Impact of Sel-Plex® dietary supplementation on growth performance, physiological response, oxidative status, and immunity-linked gene expression in Nile tilapia (Oreochromis niloticus) fingerlings challenged with Aeromonas hydrophila.

Background: Organic selenium (Sel-Plex®) supplementation holds considerable promise for improving the effectiveness of fish production. Aim: This experiment was accomplished to judge the potential benefits of Sel-Plex® nutritional additive on growth outcomes, physiological response, oxidative status, and immunity-linked gene expression in Nile tilapia (Oreochromis niloticus) fingerlings exposed to bacterial infection with Aeromonas hydrophila. Methods: Utilizing a basal diet of 30% protein, four experimental diets were prepared, each of which contained Sel-Plex® at concentrations of 0.0, 0.5, 1, and 2 mg/kg, respectively. Three replicates of 20 fish/treatment were used using 240 healthy Nile tilapia fingerlings. Fish were placed in 12 glass aquariums and separated into 4 groups at random. For the entire span of 8 weeks, diets were admitted to fish at a 3% rate of fish biomass/aquarium. After the feeding trial, pathogenic A. hydrophila was intraperitoneally injected into fish of each treatment, and fish were observed for 15 days to track the survival rate (SR) after the challenge. Results: Growth performance, physiological response, immunological parameters (phagocytic activity, phagocytic index, and lysozyme), and antioxidant parameters [catalase, superoxide dismutase (SOD), malondialdehyde, and glutathione peroxidase (GPx)] were noticeably improved in Sel-Plex® treated groups. Moreover, Sel-Plex® increased gene expression linked with the immune system in the liver (tumor necrosis factor-alpha and interleukin 1ß), to growth (insulin-like growth factor 1 and growth hormone receptor), and antioxidants (SOD and GPx). Under pathogen-challenge conditions, the employed dietary Sel-Plex® supplementation could successfully lower fish oxidative stress, offering a potential preventive additive for Nile tilapia instead of antibiotics. On the other hand, Sel-Plex® significantly enhanced each of three intestinal morphological measurements (villus width, villus length, and crypt depth), demonstrating the greatest influence on the improvement of intestinal structure overall. In the Nile tilapia control group, the infection with A. hydrophila caused noticeable degenerative alterations in the gut, hepatopancreas, spleen, and posterior kidney. The severity of the lesion was significantly reduced and significantly improved with higher Sel-Plex® concentrations. Sel-Plex® supplemented groups had 100% SRs among the A. hydrophila-challenged groups. Conclusion: It could be advised to enrich the diets of Nile tilapia fingerlings with 1-2 mg.kg-1 of Sel-Plex® to enhance growth rate, physiological response, immunological reaction, and intestinal absorptive capacity.

Ameliorative effect of BIO-MOS® as a dietary supplementation on growth performance, physiological response, oxidative status, and immunity-linked gene expression in Nile tilapia (Oreochromis niloticus) fingerlings challenged with Aeromonas hydrophila.

Background: Mannanoligosaccharides (MOS) usage in fish production has drawn more attention because of their positive benefits on disease resistance and fish performance. Aim: The ongoing research was executed to assess the potential advantages of Bio-Mos® dietary supplementation regarding the growth outcomes, physiological response, oxidative biomarkers, and immunity-linked gene expression in Nile tilapia (Oreochromis niloticus) fingerlings exposed to bacterial infection with Aeromonas hydrophila. Methods: Four experimental diets were developed using a 30% protein baseline diet, with Bio-Mos® added at variable levels; 0.0, 0.5, 1, and 2 g/kg, respectively. 240 healthy Nile tilapia fingerlings were split into 4 groups at random and assigned to 12 glass aquariums (three replicates of 20 fish/treatment). Diets were admitted at a 3% rate of fish biomass/aquarium for 8 weeks. Following the feeding trial, fish from every treatment were intraperitoneally injected with pathogenic A. hydrophila, and then observed for 15 days to record the survival rate percent (SR%) post challenge. Results: Results revealed significant improvement in growth performance, physiological response, immunological parameters (phagocytic index, phagocytic activity, and lysozyme), and antioxidant parameters [catalase, malondialdehyde, glutathione peroxidase (GSH-Px), and superoxide dismutase (SOD)] among Bio-Mos® treated groups. Moreover, Bio-Mos® increased the expression of tumor necrosis factor alpha and Interleukin 1ß, genes linked to the liver immune system. Growth-related genes (GHr), antioxidant-related genes (SOD and GSH-Px). In fish subjected to pathogens, dietary MOS supplementation could significantly lower oxidative stress, showing promise as a preventative supplement for Nile tilapia in place of antibiotics. On the other hand, Bio-Mos® considerably improved each of the three intestinal morphological measures (villus width, villus length, and crypt depth), showing the best overall intestinal structure-improving impact. The challenge with A. hydrophila caused marked degenerative alterations in the intestine, hepatopancreas, spleen, and posterior kidney of Nile tilapia, in the control group. However, lesion severity was greatly decreased and showed marked amelioration with an increased concentration of Bio-Mos®. The A. hydrophila-challenged groups revealed a 100% SR% mainly among the Bio-Mos® supplemented groups. Conclusion: It is recommended to enrich the Nile tilapia fingerlings diets with 2 g.kg-1 of MOS for better results on the growth rate, physiological response, immunological response, and intestinal absorptive capacity.

