epiglucan and Gram-Negative-Bacterial-Infections

The innate immune response is able to ward off pathogens and remember previous infections using different mechanisms; this kind of immune reaction has been called "trained immunity". Changes in cellular metabolism (aerobic glycolysis) have been observed during training with some immunostimulants like β-glucans or during viral and bacterial infections. We hypothesize that β-glucans can induce metabolic changes used by the host to fight pathogens. Accordingly, we evaluated changes in metabolic parameters in turbot that could affect their survival after a previous intraperitoneal treatment with β-glucans and subsequent administration of Viral Hemorrhagic Septicemia Virus (VHSV) or bacteria (Aeromonas salmonicida subsp. salmonicida). The results obtained support that β-glucans, VHSV and A. salmonicida induce changes in lactate, glucose and ATP levels in plasma, head kidney and liver and in the mRNA expression of enzymes related to glucose and fatty acid metabolism in head kidney. Additionally, the metabolic changes induced by β-glucans are beneficial for VHSV replication, but they are harmful to A. salmonicida, resulting in reduced mortality. β-glucans appear to have great therapeutic potential and can induce trained immunity against bacterial disease but not against viral disease, which seems to take advantage of β-glucan metabolic alterations.

Topics: Adjuvants, Immunologic; Aeromonas salmonicida; Animals; beta-Glucans; Fish Diseases; Flatfishes; Gram-Negative Bacterial Infections; Longevity; Novirhabdovirus; Random Allocation; Rhabdoviridae Infections

An eight-week feeding trial was performed to assess the effect of different dietary levels (0, 0.5, 1.0, and 1.5%) of β-glucan (sourced from Saccharomyces cerevisiae) on growth, survival, immunological parameters (immune gene expression, lysozyme, and antiprotease), total antioxidant status, thermal tolerance, and disease resistance of Tor putitora fry. Feeding of moderate doses (0.5 and 1.0%) of β-glucan significantly improved survival but not weight gain percentage as compared to that received unsupplemented control and highest dose (1.5%) of glucan. Supplementation of β-glucan in diets differentially influenced the mRNA expression of cytokine and other immune genes. For instance, transcripts of cytokines such as tnf-α and il-1β were significantly upregulated, while ifn-γ and il-10 were unaffected by β-glucan intake. Also, the relative mRNA expression of tlr-5 and hepcidin1 along with lysozyme and antiprotease activities were remained largely unchanged by dietary glucan administration. In contrast, β-glucan induced mRNA expression of defensin1 and c3 while decreased the transcript level of mhc-1. On the other hand, dietary inclusion of β-glucan markedly improved total antioxidant levels and extended the thermal tolerance limits at both the ends, as shown by increased CT

Topics: Aeromonas salmonicida; Animal Feed; Animals; Antioxidants; beta-Glucans; Cyprinidae; Disease Resistance; Fish Diseases; Gene Expression; Gram-Negative Bacterial Infections; Muramidase; Protease Inhibitors; RNA, Messenger

The effects of dietary β-glucan on innate immune responses have been shown in a number of different vertebrate species. However, there is conflicting information about the period of administration (shorter vs. longer), and it is also unclear to what extent β-glucan's effects can be observed post-treatment in fish. Thus, we fed Nile tilapia for 0 (control group; 45 days of control diet), 15 (30 days of control followed by 15 days of β-glucan), 30 (15 days of control followed by 30 days of β-glucan) or 45 days with a diet containing 0.1% of β-glucan (MacroGard®). We evaluated the growth performance at the end of the β-glucan feeding trial and the innate immune function immediately after the feeding trial and 7 and 14 days post-feeding trial. In addition, at day 10 post-feeding trial, we assessed the tilapia's resistance against a bacterial infection. No significant differences were observed in growth performance between the groups; however, fish fed with β-glucan for 30 and 45 days had higher (approx. 8%) relative weight gain compared to the control. Regardless of the administration period, fish fed with β-glucan had higher innate immune responses immediately after the feeding trial such as lysozyme activity in plasma, liver and intestine and respiratory burst compared to the control, and in general these differences were gradually reduced over the withdrawal period (up to 14 days). No differences were observed in the plasma hemolytic activity of the complement or myeloperoxidase activity in plasma or intestine. Moreover, fish from the control group had early mortalities (2 vs. 4-5 days post-infection, respectively) and a lower survival rate (60 vs. 80%, respectively) compared to fish fed with β-glucan for 15 or 30 days, and, interestingly, fish fed for 45 days with β-glucan had no mortality. This study indicates that regardless of the administration period (i.e., 15 up to 45 days), the β-glucan improved the innate immune responses and the tilapia's resistance to disease, and this protection could be observed up to 10 days post-feeding trial, adding in vivo evidence that β-glucan may contribute to a trained innate immunity. Additionally, we showed that a longer period of administration did not cause immunosuppression as previously hypothesized but promoted further growth and immune performance. These findings are relevant to the aquaculture industry and demonstrate that a longer β-glucan feeding protocol may be considered to achieve better results.

