Protective effects of sulforaphane on inflammation, oxidative stress and intestinal dysbacteriosis induced by triphenyltin in Cyprinus carpio haematopterus.

The purpose of this experiment was to study the mitigation effect of sulforaphane (SFN) on fish toxicological damage caused by triphenyltin (TPT) pollution. A total of 320 healthy fish (56.9 ± 0.4g) were randomly placed into four groups, each with four duplicates. The control group was fed the basal diet, the TPT group was exposed to 10 ng/L TPT on the basis of the control group, the SFN group was fed a diet supplemented with 10 mg/kg SFN, the SFN + TPT group was exposed to 10 ng/L TPT on the basis of the SFN group. Each tank had 20 fish and the breeding lasted for 8 weeks. The present study found that the antioxidant enzyme activity in the TPT group was significantly lower than that of the control group (P < 0.05). In addition, compared with the control group, the mRNA expression of pro-inflammatory factors (IL-6, TNF-α) were significantly induced, and the anti-inflammatory factor genes (IL-10, TGF) were significantly inhibited (P < 0.05) in TPT group. SFN relieved the changes of inflammatory factors caused by TPT, ameliorated oxidative stress, improved antioxidant enzyme (include SOD, CAT, GSH, GPx) activities (P < 0.05). 16s RNA analysis indicated that exposure to TPT caused changes in intestinal microflora. The results of the study showed that after exposure to TPT, some beneficial genera of bacteria in the gut of Rhizobiaceae, Bdellovibrio and Candidatus Alysiosphaera were decreased. The bacteria associated with intestinal inflammation including Propionibacterium, Rubrobacter, Anaerorhabdus_furcosa_group, Rikenellaceae and Eubacterium_brachy were upregulated. However, the SFN treatment group significantly down-regulated the above five inflammation-related bacteria. The above results indicated that TPT caused oxidative stress and inflammation in fish intestines, changed the intestinal microflora, and dietary SFN could improve antioxidant status, regulate inflammation and intestinal health. Therefore, SFN is a promising diet additive for improving fish damage caused by TPT contamination.

Effects of salinity stress on anxiety behavior and antioxidant capability of guppy (Poecilia reticulata).

The objective of this study was to determine the effect of salinity on anxiety behavior and liver antioxidant capacity in the guppy (Poecilia reticulata). Guppies were exposed to salinities of 0‰, 5‰, 10‰, 15‰ and 20‰ for acute stress tests, and then we analyzed the activity of antioxidant enzymes at 3, 6, 12, 24, 48, 72 and 96 h. During the experiment, the anxiety behavior of guppy was enhanced at salinities of 10‰, 15‰, and 20‰, as evidenced by a significantly higher latency time for the first passage through the upper part than that of the control group (P < 0.05). CAT activity was highest at 24 h in the treatment with the salinity of 10‰, and SOD and GPX activities were highest at 12 h into the treatment with the salinity of 10‰. The SOD and CAT activities were significantly higher than the control group after 96 h of treatment at different salinities (P < 0.05). The MDA contents of the experimental groups at salinities of 5‰ and 10‰ were not significantly different from the control group after 96 h of treatment (P > 0.05). While the MDA contents of the experimental groups at salinities of 15‰ and 20‰ were still significantly higher than the control group after 96 h of treatment (P < 0.05). The experimental results indicated that elevated salinity could lead to oxidative stress in the guppy, altering their anxiety behavior as well as the activity of antioxidant enzymes. In conclusion, drastic changes in salinity during culture should be avoided.

Protective effects of the fructooligosaccharide on the growth performance, biochemical indexes, and intestinal morphology of blunt snout bream (Megalobrama amblycephala) infected by Aeromonas hydrophila.