Roles of ß-catenin in innate immune process and regulating intestinal flora in Qi river crucian carp (Carassius auratus).

In mammals, ß-catenin participates in innate immune process through interaction with NF-κB signaling pathway. However, its role in teleost immune processes remains largely unknown. We aimed to clarify the function of ß-catenin in the natural defense mechanism of Qi river crucian carp (Carassius auratus). ß-catenin exhibited a ubiquitous expression pattern in adult fish, as indicated by real-time PCR analysis. Following lipopolysaccharide (LPS), Polyinosinic-polycytidylic acid (polyI: C) and Aeromonas hydrophila (A. hydrophila) challenges, ß-catenin increased in gill, intestine, liver and kidney, indicating that ß-catenin likely plays a pivotal role in the immune response against pathogen infiltration. Inhibition of the ß-catenin pathway using FH535, an inhibitor of Wnt/ß-catenin pathway, resulting in pathological damage of the gill, intestine, liver and kidney, significant decrease of innate immune factors (C3, defb3, LYZ-C, INF-γ), upregulation of inflammatory factors (NF-κB, TNF-α, IL-1, IL-8), and downregulation of glutathione peroxidase (GSH-Px), superoxide dismutase (SOD) and catalase (CAT) activities, increase of Malondialdehyde (MDA) content. Following A. hydrophila invasion, the mortality rate in the FH535 treatment group exceeded that of the control group. In addition, the diversity of intestinal microflora decreased and the community structure was uneven after FH535 treatment. In summary, our findings strongly suggest that ß-catenin plays a vital role in combating pathogen invasion and regulating intestinal flora in Qi river crucian carp.

Acanthopanax senticosus cultures fermented by Lactobacillus rhamnosus enhanced immune response through improvement of antioxidant activity and inflammation in crucian carp (Carassius auratus).

Lactic acid bacteria (LAB) have been recognized as safe microorganism that improve micro-flora disturbances and enhance immune response. A well-know traditional herbal medicine, Acanthopanax senticosus (As) was extensively utilized in aquaculture to improve growth performance and disease resistance. Particularly, the septicemia, skin wound and gastroenteritis caused by Aeromonas hydrophila threaten the health of aquatic animals and human. However, the effects of probiotic fermented with A. senticosus product on the immune regulation and pathogen prevention in fish remain unclear. Here, the aim of the present study was to elucidate whether the A. senticosus fermentation by Lactobacillus rhamnosus improve immune barrier function. The crucian carp were fed with basal diet supplemented with L. rhamnosus fermented A. senticosus cultures at 2 %, 4 %, 6 % and 8 % bacterial inoculum for 8 weeks. After trials, the weight gain rate (WGR), specific growth rate (SGR) were significantly increased, especially in LGG-6 group. The results confirmed that the level of the CAT, GSH-PX, SOD, lysozyme, and MDA was enhanced in fish received with probiotic fermented product. Moreover, the L. rhamnosus fermented A. senticosus cultures could trigger innate and adaptive immunity, including the up-regulation of the C3, C4, and IgM concentration. The results of qRT-PCR revealed that stronger mRNA transcription of IL-1ß, IL-10, IFN-γ, TNF-α, and MyD88 genes in the liver, spleen, kidney, intestine and gills tissues of fish treated with probiotic fermented with A. senticosus product. After infected with A. hydrophila, the survival rate of the LGG-2 (40 %), LGG-4 (50 %), LGG-6 (60 %), LGG-8 (50 %) groups was higher than the control group. Meanwhile, the pathological damage of the liver, spleen, head-kidney, and intestine tissues of probiotic fermentation-fed fish could be alleviated after pathogen infection. Therefore, the present work indicated that L. rhamnosus fermented A. senticosus could be regard as a potential intestine-target therapy strategy to protecting fish from pathogenic bacteria infection.

Excessive dietary iron exposure increases the susceptibility of largemouth bass (Micropterus salmoides) to Aeromonas hydrophila by interfering with immune response, oxidative stress, and intestinal homeostasis.

Iron is an essential cofactor in the fundamental metabolic pathways of organisms. Moderate iron intake can enhance animal growth performance, while iron overload increases the risk of pathogen infection. Although the impact of iron on the pathogen-host relationship has been confirmed in higher vertebrates, research in fish is extremely limited. The effects and mechanisms of different levels of iron exposure on the infection of Aeromonas hydrophila in largemouth bass (Micropterus salmoides) remain unclear. In this study, experimental diets were prepared by adding 0, 800, 1600, and 3200 mg/kg of FeSO4∙7H2O to the basal feed, and the impact of a 56-day feeding period on the mortality rate of largemouth bass infected with A. hydrophila was analyzed. Additionally, the relationships between mortality rate and tissue iron content, immune regulation, oxidative stress, iron homeostasis, gut microbiota, and tissue morphology were investigated. The results showed that the survival rate of largemouth bass infected with A. hydrophila decreased with increasing iron exposure levels. Excessive dietary iron intake significantly increased iron deposition in the tissues of largemouth bass, reduced the expression and activity of antioxidant enzymes superoxide dismutase, catalase, and glutathione peroxidase, increased the content of lipid peroxidation product malondialdehyde, and thereby induced oxidative stress. Excessive iron supplementation could influence the immune response of largemouth bass by upregulating the expression of pro-inflammatory cytokines in the intestine and liver, while downregulating the expression of anti-inflammatory cytokines. Additionally, excessive iron intake could also affect iron metabolism by inducing the expression of hepcidin, disrupt intestinal homeostasis by interfering with the composition and function of the gut microbiota, and induce damage in the intestinal and hepatic tissues. These research findings provide a partial theoretical basis for deciphering the molecular mechanisms underlying the influence of excessive iron exposure on the susceptibility of largemouth bass to pathogenic bacteria.