Topics: Aeromonas; Animal Feed; Animals; beta-Glucans; Cichlids; Diet; Dietary Supplements; Disease Resistance; Fish Diseases; Gram-Negative Bacterial Infections; Immunity, Innate; Random Allocation; Streptococcal Infections; Streptococcus agalactiae

Group II C-type lectin domain (CTLD) containing receptors belong to a large family of pattern recognition receptors which mainly act on the innate immunity. They are structurally related and consist of a cytoplasmic immunoreceptor tyrosine-based inhibitory motif (ITIM) and a single extracellular CTLD. Although they have been described in teleost fish, their involvement in immune responses is not well understood. In this study, four immune-related lectin-like receptors (termed CiILLR1 and CiILLR5-7), belonging to the group II CTLD receptors, were identified in grass carp (Ctenopharyngodon idella). They contain a short cytoplasmic tail and a single CTLD in the extracellular region. The CiILLR1 has a WxHxxxxxY motif similar to the WxHxxxxY motif which is required for the recognition of β-glucans by some of the group II CTLD containing lectins in mammals. Further, a modified QPD motif (EPD) known to be involved in binding to carbohydrate ligands is present in the CiILLR1, 5 and 6. However, CiILLR7 lacks these motifs. Expression analysis revealed that they were constitutively expressed in the head kidney and spleen. Moreover, CiILLR1, 5 and 6 could be up-regulated in the head kidney and spleen of fish after infection with Flavobacterium columnare and in the primary head kidney leukocytes by LPS and PHA. Expression of CiILLR1, CiILLR5 and CiILLR6 were mainly detected in the enriched lymphocytes whilst CiILLR7 was expressed in the enriched monocytes/macrophages. The results expand existing knowledge on the immune responses of the C-type lectin receptors in teleost fish.

Topics: Amino Acid Sequence; Animals; beta-Glucans; Carbohydrates; Carps; Fish Diseases; Fish Proteins; Flavobacterium; Gram-Negative Bacterial Infections; Head Kidney; Immunity, Innate; Lectins, C-Type; Leukocytes; Ligands; Lymphocytes; Macrophages; Monocytes; Sequence Alignment; Signal Transduction; Spleen; Up-Regulation

We evaluated the immune response of pacu fed with a β-glucan diet (0.5%) for 10 days. After the feeding period, fish were subjected to handling and 3 h after, inoculated with Aeromonas hydrophila. Fish were sampled before handling (baseline condition), 3, 6, and 24 h and 1 week after inoculation. A higher level of blood glucose was found in fish treated with β-glucan in baseline conditions. Handling and bacterial inoculation increased the circulating levels of cortisol and glucose and promoted the acute inflammatory response (lymphopenia and neutrophilia). β-Glucan prevented the decrease in the respiratory activity of leukocytes observed in the control group at 3 h sampling. β-Glucan did not affect the complement and lysozyme, which were activated 24 h after the bacterial challenge in control fish. A reduction in the number of leukocytes was found in fish treated with β-glucan 1 week after the challenge. We suggest two plausible hypotheses for this event: (1) it could be attributed to a depletion of the immune responses or (2) it could be due to a mobilization of the leukocytes to the spleen for antigen presenting/processing. In general, β-glucan avoided the reduction of the activity of leukocytes after stress and the bacterial challenge and increased the baseline glucose levels. Our findings confirm the immunomodulatory action of glucan and add evidence showing that glucan can have a role in stress response.