The purpose of the study was to investigate the effects of dietary fructooligosaccharide (FOS) on growth performance, biochemical indexes, intestinal morphology, and growth-related gene expression of blunt snout bream (Megalobrama amblycephala) infected by Aeromonas hydrophila (AH). Two hundred twenty-five healthy blunt snout bream with an initial body weight of 38.41 ± 0.88 g were randomly divided into five groups with three replicates: control (basal diet), model (AH + basal diet), SFOS (AH + 2 g/kg FOS), MFOS (AH + 4 g/kg FOS), LFOS (AH + 6 g/kg FOS). After 9 weeks of feeding, the results showed that the FOS-added diet abrogated AH-induced retardation, hemorrhage, and inflammatory infiltration. FOS supplementation enhanced the growth performance degradation caused by AH, and the highest growth performance was observed at MFOS. Meanwhile, the addition of FOS to feed improved the blood immunity reduced by AH. In expansion, the mucosal epithelium of intestinal villi exfoliated, exposing the lamina propria, and a few villi were genuinely harmed in the model group. Fish fed with MFOS ameliorated the damaged intestine, evidenced by well-preserved intestine architecture. Furthermore, the model group downregulated the expression of growth-related genes (growth hormone receptor (GHR), insulin-like growth factor 1 (IGF-1)). Fish fed with 2 g/kg or 4 g/kg FOS upregulated the genes specified above expressions in the liver compared with the model group. In conclusion, the results mentioned above suggested that the dietary FOS could relieve the pressure to elevate the immune damage and intestine injury induced by AH and enhance the hepatic expression of IGF-1 and GHR.

The intestinal histopathology, innate immune response and antioxidant capacity of blunt snout bream (Megalobrama amblycephala) in response to Aeromonas hydrophila.

The present study was performed to determine the effects of Aeromonas hydrophila infection on intestinal -histopathology, innate immune response and changes in antioxidant capacity of blunt snout bream (Megalobrama amblycephala). A series of histopathological changes, innate immune enzyme activities, antioxidant enzyme activities, and the corresponding mRNA relative genes expressions in intestines were measured at 0, 1, 2, and 3 weeks post-treatment of Aeromonas hydrophila (1✕107 CFU mL-1) infection. The results showed that Aeromonas hydrophila induced changes in intestinal morphology, including the decreased muscularis thickness, the proliferated goblet cells, and the atrophied intestine villi height. Moreover, the innate immune enzymes activities in serum such as acid phosphatase, alkaline phosphatase, lysozyme activities and immunoglobulin M were significantly reduced after infection at 1week, 2week and 3week. The contents of complement 3 and complement 4 were significantly decreased after infection as well. In addition, the antioxidant enzymes activities, including superoxide dismutase, catalase and glutathione peroxidase in the experimental groups were significantly decreased compared with the control group, whereas the content of malondialdehyde was significantly increased after infection at 1week, 2week and 3week. Furthermore, the mRNA relative expressions of the inflammatory cytokines such as tumor necrosis factor-α, interleukins-1ß, interferon-γ, and interleukins-6 were significantly increased after infection with Aeromonas hydrophila. The TJ-related gene expressions in the intestine of zonula occluden-1, occludin, occludin-1, occludin-2 were significantly reduced throughout the infection period. The mRNA relative expressions of signal transducers and activators of transcription 4 and janus kinase-3 in the intestine were significantly ascended compared with the non-infected group. Overall, the results elucidated that the intestine tissue injury and innate immune response reduction, as well as antioxidant capacity attenuation were occurred against Aeromonas hydrophila infection of the blunt snout bream.

The effects of TPT and dietary quercetin on growth, hepatic oxidative damage and apoptosis in zebrafish.

The objective of this study was to determine the effects of triphenyltin (TPT) and dietary quercetin on the growth, oxidative stress and apoptosis in zebrafish. A total of 240 fish were divided into 4 groups with three replicates as follows: fish were fed with the basal diet as the control group (D1), only 10 ng/L TPT (D2), 10 ng/L TPT + 100 mg/kg quercetin (D3), and only 100 mg/Kg quercetin as the D4 group. At the end of the study period (56 d), the results showed that the growth performance of the fish that were fed 100 mg/kg quercetin was significantly higher than that of fish that were exposed to 10 ng/L TPT. Quercetin ameliorated oxidative stress, which decreased malondialdehyde (MDA) and nitric oxide (NO) levels and improved antioxidant enzyme activities. The mRNA expressions of the key apoptotic gene and pro-inflammatory cytokines were significantly induced by TPT exposure. However, dietary quercetin prevented a marked increase in the Bax, caspase3 and caspase9 transcript abundances that were induced by TPT. In addition, the quercetin treatments decreased inflammation by regulating the NF-kB signalling pathway. In conclusion, our findings suggested that TPT induced oxidative stress and apoptosis in zebrafish and that the pretreatment with quercetin showed an ameliorative role. Dietary 100 mg/ kg quercetin helps to prevent oxidative damage, apoptosis and inflammation in TPT treated zebrafish.