In vivo and ex vivo studies support the immunostimulant and immunoprotective effect of Damiana (Turnera diffusa Willd) in Almaco Jack (Seriola rivoliana).

Damiana (Turnera diffusa Willd) was evaluated in vitro for antioxidant and antibacterial activities against Staphylococcus aureus and Streptococcus pyogenes (as a preliminary screening assessment) by high-performance thin-layer chromatography (HPTLC)-Direct bioautography. A study was performed in vivo to evaluate the effects of Damiana enriched diets at 0.5 % on immune parameters in mucus and serum and gene expression in Almaco Jack (Seriola rivoliana) intestine after two and four weeks; an infection with Aeromonas hydrophila at 1x107 colony forming units (CFU) followed and an ex vivo study was carried out using head-kidney leukocytes. Ferric reducing ability of plasma (FRAP) and 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid (ABTS) assays showed high antioxidant activities in Damiana leaves; even in the ABTS assay, Damiana at 300 µg/mL showed similar activity to ascorbic acid - the standard control. Damiana exhibited strong in vitro antimicrobial activity against S. aureus and S. pyogenes. In vivo studies showed a strong enhancement of myeloperoxidase, nitric oxide, superoxide dismutase, and catalase activities in mucus and serum of S. rivoliana supplemented with Damiana; their immunological response enhanced after infection with A. hydrophila. IL-1ß, TNF-α, and IL-10 gene expressions upregulated in the fish intestine challenged with the bacterium. Piscidin and macrophage (MARCO) receptor gene expression up-regulated at week 4 and down-regulated after infection. Intestinal histology results confirm that Damiana not cause inflammation or damage. Finally, the ex vivo study confirmed the immunostimulant and protective effects of Damiana through increased phagocytic, respiratory burst, myeloperoxidase activities and nitric oxide generation before and upon the bacterial encounter. These results support the idea that Damiana has the potential as an immunostimulant additive for diets in aquaculture by enhancing immune parameters and protecting Almaco Jack against A. hydrophila infections upon four weeks of supplementation.

Chicken egg lysozyme enhanced the growth performance, feed utilization, upregulated immune-related genes, and mitigated the impacts of Aeromonas hydrophila infection in Nile tilapia (Oreochromisniloticus).

Functional supplements, including lysozyme, are highly approved as immunostimulant and antibacterial agents with a high potential for use in aquaculture. In this regard, Nile tilapia was treated with lysozyme at 0, 0.5, 1, 1.5, and 3 g/kg for 60 days, then challenged with Aeromonas hydrophila. Fish were stocked in 15 glass aquaria (70 L each) with an equal initial weight of 10.72 ± 0.71 g per fish and 15 fish per aquarium. The regression analysis revealed that dietary lysozyme supplementation at 1.83-2 g/kg enhanced the growth performance, protein efficiency ratio, and protein productive value while reducing the feed conversion ratio of tilapia. Markedly, tilapia treated with lysozyme had a low mortality rate (30-50 %) compared to the control, which recorded a 70 % mortality rate after 15 days of challenge with A. hydrophila. The regression analysis also revealed that the highest lysozyme activity of tilapia-fed lysozyme for 60 days is achieved by 2.05 g/kg lysozyme. The expression of Nf-κb, IL-1ß, and IL-8 genes is upregulated in tilapia-fed lysozyme at 0.5, 1, 1.5, and 3 g/kg for 60 days before and after A. hydrophila infection. The expression of GPX and CAT genes was higher in tilapia-fed lysozyme at 0.5, 1, 1.5, and 3 g/kg for 60 days before and after A. hydrophila infection. Before infection, the relative transcription of the lysozyme and C3 was upregulated in tilapia-fed lysozyme at 0.5, 1, 1.5, and 3 g/kg. However, lysozyme gene expression in tilapia treated with 0.5 g/kg lysozyme had no significant differences from those fed 0 g/kg lysozyme. After infection, the relative transcription of the lysozyme gene was upregulated in tilapia fed 1 and 1.5 g/kg, while tilapia fed 1 g/kg lysozyme had the highest C3 gene transcription. After infection, the hepatocytes in the livers of fish fed 0 g/kg lysozyme exhibited a noticeable fatty alteration, along with congestion, a light infiltration of inflammatory cells, and the start of necrosed cell regeneration. However, the livers of fish that received lysozyme were normal except for infiltrations of perivascular and interstitial mononuclear cells, depending on the supplementation dose. In conclusion, dietary lysozyme is recommended at 1.83-2.05 g/kg to gain high growth performance, immune response, and high resistance to A. hydrophila in Nile tilapia.