Topics: Aeromonas hydrophila; Animals; beta-Glucans; Blood Glucose; Characiformes; Fish Diseases; Gram-Negative Bacterial Infections; Leukocytes

To study the mechanism of β-glucan in immune protection, rainbow trout were fed diets with or without 0.2% β-glucan for 42 days and then infected with Aeromonas salmonicida. After that, spleen tissues were sampled on 4- and 6-days post infection (dpi). Transcriptome analysis was compared between control group (CG, without β-glucan addition) and 0.2% β-glucan group (BG). In CG vs BG, 378 and 406 DEGs were identified on 4 dpi and 6 dpi respectively; furthermore, 46 DEGs were shared on 4 dpi and 6 dpi, enriching in GO terms, such as complement activation, inflammatory response, and metabolic process. KEGG pathway analysis revealed that some DEGs in CG vs BG were involved in immune or metabolic signaling pathways such as complement and coagulation cascades, toll-like receptor signaling pathway, NF-κB signaling pathway, antigen processing and presentation, and platelet activation on 4 or 6 dpi. DEGs, such as fgg, fgb, f5, c9, c3, c5, tlr5, and myd88, were analyzed in CG vs BG on 4 dpi and 6 dpi, implying their potential roles in β-glucan-modulated immunity. These results are beneficial to understand the mechanism of β-glucan in resisting bacteria in fish.

Topics: Adaptive Immunity; Aeromonas salmonicida; Animal Feed; Animals; beta-Glucans; Diet; Dietary Supplements; Dose-Response Relationship, Drug; Fish Diseases; Gene Expression Profiling; Gram-Negative Bacterial Infections; Immunity, Innate; Oncorhynchus mykiss; Protective Agents; Random Allocation; Spleen; Transcriptome

Interleukin-6 (IL-6) is a pleiotropic cytokine with crucial immunoregulatory functions in both innate and adaptive immune responses. However, the IL-6 sequence and function remain unknown in sturgeon, one chondrostean fish. In the present study, we identified an interleukin-6 homolog from Siberian sturgeon (Acipenser baeri), named AbIL-6. Its open reading frame (ORF) was 657 nucleotides in length, encoding a polypeptide of 218 amino acids, which contains a signal peptide and the IL-6 family domain. Phylogenetic analysis showed that sturgeon IL-6 had close relationship with both teleost and chondrichthyes IL-6s. Abil-6 mRNA was highly expressed in spleen, brain and liver tissues of healthy sturgeon, and significantly up-regulated in the spleen, head kidney and liver by A.hydrophila (A.h) challenge. Heat-killed A.h and LPS effectively stimulated Abil-6 transcripts in primary spleen cells in vitro. In order to understand the bioactivity and influence of AbIL-6 on immune responses, recombinant AbIL-6 (rAbIL-6) was synthesized by prokaryotes and demonstrated to promote the proliferation of spleen cells and head kidney cells in vitro. Additionally, intraperitoneal injection of rAbIL-6 induced significantly higher expression of four immuno-related genes including il-1β, cxcl10, mhcIIβ and igm. rAbIL-6 improved the survival rate and reduced the tissue bacterial load after A.h infection. Taken together, these results suggest that AbIL-6 plays an important role in inflammatory responses and immune defense against bacterial infection of sturgeon.

Topics: Adaptive Immunity; Aeromonas hydrophila; Amino Acid Sequence; Animals; Base Sequence; beta-Glucans; Fish Diseases; Fish Proteins; Fishes; Gene Expression Profiling; Gene Expression Regulation; Gram-Negative Bacterial Infections; Interleukin-6; Lipopolysaccharides; Perciformes; Phylogeny; Poly I-C; Sequence Alignment