Modulatory effect of fructooligosaccharide against triphenyltin-induced oxidative stress and immune suppression in goldfish (Carassius auratus).

Triphenyltin (TPT) is a widely used pesticide that is highly toxic to a variety of organisms, including humans, and is a potential contributor to environmental pollution. The present study was conducted to evaluate the oxidative stress and immunotoxicity induced by TPT in goldfish (Carassius auratus) and the protective effects of fructooligosaccharide (FOS). Goldfish (mean weight of 13.3 ± 0.2 g) were randomly divided into six groups with three replicates: (G1) the control group, (G2) the 10 ng/L TPT group, (G3) the 0.4% FOS group, (G4) the 10 ng/L TPT + 0.4% FOS group, (G5) the 0.8% FOS group, and (G6) the 10 ng/L TPT + 0.8% FOS group. The results showed that 10 ng/L TPT induced oxidative stress and significantly decreased the activities of antioxidant enzymes, such as superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx), in the liver and the gene expression of SOD, GPx, metallothionein (MT), and peroxiredoxin-4 (Prdx-4). The concentration of malondialdehyde (MDA) and the gene expression of cytochrome P450 (CYP) and glutathione S-transferase (GST) in the liver were significantly increased in the TPT-treated group. Exposure to 10 ng/L TPT in water induced immune suppression and significantly decreased the activities of immune enzymes, such as lysozyme, myeloperoxidase (MPO), alternative complement (ACH50), acid phosphatase (ACP) and alkaline phosphatase (AKP), in the serum. TPT could stimulate the fish to generate large amounts of proinflammatory cytokines, including increased tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), interleukin-1ß (IL-1ß), and nitric oxide (NO) levels and TNF-α, IL-6, IL-1ß, and NF-κB mRNA expression. However, TPT-induced toxicity was significantly ameliorated in the groups treated with FOS, and FOS partly prevented alterations in the activities of antioxidant enzymes and the expression of antioxidant- and ROS scavenger-related genes. In addition, TPT-induced immune toxicity was significantly ameliorated in the groups treated with FOS. FOS markedly suppressed TNF-α, IL-6, IL-1ß, and NO production and TNF-α, IL-6, and IL-1ß mRNA expression in the TPT-treated groups. The study indicated that TPT-induced oxidative stress may play a critical role in inhibiting immunity. However, FOS administration attenuates TPT-induced oxidative stress and immune suppression in goldfish.

Quercetin affects shoaling and anxiety behaviors in zebrafish: Involvement of neuroinflammation and neuron apoptosis.

Quercetin, a potential fish food supplement, has been reported to process many beneficial properties. However, some negative effects of quercetin have been observed, which pointed out necessity for additional studies to evaluate its safety. Therefore, the present study investigated effects of quercetin (0.01, 0.1, 1, 10, 100 and 1000 µg/L) on shoaling and anxiety behaviors through novel tank tests in zebrafish (Danio rerio). Furthermore, oxidative stress, neuroinflammation and apoptosis in the brains were examined to learn more about mechanisms of action related to quercetin. The results showed that quercetin at the lower concentrations exerted beneficial effects on shoaling and anxiety behaviors. On the contrary, when quercetin was up to 1000 µg/L, it exerted detrimental effects shown as decreases of movement and increases of anxiety behaviors. Generally, U-shaped responses of antioxidant enzyme activities (superoxide dismutase and catalase), and inversed U-shaped responses of inflammatory mediators (cyclooxygenase-2) and cytokines (interleukin-1ß, interleukin-6, interleukin-10, and tumor necrosis factor α) to quercetin treatment were found in the brains. In addition, quercetin at the lower concentrations attenuated cell apoptosis, while even more apoptosis was found at the 1000 µg/L quercetin group. In conclusion, quercetin could exert beneficial or detrimental effects on the shoaling and anxiety behaviors depending on the treatment concentrations, and the underlying mechanisms are potentially associated with neuroinflammation and neuron apoptosis.

Molecular cloning, expression HSP70 and its response to bacterial challenge and heat stress in Microptenus salmoides.