Chlorella vulgaris extract conjugated magnetic iron nanoparticles in nile tilapia (Oreochromis niloticus): Growth promoting, immunostimulant and antioxidant role and combating against the synergistic infection with Ichthyophthirius multifiliis and Aeromonashydrophila.

Nile tilapia reared under intensive conditions was more susceptible for Ichthyophthirius multifilii (I. multifiliis) infection eliciting higher mortality, lower productive rate and further bacterial coinfection with Aeromonas hydrophila (A. hydrophila). The higher potency of magnetic field of iron oxide nanoparticles (NPs) can kill pathogens through inhibiting their viability. Herein, coating of Chlorella vulgaris extract (ChVE) with magnetic iron oxide NPs (Mag iron NPs) can create an external magnetic field that facilitates their release inside the targeted tissues. Thus, the current study is focused on application of new functionalized properties of Mag iron NPs in combination with ChVE and their efficacy to alleviate I. multifiliis and subsequent infection with A. hydrophila in Nile tilapia. Four hundred fingerlings were divided into: control group (with no additives), three groups fed control diet supplemented with ChVE, Mag iron NPs and ChVE@Mag iron NPs for 90 days. At the end of feeding trial fish were challenged with I. multifiliis and at 9 days post challenge was coinfected by A. hydrophila. A remarkable higher growth rate and an improved feed conversion ratio were detected in group fed ChVE@Mag iron-NPs. The maximum expression of antioxidant enzymes in skin and gills tissues (GSH-Px, CAT, and SOD) which came in parallel with higher serum activities of these enzymes was identified in groups received ChVE@Mag iron-NPs. Furthermore, group fed a combination of ChVE and Mag iron-NPs showed a boosted immune response (higher lysozyme, IgM, ACH50, and MPO) prior to challenge with I. multifiliis. In contrast, fish fed ChVE@Mag iron-NPs supplemented diet had lower infection (decreased by 62%) and mortality rates (decreased by 84%), as well as less visible white spots (decreased by 92 % at 12 dpi) on the body surfaces and mucous score. Interestingly, post I. multifiliis the excessive inflammatory response in gill and skin tissues was subsided by feeding on ChVE@Mag iron-NPs as proved by down regulation of IL-1ß, TNFα, COX-2 and iNOS and upregulation of IL-10, and IgM, IgT and Muc-2 genes. Notably, group exposed to I. multifiliis-showed higher mortality when exposed to Aeromonas hydrophilia (increased by 43 %) while group fed ChVE@Mag iron-NPs exhibited lower morality (2%). Moreover, the bacterial loads of A. hydrophilia in fish infected by I. multifiliis and fed control diet were higher than those received dietary supplement of ChVE, Mag iron-NPs and the most reduced load was obtained in group fed ChVE@Mag iron-NPs at 7 dpi. In conclusion, ChVE@Mag iron-NPs fed fish had stronger immune barrier and antioxidant functions of skin and gills, and better survival following I. multifiliis and A. hydrophilia infection.

Effect of dietary koumine on the immune and antioxidant status of carp (Cyprinus carpio) after Aeromonas hydrophila infection.

Koumine (KM) has anxiolytic, anti-inflammatory and growth-promoting effects in pigs and sheep. Based on the growth-promoting and immunological effects of koumine, the present study was conducted on Cyprinus carpio (C. carpio) with four KM concentrations: 0 mg/kg, 0.2 mg/kg, 2 mg/kg, and 20 mg/kg for 10 weeks, followed by a 1-week Aeromonas hydrophila (A. hydrophila) infection experiment. The effect of KM on the immunity of A. hydrophila infected carp was analyzed by histopathology, biochemical assay, and qRT-PCR to assess the feasibility of KM in aquaculture. The results showed that the presence of KM alleviated pathogen damage to carp tissues. At 2 mg/kg and 20 mg/kg concentrations of KM successively and significantly elevated (p < 0.05) the SOD activities in the intestinal tract, hepatopancreas and kidney of carp. The expression levels of hepatopancreatic antioxidant genes Nrf2 and IGF-1 were significantly up-regulated in the same group (p < 0.05), while the expression levels of immune genes IL-8 and IL-10 were down-regulated. In summary, KM at concentrations of 2 mg/kg and 20 mg/kg could regulate the expression of antioxidant and immune genes in various tissues in an orderly and rapid manner, and significantly improve the antioxidant and immune abilities of carp, which is conducive to the improvement of the resilience of carp.

Sweet orange peel waste as a feed additive in growth promoting and protective effect against Aeromonas hydrophila of juvenile bagrid catfish Mystus nemurus.

Sweet orange Citrus sinensis peel is a phytobiotic agricultural waste with bioactive compounds that have potential functional properties as a growth promoter and immune stimulator. This study aims to evaluate the dietary effects of sweet orange peel (SOP) as a feed additive on growth enhancement of juvenile bagrid catfish Mystus nemurus and their disease resistance ability against Aeromonas hydrophila infection. Four experimental diets were formulated to contain 0 (SOP0, control), 4 (SOP4), 8 (SOP8) and 12 g/kg (SOP12) SOP. After 90 d of the feeding experiment, improvement in weight gain, specific growth rate, feed conversion ratio, and protein efficiency ratio were observed in the fish fed with SOP4. While fish survival was not significantly affected, hepatosomatic and viscerosomatic indices were significantly higher in fish fed with SOP12. Muscle protein was higher in fish fed with SOP4, SOP8, and SOP12 than in control but muscle lipids showed an opposite trend. A 14-d post-challenge test against A. hydrophila revealed no significant effect on the fish survival. Nevertheless, fish fed SOP4 encountered delayed bacterial infection compared to other treatments and fish fed with SOP0 and SOP4 performed numerically better survival. Infected fish showed skin depigmentation, haemorrhagic signs at the abdomen and anus, internal bleeding, and stomach and intestine enlargement. In conclusion, SOP4 could be recommended as a growth promoter while slightly delaying A. hydrophila infection in M. nemurus.