A trial was operated to assess the potential of using Lactobacillus plantarum L-137 (L-137) and/or β-glucan (BG) in improving the resistance of Nile tilapia against Aeromonas hydrophila. Control diet and 3 diets supplemented with L-137, BG or L-137 + BG were prepared. Final body weight, specific growth rate, superoxide dismutase, and catalase showed considerably (P < .05) increased values in L-137 or L-137/BG groups, while glutathione peroxidase increased significantly (P < .05) only in L-137/BG group. Fish fed L-137 and/or BG diets showed that feed conversion ratio and malonaldehyde levels were significantly decreased (P < .05). Also, both L-137 and BG helped Nile tilapia to have high phagocytosis activity and relative expression of tumor necrosis factor-alpha (TNF-α) and interleukin 1 beta (IL-1β) and interferon-gamma (INF-γ) genes. After A. hydrophila challenge, the intestinal villi epithelium of the L-137/BG group was intact and denser than the other groups. The hepatopancreas and spleen of the control group displayed severe necrosis in hepatocytes and congestion of blood sinusoids in addition to diffuse vacuolation. Regarding the L-137, BG and L-137/BG groups, there was a moderate and normal degree of vacuolation with focal necrosis and mild to moderate degree of congestion of blood sinusoids. Red blood cells, hemoglobin, and albumin showed meaningfully (P < .05) increased values in L-137 or L-137/BG groups. TNF-α, IL-1β, and INF-γ expressions were upregulated by L-137 and/or BG. The obtained results revealed the ability of L-137 and/or BG to protect Nile tilapia from the effects of A. hydrophila infection by the motivation of the immune, antioxidative, and antiinflammation responses.

Topics: Adjuvants, Immunologic; Aeromonas hydrophila; Animal Feed; Animals; Antioxidants; beta-Glucans; Diet; Disease Resistance; Fish Diseases; Fish Proteins; Gram-Negative Bacterial Infections; Lactobacillus plantarum; Probiotics

Dietary α-lipoic acid (LA), β-glucan (Gluc) and l-carnitine (L-Ca) are commonly used additives to promote fish growth and stress resistance in aquaculture production. However their mechanisms and efficiencies in helping fish to resist diseases have not been compared before. In this study, we fed Nile tilapia (Oreochromis niloticus) with diets containing appropriate doses of LA, Gluc and L-Ca for five weeks and further intraperitoneally injected the fish with Aeromonas hydrophila. After dietary treatment, none of the additives affected the fish growth, but dietary Gluc and L-Ca reduced protein and lipid body contents in fish, respectively. After A. hydrophila challenge, all fish treated with the three dietary additives showed higher survival rate, but those fed on dietary L-Ca had lower survival than those fed on LA and Gluc diets, indicating high protection efficiency of LA and Gluc. The protective mechanisms of the three feed additives were quite different under A. hydrophila infection. Dietary LA induced higher total antioxidant capacity and higher mRNA expression of anti-oxidative genes than other additives in liver and also activated partly the immune function in serum and spleen. Gluc largely increased the immune function by activating the immunity enzymes in serum, inducing inflammation in liver and increasing the expression of immune genes in spleen and head kidney. Gluc also increased partly the antioxidant capacity in serum and liver and lipid catabolism in liver. L-Ca largely increased lipid catabolism in liver while it increased partly the antioxidant capacities in serum and liver. Taken together, these results indicate that, dietary LA, Gluc and L-Ca have various protective mechanisms and differ in their efficiencies on resisting A. hydrophila infection in Nile tilapia.

Topics: Aeromonas hydrophila; Animal Feed; Animals; beta-Glucans; Carnitine; Cichlids; Diet; Dietary Supplements; Fish Diseases; Gram-Negative Bacterial Infections; Protective Agents; Thioctic Acid