The gene encoding HSP70 was isolated from Microptenus salmoides by homologous cloning and rapid amplification of cDNA ends (RACE). The HSP70 transcripts were 2116 bp long and contained 1953 open reading frames encoding proteins of 650 amino acids with a molecular mass of 71.2 kDa and theoretical isoelectric point of 5.22. The qRT-PCR analysis showed that the HSP70 gene was differentially expressed in various tissues under normal conditions, and the highest HSP70 level was observed in the spleen and the lowest levels in the muscle and heart. The clear time-dependent expression level of HSP70 was observed after bacterial challenge and heat stress. A significant increase in HSP70 expression level was detected and reached a maximum at 3 h and 6 h in liver, spleens and gill tissues after Aeromonas hydrophila infection and heat stress, respectively (P < 0.05). As time progressed, the expression of HSP70 transcript was downregulated and mostly dropped back to the original level at 48 h. The concentration of cortisol, aspartate aminotransferase (AST) and alanine aminotransferase (ALT) increased as the time of stress progressed, with the highest level found on 3 h and later declined rapidly and reached to the control levels at the 48 h. Those results suggested that HSP70 was involved in the immune response to bacterial challenge and heat stress. The cloning and expression analysis of the HSP70 provide theoretical basis to further study the mechanism of anti-adverseness in Microptenus salmoides.

The effects of quercetin on immunity, antioxidant indices, and disease resistance in zebrafish (Danio rerio).

The main purpose of this study was to evaluate the immunity, antioxidant indices, and disease resistance of quercetin in zebrafish (Danio rerio). A total of 630 fish were assigned to 21 tanks with 30 fish/tank, and they were exposed to 0, 0.01, 0.1, 1, 10, 100, and 1000 µg/L quercetin, respectively, for 56 days. Results indicated that the immune indices including acid phosphatase (ACP), myeloperoxidase (MPO), lysozyme activities, and Complement 3 (C3), C4, IgM contents were significantly higher in 1 µg/L quercetin group than these parameters in the control group (P < 0.05). TNF-α and IL-8 mRNA expressions significantly decreased as the levels of quercetin increased up to 1 µg/L and increased thereafter (P < 0.05). 1 and 10 µg/L quercetin groups showed significantly lower TNF-α and IL-8 mRNA levels than the quercetin-free group. Transforming growth factor-ß and IL-10 mRNA levels showed an obviously opposite trend with TNF-α expression. The SOD, GPX, CAT, T-AOC activities, and SOD and GPX gene expression in the liver were enhanced with increasing quercetin up to 1 µg/L, and decreased thereafter. MDA contents were affected by quercetin, in which 1 and 10 µg/L quercetin had a significantly lower level than that of the control group (P < 0.05). Defensin and Leap-II mRNA expression in the liver were the highest for fish exposed to 1 µg/L quercetin. The fish that exposed to 1 µg/L quercetin also showed a significantly higher survival rate than these of fish exposed to 0, 0.01, and 1000 µg/L quercetin (P < 0.05). In conclusion, the optimal level of quercetin promotes immunostimulant properties, antioxidant indices, and disease resistance of zebrafish.

Molecular characterization of p38 MAPK from blunt snout bream (Megalobrama amblycephala) and its expression after ammonia stress, and lipopolysaccharide and bacterial challenge.

p38 mitogen-activated protein kinase (MAPK) is an important protein which plays a key role in regulating the innate immunity, so exploring its molecular characterization is helpful in understanding the resistance against microbial infections in cultured fish. Here, a full-length cDNA of p38 MAPK was cloned from liver of blunt snout bream (Megalobrama amblycephala) which covered 2419 bp with an open reading frame of 1086 bp encoding 361 amino acids. p38 MAPK contained the characteristic structures of Thr-Gly-Tyr (TGY) motif and substrate binding site Ala-Thr-Arg-Trp (ATRW), which are conserved in MAPK family. To investigate p38 MAPK functions, two in vivo experiments were carried out to examine its expression following ammonia exposure and bacterial challenge. Also, an in vitro experiment was conducted to assess the role of p38 MAPK in inflammation of primary hepatocytes induced by lipopolysaccharide (LPS). The results showed the ubiquitous expression of p38 MAPK in all the tested tissues with varying levels. p38 MAPK mRNA expression was significantly up-regulated by ammonia stress and Aeromonas hydrophila challenge, and altered in a time-dependent manner. Moreover, the results indicated that the inflammatory response induced by LPS in hepatocytes is p38 MAPK dependent as knockdown of p38 MAPK using siRNA technology depressed the expression of IL-1ß and IL-6. The findings in this study showed that p38 MAPK has anti-stress property, and plays key role in protection against bacterial infection and inflammation in blunt snout bream.