Potentials of Aloe barbadensis inclusion in fish feeds on resilience to Aeromonas hydrophila infection in freshwater fish Labeo rohita.

Aquatic bacterial pathogens can cause severe economic loss in aquaculture industry. An opportunistic pathogen, Aeromonas hydrophila is responsible for Motile Aeromonas Septicemia, leading to high mortality rates in fish. The present study was focused on the efficacy of Aloe barbadensis replacing fishmeal diets on hematological, serum biochemical, antioxidant, histopathological parameters, and disease resistance against A. hydrophila infection in Labeo rohita. Isonitrogenous fishmeal replaced diets (FMR) were prepared with varying levels of A. barbadensis at D1 (0%) (control), D2 (25%), D3 (50%), D4 (75%) and D5 (100%) then fed to L. rohita. After 60 days of post-feeding, the experimental fish were challenged with A. hydrophila. Blood and organs were collected and examined at 1- and 15-days post infection (dpi). The results demonstrated that on 1 dpi, white blood cells (WBC), total protein, cholesterol and low-density lipoprotein (LDL) levels were significantly increased in D3 diet fed groups. The D2 and D3 diet fed group showed decreasing trends of serum glutamic pyruvic transaminase (SGPT) and antioxidant enzymes activity on 15 dpi. The histopathological architecture results clearly illustrated that the D3 diet fed group had given a higher protective effect by reducing the pathological changes associated with A. hydrophila infection in liver, intestine and muscle. Higher percentage of survival rate was also observed in D3 diet fed group. Therefore, the present study suggested that the dietary administration of A. barbadensis up to 50% fishmeal replacement (D3 diet) can elicit earlier antioxidant activity, innate immune response and improve survival rate in L. rohita against A. hydrophila infection.

Effects of the combination of chitosan and Acinetobacter KU011TH on the growth and health performances and disease resistance of juvenile hybrid catfish (Clarias gariepinus × C. macrocephalus).

Aquatic animal health management has become a crucial component in the goal of increasing catfish aquaculture productivity. Additionally, hybrid catfish (Clarias gariepinus × C. macrocephalus) has been promoted as a highly profitable freshwater fish in Asia. Interestingly, the crucial diseases induced by Aeromonas hydrophila have been reported to greatly impede catfish production. To overcome this challenge, the aim was to investigate the effects of the oral administration of potentially synbiotic chitosan (CH) and Acinetobacter KU011TH (AK) on the growth performance, immunological responses, and disease resistance of hybrid catfish against A. hydrophila. The control group was fed a basal diet (A), the diet fed to treatment group B was supplemented with 20 mL of CH/kg diet (B), and the experimental feed fed to groups C-D was mixed with 1 × 108, 1 × 109 and 1 × 1010 CFU/mL AK coated with 20 mL of CH/kg diet. Five different groups of juvenile hybrid catfish were continuously fed the 5 formulated feeds for 4 weeks. The results revealed that all tested feeds did not significantly enhance the hybrid catfish's average daily gain, specific growth rate, feed conversion ratio, hematocrit and erythrocyte counts. Interestingly, the application of CH and AK significantly increased the leukocyte counts, respiratory burst, lysozyme activity, alternative complement pathway hemolytic activity, and bactericidal activity (P < 0.05). The expression levels of the immune-related genes in the whole blood, head kidney, and spleen were significantly increased after CH-AK application (P < 0.05), but this finding was not observed in the liver (P > 0.05). Additionally, after 14 days of A. hydrophila peritoneal injection, the fish in group C showed significantly higher survival rates of approximately 70.0 % compared with the control fish in groups B, D, and E (52.5 %, 40.0 %, 45.0 %, and 45.0 %, respectively) (P < 0.05). These results collectively suggest that short-term application of the diet fed to group C effectively boosted the immune responses and disease resistance of hybrid catfish against A. hydrophila.

Silymarin enhances the response to oxytetracycline treatment in Oreochromis niloticus experimentally infected with Aeromonas hydrophila.