β-glucan is one of the most potent immunostimulants enhancing innate immune activity, disease resistance and growth performance of many aquatic organisms. Nevertheless, there are few studies on feeding regimens of β-glucan that correlate to immune response and disease resistance and are important considerations for practical β-glucan utilization. Thus, the effect of β-glucan and feeding duration on innate immunity and disease resistance was investigated to establish an optimal feeding regimen of β-glucan for Nile tilapia (Oreochromis niloticus Linn.). A variety of β-glucan feeding regimens were evaluated, including: i) feeding for 2 weeks, ii) feeding for 4 weeks, and iii) feeding every-other-week, with the objective of establishing the optimal feeding regimen that enhanced innate immunity and disease resistance. Innate immunity parameters were determined every week for eight weeks. Alternative complement activity of all β-glucan groups was significantly (P < 0.05) increased at the end of the first week, and then fluctuated but was not significantly (P > 0.05) different to the control until the end of the trial. Increased lysozyme activity was only detected at the end of the second week in all β-glucan-treated groups, and then decreased to the control level during most of the sampling periods. Phagocytosis percentage was increased and prolonged by β-glucan feeding, while the phagocytic index was not. Apart from innate immunity, β-glucan-fed fish demonstrated enhanced disease resistance against Aeromonas hydrophila and Flavobacterium columnare challenge at only the end of the fourth week of the trial. The growth performance of β-glucan-fed fish was not significantly (P > 0.05) different among the experimental groups and control. Taken together, the result indicated that all β-glucan-feeding regimens resulted in quite similar outcomes with respect to innate immunity stimulation, disease resistance and growth performance. This novel result suggests that an every-other-week regimen is the optimal choice for Nile tilapia cultivation as an economic cost saving benefit. This is the first study to determine the optimal feeding-regimen of β-glucan to enhance innate immunity and increase resistance to infection by pathogenic bacteria in Nile tilapia.

Topics: Aeromonas hydrophila; Animal Feed; Animals; Aquaculture; beta-Glucans; Cichlids; Diet; Disease Resistance; Fish Diseases; Flavobacteriaceae Infections; Flavobacterium; Gram-Negative Bacterial Infections; Immunity, Innate

Interleukin-8, otherwise known as CXCL8, is a CXC chemokine that plays a pivotal regulatory role in immune and inflammation responses of animals. Here, we identified an interleukin-8 homologue from Siberian sturgeon (Acipenser baeri), named AbIL-8, which belongs to the lineage 1 group of teleost fish IL-8s. The cDNA of Abil-8 is 1130 bp in length, containing a 5'- untranslated region (UTR) of 50 bp, a 3'- UTR of 783 bp, and an open reading frame (ORF) of 297 bp that encodes a protein consisting of 98 amino acids. The deduced AbIL-8 contained five cysteines, four of which are highly conserved, and an ELR motif typical of known mammalian CXC chemokines was also found preceding the CXC motif. Our phylogenetic analysis showed that AbIL-8 clustered with the CXCL8_L1 sequences from other teleosts, being clearly distinct from those of either birds or mammals. Abil-8 mRNA was constitutively expressed in all tested tissues and significantly up-regulated in the liver and spleen tissues by the bacteria Aernomas hydrophila. The in vitro experiment using primary spleen cells stimulated with heat-killed Aernomas hydrophila or lipopolysaccharide (LPS) revealed a similar expression pattern to that found in vivo, whereas stimulation on spleen cells with β-glucan or polyI:C elicited negligible changes in levels of Abil-8 mRNA. Purified recombinant AbIL-8 not only exhibited chemotactic activity for lymphocytes and monocytes in peripheral blood leukocytes (PBLs) and, to a lesser extent, spleen cells, but also stimulated the proliferation of spleen cells at 10 ng/mLor above. Furthermore, intraperitoneal injection of rAbIL-8 also up-regulated the expression of immuno-related genes (IL-6, IgM and MHCIIβ) at 24 h. Collectively, these results enhance our understanding of how IL-8 functions in the regulation of the immune responses in sturgeon.

Topics: Aeromonas hydrophila; Amino Acid Sequence; Animals; Base Sequence; beta-Glucans; Fish Diseases; Fish Proteins; Fishes; Gene Expression Profiling; Gene Expression Regulation; Gram-Negative Bacterial Infections; Immunity, Innate; Interleukin-8; Lipopolysaccharides; Phylogeny; Poly I-C; Sequence Alignment