Triphenyltin exposure affects mating behaviors and attractiveness to females during mating in male guppies (Poecilia reticulata).

The impacts of triphenyltin (TPT) on ecological health have been of great concern due to their widespread use and ubiquity in aquatic ecosystems. However, little is known about the effects of TPT on the reproductive behaviors of fishes. Therefore, the present study was conducted to investigate the effects of TPT at environmentally relevant concentrations (0, 1 and 10 ng Sn/L) on the mating behaviors and the attractiveness to females during mating in male guppies (Poecilia reticulata). The results showed that TPT exposure disturbed the mating behaviors; the TPT-exposed male fish performed more sneaking attempts, but no changes in sigmoid courtship were displayed. The increases in sneaking attempts might be related to increases in testosterone levels induced by TPT exposure. In the context of a competing male, the TPT-exposed males were less attractive to females during mating. The decreases in attractiveness might be related to decreases in carotenoid-based coloration, shown as decreases in caudal fin redness values and skin carotenoid contents. In addition, TPT-induced total antioxidant capacities, the activities of superoxide dismutase and catalase, and the contents of malondialdehyde in liver and intestinal tissues indicated increases in oxidative stress. Both oxidative stress and coloration are linked to carotenoids. Thus, we speculated that the TPT-exposed males might use carotenoids to cope with increases in oxidative stress at the expense of carotenoid-based coloration. The disruption of mating behaviors and the decrease in attractiveness to females in male fish could result in reproductive failure. The present study underscores the importance of using behavioral tests as a sensitive tool in assessing the impact of pollutants present in aquatic environments.

Molecular cloning, expression and antibacterial activity of goose-type lysozyme gene in Microptenus salmoides.

It is well known that lysozymes are key proteins to teleosts in the innate immune system and possess high bactericidal properties. In the present study, a g-type lysozyme gene was cloned from Microptenus salmoides. The g-type sequence consisted of 582 bp, which translated into a 193 amino acid (AA) protein (GenBank accession no: MH087462). The predicted molecular weight and theoretical isoelectric point were 21.36 kDa and 6.91 respectively and no signal peptide was observed. The qRT-PCR analysis showed that the g-type lysozyme gene was differentially expressed in various tissues under normal conditions and the highest g-type lysozyme level was observed in liver, gill and spleen while there seemed to be low expression in the muscle, heart and head-kidney. The expression of g-type lysozyme was differentially upregulated in the spleen, gill and intestine after stimulation with heat stress and Aeromonas hydrophila (A. hydrophila). Under heat stress and A. hydrophila injection, the g-type lysozyme mRNA levels all in spleens, gill and intestine tissues increased significantly (P < 0.05), with the maximum levels attained at 12 h, 24 h (or 12 h) and 24 h. Thereafter, they all decreased significantly (P < 0.01) and the expression in gill returned to nearly the basal value within 72 h. Those results suggested that g-type lysozyme was involved in the immune response to heat stress and bacterial challenge. The cloning and expression analysis of the g-type lysozyme provide theoretical basis to further study the mechanism of anti-adverseness in Microptenus salmoides. The g-type lysozyme gene perhaps also played an important role in the immune responses against bacterial invasion.

Transcriptomic analyses of tributyltin-induced sexual dimorphisms in rare minnow (Gobiocypris rarus) brains.