Many governments have approved the use of oxytetracycline as an antibiotic additive to food fish, with oxytetracycline now routinely used in many nations. However, oxytetracycline is known to have immunosuppression impacts. We, therefore, evaluated the immunological, antioxidative, and histopathological status of Nile tilapia fed a diet containing silymarin (100 mg/kg fish feed) for 0, 2, 4, 6, and 8 weeks. The protective effects of silymarin against Aeromonas hydrophila (A. hydrophila) infection and oxytetracycline treatment were evaluated. Blood parameters (erythrocyte count, white blood cell count, hemoglobin, and packed cell volume) improved over time in fish fed on dietary silymarin. Serum levels of alanine aminotransferase (ALT) were lower in fish fed on dietary silymarin, whereas serum levels of aspartate transferase (AST)and alkaline phosphatase (ALK) were unchanged. Dietary silymarin affected serum lipid profiles as decreases in serum triglyceride and low-density lipoprotein cholesterol levels and a trend toward lower cholesterol levels, whereas serum high-density lipoprotein cholesterol levels were increased compared to fish fed on the control diet. Dietary silymarin resulted in an increase of serum total protein levels and globulin fractions. Significant and progressive increases in catalase and glutathione peroxidase levels were observed after six weeks of feeding on a dietary silymarin before decreasing to control levels at the end of the experimental period. Fish fed on dietary silymarin, interleukin-1 and fish tumor necrosis factor-alpha were upregulated in hepatic tissues; however, interleukin-10 levels decreased to comparable levels to controls after eight weeks. Fish infected with A. hydrophila displayed septicemia (opaque eye, hemorrhagic ulcers, dentated fins, hepatomegaly, and splenomegaly). Reduced mortality was observed in Nile tilapia infected with A. hydrophila and fed a diet containing silymarin, indicating that silymarin improves fish responses to oxytetracycline with a 37% reduction in mortality.

Growth performance, hematology parameters and Aeromonas hydrophila challenge of pirarucu (Arapaima gigas) in response to dietary protein levels.

The pirarucu (Arapaima gigas) is a fast-growing Amazonian species of high commercial value. The present study aimed to determine the dietary crude protein (CP) level to promote maximum zootechnical performance for pirarucu fingerlings and as their resistance to Aeromonas hydrophila, as well as evaluate their hematological parameters. Pirarucu fingerlings (2.4 ± 0.08 g, 6.8 ± 0.52 cm) were distributed in 18 tanks (140 L, 40 fish per tank, n = 3) and fed six experimental diets consisting of increasing levels of CP: 300, 400, 450, 500, 550 and 650 g kg-1 in a completely randomized design. Data were submitted to one-way ANOVA (p < 0.05) and the ideal CP level for weight gain was determined using polynomial regression analysis. The dietary CP levels were evaluated using a quadratic polynomial regression and the level of 595 g kg-1 was determined for the best weight gain. The hematocrit of fish fed 300 g kg-1 was higher than in the other groups. No mortalities were observed after the 15-day bacterial challenge; however, number of pirarucu with bacterial damage on the pirarucu caudal fin was higher in the group that was fed the diet with 300 g kg-1. A dietary protein level of 618 g kg-1 is therefore recommended for providing maximum weight gain and immunological resistance in pirarucu fingerlings weighing 2.4-112.5 g.

Vitamin D Promotes Mucosal Barrier System of Fish Skin Infected with Aeromonas hydrophila through Multiple Modulation of Physical and Immune Protective Capacity.

The vertebrate mucosal barrier comprises physical and immune elements, as well as bioactive molecules, that protect organisms from pathogens. Vitamin D is a vital nutrient for animals and is involved in immune responses against invading pathogens. However, the effect of vitamin D on the mucosal barrier system of fish, particularly in the skin, remains unclear. Here, we elucidated the effect of vitamin D supplementation (15.2, 364.3, 782.5, 1167.9, 1573.8, and 1980.1 IU/kg) on the mucosal barrier system in the skin of grass carp (Ctenopharyngodon idella) challenged with Aeromonas hydrophila. Dietary vitamin D supplementation (1) alleviated A. hydrophila-induced skin lesions and inhibited oxidative damage by reducing levels of reactive oxygen species, malondialdehyde, and protein carbonyl; (2) improved the activities and transcription levels of antioxidant-related parameters and nuclear factor erythroid 2-related factor 2 signaling; (3) attenuated cell apoptosis by decreasing the mRNA and protein levels of apoptosis factors involved death receptor and mitochondrial pathway processes related to p38 mitogen-activated protein kinase and c-Jun N-terminal kinase signaling; (4) improved tight junction protein expression by inhibiting myosin light-chain kinase signaling; and (5) enhanced immune barrier function by promoting antibacterial compound and immunoglobulin production, downregulating pro-inflammatory cytokine expression, and upregulating anti-inflammatory cytokines expression, which was correlated with nuclear factor kappa B and the target of rapamycin signaling pathways. Vitamin D intervention for mucosal barrier via multiple signaling correlated with vitamin D receptor a. Overall, these results indicate that vitamin D supplementation enhanced the skin mucosal barrier system against pathogen infection, improving the physical and immune barriers in fish. This finding highlights the viability of vitamin D in supporting sustainable aquaculture.

Fructooligosaccharide and Bacillus subtilis synbiotic combination promoted disease resistance, but not growth performance, is additive in fish.

