity, as determined by Ace and Simpson index values, was detected for bacterial communities in the EM group ponds. Furthermore, PCoA clearly grouped the bacterial communities according to the two culture modes throughout the culture period. These results indicate that ecological aquaculture with submerged macrophytes can improve water quality, control Cyanobacterial blooms, and affect the diversity and composition of bacterial communities. These valuable effects seem to be beneficial and consistent to maintaining aquaculture ecosystem stability.

Optimum feeding frequency of juvenile largemouth bass (Micropterus salmoides) reared in in-pond raceway recirculating culture system.

This study was conducted to determine the effects of feeding frequency on the growth, serum biochemical parameters, antioxidant status and hepatic growth hormone (GH), insulin-like growth factor I (IGF-I), lipoprotein lipase (LPL) and hepatic lipase (HL) gene expression levels of juvenile largemouth bass (Micropterus salmoides) reared in an in-pond raceway recirculating culture system (IPRS). Fish (initial body weight 5.0 ± 0.4 g) were hand-fed with a commercial diet under one of three different feeding frequency treatments (2, 3 or 4 meals/day) for 120 days. The results indicated that no significant differences were observed in the final body weight, weight gain and specific growth rate of fish fed different feeding frequencies on 30 days and 60 days (P > 0.05). Fish fed 2 times/day had higher growth than that fed 4 times/day on 90 days but had higher growth than those fed 3 and 4 times/day on 120 days. No significant differences were found in serum alanine aminotransferase (ALT), aspartate transaminase (AST), alkaline phosphatase (ALP) activities, total protein (TP), lysozyme and triglyceride (TG) content, hepatic total antioxidant capacity (T-AOC), glutathione peroxidase (GSH-PX) activities and malondialdehyde (MDA) content among fish fed different feeding frequency (P > 0.05). Serum glucose (Glu) content and catalase (CAT) activity decreased, while total cholesterol (TC) content increased with increasing feeding frequency. Fish fed 2 times/day had higher hepatic total superoxide dismutase (T-SOD) than that fed 4 times/day on 60 days, 90 days and 120 days (P < 0.05). Fish fed 2 times/day had higher IGF-1 gene mRNA expression on 30 days, 60 days and 120 days (P < 0.05), while no significant difference on 90 days. No significant difference was found in GH gene mRNA expression on 30 days and 60 days, while fish fed 4 times/day had lower values than that fed 2 times/day on 90 days and 120 days (P < 0.05). Fish fed 2 times/day had significantly higher LPL mRNA expression level than that fed 4 times/day on 60 days and 90 days and had significantly higher HL mRNA expression level on 60 days, 90 days and 120 days (P < 0.05). Based on growth, physiology, hepatic gene expression levels, labour costs and intensity, the optimal feeding frequency of largemouth bass (average body weight 5.0 ± 0.4 g) reared in IPRS is 2 times/day. These data are very necessary for the optimizing of culture conditions and feeding management strategy in IPRS culture operations.