The brain of fish displays sexual dimorphisms and exhibits remarkable sexual plasticity throughout their life span. Although reproductive toxicity of tributyltin (TBT) in fish is well documented in fish, it remains unknown whether TBT interrupts sexual dimorphisms of fish brains. In this work, brain transcriptomic profiles of rare minnow (Gobiocypris rarus) was characterized and sex-biased genes were identified using RNA sequencing. Functional annotation and enrichment analysis were performed to reveal differences of gene products and pathways between the brains of male and female fish. Furthermore, transcriptomic responses of male and female brains to TBT at 10 ng/L were also investigated to understand effects of TBT on brain sexual dimorphisms. Only 345 male-biased and 273 female-biased genes were found in the brains. However, significant female-biased pathways of circadian rhythm and phototransduction were identified in the brains by enrichment analysis. Interestingly, following TBT exposure in the female fish, the circadian rhythm pathway was significantly disrupted based on enrichment analysis, while in the male fish, the phototransduction pathway was significantly disrupted. In the female fish, expression of genes (Per, Cry, Rev-Erb α, Ror, Dec and CK1δ/ε) in the circadian rhythm pathway was down-regulated after TBT exposure; while in the male fish, expression of genes (Rec, GNAT1_2, GNGT1, Rh/opsin, PDE and Arr) in the phototransduction pathway was up-regulated after TBT exposure. Overall, our results not only provide key data on the molecular basis of brain sexual dimorphisms in fish, but also offer valuable resources for investigating molecular mechanisms by which environmental chemicals might influence brain sexual plasticity.

Dibutyltin depressed immune functions via NF-κB, and JAK/STAT signaling pathways in zebrafish (Danio rerio).

Dibutyltin (DBT) is the degradation products of TBT, which is generally considered higher toxicity than TBT in the immune system. In order to learn more about the mechanisms of immune-toxic of DBT, we exposed zebrafish (Danio rerio) to 0, 1, 10 and 100 ng/L DBT for 8 weeks. At the end of the experiment, we determined the immune parameters and immune-related genes. The results showed that with an increase in TBT dose, lysozyme activities and IgM, C3, C4 content in intestine, skin and spleen were all significantly inhibited by the DBT exposure. Fish exposed to 10 ng/L and 100 ng/L showed significantly lower lysozyme activities and IgM, C3, C4 content than those of the control group. Zebrafish exposed to 10 ng/L and 100 ng/L DBT, the mRNA transcript levels of interleukin-1ß (IL-1ß), interleukin-6 (IL-6), interleukin-8 (IL-8), tumor necrosis factor-α (TNF-α), interferon γ2 (INFγ2), nuclear factor-κB p65 (NF-kB p65), inhibitor protein-κBα (IκBα), IκB kinases ß (IKKß), Janus family of protein tyrosine kinases (JAKs) and the signal transducers and activators of transcription proteins (STATs) all increased with the DBT levels in the intestine and spleen. Those parameters showed significantly higher values in 10 ng/L and 100 ng/L than those of fish in the control group. However, no significant difference was found in IκB kinases α (IKKα) and IκB kinase γ (IKKγ) mRNA levels in the intestine and spleen. These data imply that DBT might be via suppression on IKKß/IkBa/NF-kBp65 and JAK/STAT signaling pathways to regulate the immunity of zebrafish.

Lipid accumulation, oxidative stress and immune-related molecules affected by tributyltin exposure in muscle tissues of rare minnow (Gobiocypris rarus).

Tributyltin (TBT) is reported to induce adipogenesis in fish, which might affect nutritional qualities and health status. Muscle tissues account for the majority of body mass, and have been described as a major site of fat deposition and an immunologically active organ. Therefore, the present study aims to evaluate whether chronic exposures of TBT, at environmental concentrations of 1, 10 and 100 ng/L, affects lipid accumulation, oxidative stress and immune status in muscle tissues of rare minnow (Gobiocypris rarus). After 60 d of exposure, TBT increased contents of total lipid, total cholesterol, triglyceride and fatty acids in muscle tissues. Interestingly, TBT exposure disrupted fatty acid composition and increased contents of unsaturated fatty acids (such as eicosapentaenoic acid and docosahexaenoic acid) in muscle tissues, which might be a response to preserve membrane functions from TBT exposure. Meanwhile, the concentrations of hepatic fatty acid desaturase 2 (Δ6-desaturase) and stearoyl-CoA desaturase (Δ9-desaturase) were increased after TBT exposure, which might contribute the increase of unsaturated fatty acids. Furthermore, TBT increased muscle lipid peroxidation products, antioxidant enzymes (superoxide dismutase, catalase and glutathione peroxidase), and the expression of immune-related molecules (tumor necrosis factor alpha, interleukin 1 beta and nuclear factor kappa B) in muscle tissues. The disruption of TBT on the lipid accumulation, oxidative stress and immune-toxic effects in muscle tissues of fish might reduce nutritional qualities, and affect growth and health status, which might pose a constant and serious threat to fish and result in economic loss in aquaculture.