Species diversification from major to minor carps for their sturdiness and initial higher growth, and also a quest for antibiotic-free aqua farming in the subcontinent, mandates search for and evaluation of alternatives. An experiment was performed to investigate the potential of fructooligosaccharide (FOS) and Bacillus subtilis (BS) (alone or as synbiotics) in promoting growth and immunity against infections in Labeo fimbriatus fingerlings. Six iso-nitrogenous and iso-lipidic diets containing combinations of two levels of FOS (0% and 0.5%) and three levels of BS (0, 104, 106 CFU/g feed) were fed to fish for 60 days. At the end of the feeding trial, twenty-four fish from each group were injected intra-peritoneally with pathogenic strain of Aeromonas hydrophila O:18 to test the immunoprotective efficacy of the supplements against bacterial infection. BS, but not FOS, significantly improved (P < 0.05) growth and feed utilisation attributes like percentage weight gain (PWG), specific growth rate (SGR) and feed conversion ratio (FCR). There were interactive effects of FOS and BS on PWG, SGR and FCR; however, the effects were not additive in nature. These beneficial effects of BS, alone or in combination with FOS, were corroborated by increased protease activity, microvilli density and diameter and number of goblet cells. Overall beneficial effects of FOS and BS included improved erythrocyte (RBC), hemoglobin (Hb), total protein and globulin levels. Total leucocyte (WBC) count and immunological parameters like respiratory burst activity of leucocytes (NBT reduction), lysozyme activity, albumin: globulin ratio and post-challenge survival were significantly improved by both FOS and BS, and their dietary combination yielded the highest improvement in these parameters. Synergistic effects of FOS and BS as dietary supplements indicate that a combination of 106 CFU/g BS and 0.5% FOS is optimal to improve growth, feed utilisation, immune functions, and disease resistance in L. fimbriatus fingerlings.

Dietary Bacillus velezensis R-71003 and sodium gluconate improve antioxidant capacity, immune response and resistance against Aeromonas hydrophila in common carp.

This study aimed to evaluate the effects of dietary supplementation with Bacillus velezensis R-71003 combined with sodium gluconate on antioxidant capacity, immune response and resistance against Aeromonas hydrophila in common carp. In addition, the biocontrol potential of the secondary metabolites of B. velezensis R-71003 was also evaluated to analyze the possible mechanism of B. velezensis R-71003 against A. hydrophila. The results indicated that the antibacterial crude extract of B. velezensis R-71003 can destroy the cell wall of A. hydrophila. Moreover, the results showed that dietary B. velezensis R-71003 could promote antioxidant capacity, which significantly increased the activities of CAT and SOD and decreased the content of MDA. Additionally, B. velezensis R-71003 supplementation significantly enhanced the immunity of common carp, as measured by the mRNA expression levels of cytokine-related genes (TNF-α, TGF-ß, IL-1ß and IL-10). In addition, dietary B. velezensis R-71003 exhibited an upregulation of IL-10 and a downregulation of IL-1ß, coupled with higher survival rates when challenged with A. hydrophila compared to the positive group. Furthermore, compared to prechallenge, the mRNA expression levels of TLR-4, MyD88, IRAK1, TRAF6, TRIF and NF-κB in the head kidney of common carp were significantly increased after challenge. The fish fed the B. velezensis R-71003 diet showed lower expression of TLR-4, MyD88, IRAK1, TRAF6, TRIF and NF-κB after the challenge than those fed the control diet. Thus, this study revealed that B. velezensis R-71003 can improve the resistance of common carp to pathogenic bacteria by destroying bacterial cell walls and improving fish immunity by activating the TLR4 signaling pathway. Importantly, this study indicated that sodium gluconate has a positive effect on B. velezensis R-71003 in enhancing the anti-infection ability of common carp. The results of this study will lay the foundation for the application of B. velezensis R-71003 in combination with sodium gluconate as an alternative to antibiotics in aquaculture.

Characterization of hepcidin gene and protection of recombinant hepcidin supplemented in feed against Aeromonas hydrophila infection in Yellow River carp (Cyprinus carpio haematopterus).

Hepcidin is a small peptide of defensins with antibacterial activity, and plays an important role in innate immunity against pathogenic microorganisms, which can also participate in the regulation of iron metabolism. The hepcidin gene in Yellow River carp (Cyprinus carpio haematopterus) (CcHep) was cloned and identified. The total length of CcHep cDNA was 480 bp, containing an open reading frame (ORF) that encoded 91 amino acids (aa), which contained a 24-aa signal peptide, a 42-aa propeptide, and a 25-aa mature peptide. The mature peptide had a typical RX (K/R) R motif and eight conserved cysteine residues forming four pairs of disulfide bonds. Homology and phylogenetic tree analysis showed that CcHep had the closest relationship with that of crucian carp. The expression levels of hepcidin mRNA in healthy and Aeromonas hydrophila stimulated fish were measured by real-time fluorescence quantitative PCR. The results showed that CcHep mRNA was expressed in different tissues of healthy fish with the highest relative expression level in liver, followed by kidney and intestine, and the lowest expression level was observed in heart. The hepcidin gene was extremely significantly up-regulated in head kidney, intestine, liver, skin, spleen, and gill at 6 h and 12 h after A. hydrophila infection. Furthermore, the immunoregulation effect of dietary recombinant protein was evaluated. The recombinant hepcidin protein (rCcHep) was successfully expressed by Pichia pastoris X-33 and showed strong antibacterial activity against A. hydrophila, Escherichia coli, Vibrio anguillarum and Bacillus subtilis in vitro. In order to evaluate the preventive effect of rCcHep, fish were fed with basal diet or diet supplemented with different doses of rCcHep, and then challenged with A. hydrophila. The results showed that immune genes were up-regulated to varying degrees, and feed additive groups exhibited a significantly improved up-regulation expressions of Lysozyme, Toll-like receptor 5 (TLR 5), Major histocompatibility complex classⅡ (MHCⅡ), while inhibited up-regulation expressions of Interleukin 1ß (IL-1ß), Interleukin 8 (IL-8), and Tumor necrosis factor α (TNF-α) in liver and spleen compared to the control. Meanwhile, the relative immune protection rate in 120 mg/kg feed additive group was 28%, and the bacterial clearance rate in tissues of this group was higher than that of the control. Collectively, these results indicated that rCcHep had antibacterial activity and showed an immune protection effect against A. hydrophila, and could be considered as a dietary supplement to apply in aquaculture.