Effects of Lactobacillus delbrueckii on immune response, disease resistance against Aeromonas hydrophila, antioxidant capability and growth performance of Cyprinus carpio Huanghe var.

The aim of the present study was to investigate effects of dietary Lactobacillus delbrueckii (L. delbrueckii) on immune response, disease resistance against Aeromonas hydrophila (A. hydrophila), antioxidant capability and growth performance of Cyprinus carpio Huanghe var. 450 fish (mean weight of 1.05 ± 0.03 g) were randomly distributed into five groups that fed diets containing different levels of L. delbrueckii (0, 1 × 105, 1 × 106, 1 × 107 and 1 × 108 CFU g-1) for 8 weeks. The results showed that intestinal immune parameters such as lysozyme, acid phosphatase, and myeloperoxidase activities, immunoglobulin M content, and the survival rate were improved in fish fed with 1 × 106 and 1 × 107 CFU g-1L. delbrueckii. In addition, 1 × 107 CFU g-1L. delbrueckii supplementation down-regulated mRNA levels of TNF-α, IL-8, IL-1ß and NF-κBp65, and up-regulated IL-10 and TGF-ß mRNA levels in the intestine. The survival rate was significantly (P < 0.05) higher (68.33%) in fish fed 1 × 106 CFU g-1L. delbrueckii than the control diet-fed group (40%) after challenge by A. hydrophila. Fish fed with diet containing 1 × 106 CFU g-1L. delbrueckii showed higher antioxidant enzyme activities such as superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPX), and total antioxidant capacity (T-AOC) and lower MDA concentrations than those of the control group (P < 0.05). The relative gene expression (SOD, CAT, GPX) showed the same trend with their activities. In addition, the growth performance was significantly improved in fish fed with the diet containing 1 × 106 and 1 × 107 CFU g-1L. delbrueckii (P < 0.05). These results demonstrated that dietary optimal levels of L. delbrueckii enhanced immunity, disease resistance against A. hydrophila antioxidant capability and growth performance in Cyprinus carpio Huanghe var.

Effect of tributyltin on antioxidant ability and immune responses of zebrafish (Danio rerio).

Tributyltin (TBT) is a toxic compound released into aquatic ecosystems through antifouling paints. This study was designed to examine the effects of TBT on antioxidant ability and immune responses of zebrafish (Danio rerio). Three hundred sixty healthy zebrafish were randomly grouped into four groups and exposed to different doses of TBT (0, 1, 10 and 100ngL-1). At the end of 8 weeks, the fish were sampled, and antioxidant capability, immune parameters and immune-related genes were assessed. The results showed that with an increase in TBT dose, the concentration of malonaldehyde in the liver was significantly increased (p<0.05), whereas the activities of total superoxide dismutase, catalase and glutathione peroxidase were significantly decreased (p<0.05) compared to the control. The activity and expression of lysozyme and the content of immunoglobulin M were significantly decreased compared to those of the fish exposed to 0ngL-1 TBT (p<0.05). However, the expression of the HSP70, HSP90, tumor necrosis factor-α(TNF-α), interleukins (IL-1ß, IL-6), and nuclear factor-kappa B p65 (NF-κ B p65) genes were all enhanced with an increase in TBT dose. The results indicated that TBT induced oxidative stress and had immunotoxic effects on zebrafish.

Cloning, characterization and mRNA expression of interleukin-6 in blunt snout bream (Megalobrama amblycephala).

In the present study, the interleukin-6 gene (IL-6) cDNA in blunt snout bream (Megalobrama amblycephala) was identified and its expression profiles under ammonia stress and bacterial challenge were investigated. The IL-6 sequence consisted of 1045 bp, including a 696 bp ORF which translated into a 232 amino acid (AA) protein. The protein contained a putative signal peptide of 24 AA in length. IL-6 expression analysis showed that the it is differentially expressed in various tissues under normal conditions and the highest IL-6 level was observed in the intestine tissue, followed by the liver, and then in the gills. Under ammonia stress, the IL-6 mRNA level both in spleens and intestine increased significantly (P < 0.05), with the maximum levels attained at 6 h, 12 h (72, 10-fold, respectively). Thereafter, they all significantly decreased (P < 0.01) and returned to the basal value within 48 h. Whereas, in livers it slightly decreased at 3 h firstly (0.5-fold), and then significantly (P < 0.05) increased with the maximum level attained 12 h (3-fold). Further expression analysis showed that the mRNA level of IL-6 in spleens, intestine and livers of blunt snout bream all increased significantly (P < 0.05), with maximum values attained at 6 h, 3 h, 6 h (10, 6, 18-fold, respectively) after Aeromonas hydrophila (A. hydrophila) injection, and then decreased to the basal value within 24 h which suggested that IL-6 was involved in the immune response to A. hydrophila. The cloning and expression analysis of the IL-6 provide theoretical basis to further study the mechanism of anti-adverseness and expression characteristics under stress conditions in blunt snout bream.

Effects of fructooligosaccharide on immune response, antioxidant capability and HSP70 and HSP90 expressions of blunt snout bream (Megalobrama amblycephala) under high ammonia stress.

This study aimed to determine the effects of fructooligosaccharide (FOS) on immune response, antioxidant capability and HSP70 and HSP90 mRNA expressions of blunt snout bream (Megalobrama amblycephala) under high ammonia stress. A total of 360 fish were randomly distributed into three groups (each with four replicates) and were fed three levels of FOS (0, 0.4 and 0.8 %) for 8 weeks. After the feeding trial, 24 fish per tank were exposed to ammonia at 10 mg L(-1). After stress, plasma cortisol and glucose levels of fish fed 0.4 % FOS were all significantly lower than that of the control group at 6 and 3 h, respectively. Plasma lysozyme and alternative complement pathway (ACH50) activities as well as nitrogen monoxide (NO) levels all increased significantly with the maximum levels being attained at 6, 6 and 3 h, respectively. Thereafter, these parameters all decreased significantly. In addition, fish fed 0.4 % FOS showed higher immune parameters under stress compared with that of control group. In addition, liver superoxide dismutase and catalase activities of fish fed 0.4 % FOS were both significantly higher than that of the control group before and after stress, while the opposite was true for malondialdehyde content. After stress, the expression of HSP70 and HSP90 of fish fed FOS was significantly higher than that of the control group at 6 and 12 h, respectively. After 12 h stress, the cumulative mortality of fish fed FOS was significantly lower than that of the control. The results indicated that the supplementation of 0.4 % FOS could increase the nonspecific immunity, antioxidant capacity and HSP70 and HSP90 expression of blunt snout bream and enhance its resistance to high ammonia stress.

Effects of dietary fructooligosaccharide levels and feeding modes on growth, immune responses, antioxidant capability and disease resistance of blunt snout bream (Megalobrama amblycephala).

This study aimed to determine the effects of fructooligosaccharide (FOS) levels and its feeding modes on growth, immune response, antioxidant capability and disease resistance of blunt snout bream (Megalobrama amblycephala). Fish (12.5 ± 0.5 g) were subjected to three FOS levels (0, 0.4% and 0.8%) and two feeding modes (supplementing FOS continuously and supplementing FOS two days interval 5 days) according to a 3 × 2 factorial design. At the end of 8-week feeding trial, fish were challenged by Aeromonas hydrophila with concentration of 1 × 10(5) CFU mL(-1) and mortality was recorded for the next 96 h. Fish fed 0.4% FOS continuously (D2) and fish fed the basal diet for 5 days followed by 0.8% FOS for 2 days (D5) showed admirable growth performance. The highest plasma lysozyme, acid phosphatase and myeloperoxidase activities as well as complement component 3, total protein and immunoglobulin M (IgM) levels were all observed in fish fed D5. They were significantly higher (P < 0.05) than those of the control group and/or fish fed 0.8% FOS continuously, but exhibited no statistical difference (P > 0.05) with that of fish fed D2. A similar trend was also observed in antioxidant capability as well as the expression of Leap-I and Leap-Ⅱ. Mortality showed an opposite trend with the immune response with the lowest rate observed in fish fed D5. The results indicated that diet supplementing FOS in appropriate levels and feeding modes could improve the growth, immune response and antioxidant capability of fish, as might consequently lead to enhanced disease resistance. It can be speculated that the basal diet for 5 days followed by 0.8% FOS for 2 days was most suitable for blunt snout bream.