Dietary Syzygium cumini leaf extract influences growth performance, immunological responses and gene expression in pathogen-challenged Cyprinus carpio.

This study evaluated the effects of jamun leaf extract (JLE) as a feed supplement on growth performance, haemato-immunological, oxidative stress-related parameters, and cytokine gene expression in Cyprinus carpio challenged with Aeromonas hydrophila.. Diets containing four different JLE concentrations, that is, 0 (basal diet), 5 (JLE5), 10 (JLE10), and 15 g kg-1 (JLE15), were fed to carp (6.17 ± 0.43 g) for eight weeks. Growth performance was significantly higher in JLE10. Haemato-immunological and antioxidant parameters were determined in fish at 48 h post-challenge with A. hydrohila. The cumulative survival was highest in JLE10 (69.69%) 14 days post-challenge. Serum protein (2.18 ± 0.06 g dL-1), lysozyme (32.38 ± 1.2 U mL-1), alternative complement pathway (70.43 ± 1.61 U mL-1), phagocytic activity (21.18 ± 0.48%), respiratory burst activity (0.289 ± 0.09 OD630nm), and immunoglobulin levels (6.67 ± 0.36 U mg mL-1) were significantly higher in JLE10 than in the control. Serum alanine aminotransferase (44.06 ± 1.62 Unit mL-1), aspartate aminotransferase (31.58 ± 1.82 Unit mL-1), and malondialdehyde (2.57 ± 0.26 nmol mL-1) levels were lower in JLE10 than in the control (p < 0.05), whereas myeloperoxidase activity was significantly higher in JLE5 and JLE10 than in the control. Superoxide dismutase levels in the serum were higher (p < 0.05) in JLE5 and JLE10 than in the other groups. Gene expression analysis revealed that the mRNA expression of pro-inflammatory cytokines TNF-α and IL-1ß was upregulated (p < 0.05) in the liver, head-kidney, and intestine of challenged carp in JLE10. The signalling molecule NF-κB p65 was upregulated in lymphoid organs in JLE10 but not in the liver. The anti-inflammatory cytokine IL-10 was significantly downregulated in challenged carp in JLE10 compared with that in the control. Quadratic regression analysis showed that optimal dietary JLE was estimated to be 9.03-10.15 g kg-1 to maximize the growth performance. Results of the present study revealed that dietary JLE at 10 g kg-1 can significantly improve the immunity and disease resistance of C. carpio. Thus, JLE is a promising food additive for carp aquaculture.

The influence of dietary Coenzyme Q10 on growth performance, antioxidant capacity and resistance against Aeromonas hydrophila of juvenile European eel (Anguilla anguilla).

The present study was conducted to investigate the effects of dietary Coenzyme Q10 (CoQ10) on the growth performance, body composition, digestive enzyme activity, antioxidant capacity, intestinal histology, immune-antioxidant gene expression and disease resistance of juvenile European eel (Anguilla anguilla). Fish were fed a diet supplemented with CoQ10 at concentrations of 0, 40, 80 and 120 mg/kg for 56 days. The results indicated that dietary CoQ10 supplementation did not significantly affect final body weight (FBW), survival rate (SR), weight gain (WG), feed rate (FR), viscerosomatic index (VSI) or hepatosomatic index (HSI) among all experimental groups. However, the highest FBW, WG and SR were found in the 120 mg/kg CoQ10 group. Dietary 120 mg/kg CoQ10 markedly improved feed efficiency (FE) and the protein efficiency ratio (PER). The crude lipid in the body and triglycerides (TG) and total cholesterol (TC) in serum were obviously lower in the 120 mg/kg CoQ10 group than in the control group. For digestive enzymes, protease activity in the intestine was markedly boosted in the 120 mg/kg CoQ10 group. The serum activities of SOD, CAT and GST in the 120 mg/kg CoQ10 group were significantly higher than those in the control group. Dietary 120 mg/kg CoQ10 efficiently enhanced superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) and glutathione S-transferase (GST) activities in the liver, while the malondialdehyde (MDA) content was significantly decreased. No significant histological changes in the liver were identified in any group. Dietary supplementation with 120 mg/kg CoQ10 improved antioxidant capacity and immunity by upregulating the expression of cyp1a, sod, gst, lysC, igma1, igmb1 and irf3 in the liver. Furthermore, the cumulative survival rate of juvenile European eel against challenge with Aeromonas hydrophila was significantly elevated in the 80 and 120 mg/kg CoQ10 supplemented groups. Conclusively, our study suggested that supplementing the diet of juvenile European eel with CoQ10 at a concentration of 120 mg/kg could promote their feed utilization, fat reduction, antioxidant capacity, digestibility, immune-antioxidant gene expression and resistance to Aeromonas hydrophila without negative effects on fish health status.