In this study, we show that β-glucan can modulate cortisol release in fish. We simulated a common situation in aquaculture: the transport of fish followed by contact with an opportunistic pathogen and observed what effect glucan had on the immune and stress response in these conditions. Pacu (Piaractus mesopotamicus) were fed with a diet containing β-glucan (0.1%) for 15 days prior to transport followed by an injection with heat-killed Aeromonas hydrophila. We sampled fish before transport, at arrival and at 3 and 24 h after bacterial injection. β-Glucans are used in aquaculture and have a known immunostimulatory effect, which was observed in this study. The results showed that β-glucan modulated the plasma cortisol levels differently by increasing these levels up to 24 h after transport and preventing the increase caused by bacterial inoculum injection. In addition, β-glucan enhanced the activity of the complement system at 24 h and reduced the monocytes and lymphocytes number in peripheral blood at 3 and 24 h after bacterial inoculation. Our results suggest that β-glucan modulated a bidirectional interaction between the stress and the immune responses. The modulation of cortisol levels and the immunostimulation by β-glucan at different moments in our study suggest the compound has a protective effect by avoiding higher levels of the hormone and improving resistance against bacterial infection in pacu. These results add evidence to support the use of β-glucan as an immunomodulator in the aquaculture industry.

Topics: Adjuvants, Immunologic; Aeromonas hydrophila; Animal Feed; Animals; beta-Glucans; Characiformes; Diet; Dietary Supplements; Fish Diseases; Gram-Negative Bacterial Infections; Hydrocortisone; Immunity, Innate; Random Allocation; Stress, Physiological

We tested the efficacy of a commercial product (Glucan-MOS

Topics: Aeromonas hydrophila; Animal Feed; Animals; beta-Glucans; Characiformes; Diet; Dietary Supplements; Dose-Response Relationship, Drug; Fish Diseases; Gram-Negative Bacterial Infections; Mannans; Random Allocation; Stress, Physiological

β-Glucans are naturally occurring polysaccharides that are produced by bacteria, fungi and yeast. They are considered immunostimulants in fish acting on non-specific defense mechanism. Yeast-derived glucans from cell wall (Sterigmatomyces halophilus, β-Gluc/Sh) have been used for this purpose in this study. Therefore, an in vitro assay using peripheral blood leucocytes (PBLs) from Pacific red snapper was performed to evaluate the stimulant effects of β-Gluc/Sh and zymosan A (positive control) for 12 and 24 h and after bacterial challenge with Aeromonas hydrophila at 24 h. In addition, structural characterization of this marine yeast glucan was performed by proton nuclear magnetic resonance (NMR) revealing structures containing (1-6)-branched (1-3)-β-D-glucan. PBLs responded positively to β-Gluc/Sh where cell viability was higher than 80%. After challenge, β-Gluc/Sh was able to inhibit cytotoxicity caused by A. hydrophila, highlighting that the PBLs incubated with β-Gluc/Sh significantly increased the non-specific immune response, such as phagocytic activity, respiratory burst, nitric oxide and peroxidase activities followed by an increase in superoxide dismutase and catalase activities after 12 and 24 h post-stimulation and after challenge with the pathogen. Regarding induction of antioxidant gene expression, it was more pronounced in stimulated β-Gluc/Sh leucocytes compared to other groups at all experimental times of the trial and after bacterial challenge. Indeed, our results clearly showed the ability of leucocytes to strongly react to β-Gluc/Sh with an increase in cytokine gene expression, particularly the IL-1β, IL-10 and IL-17 genes. These results confirm that S. halophilus yeast-derived β-glucan, isolated from an extreme marine environment, is beneficial for increasing innate immune response and enhancing resistance against A. hydrophila in vitro.

Topics: Aeromonas hydrophila; Animal Feed; Animals; Basidiomycota; beta-Glucans; Diet; Dietary Supplements; Disease Resistance; Fish Diseases; Gram-Negative Bacterial Infections; Immunity, Innate; In Vitro Techniques; Leukocytes; Perciformes

This study investigated the effects of two β-glucan molecules with different purities and isolated by different biotechnological processes on the immune response of matrinxã (Brycon amazonicus) prior and after challenge with Aeromonas hydrophila. In this sense, we evaluated serum cortisol and plasma glucose levels, the number of leukocytes (lymphocytes, neutrophils and monocytes), as well as the respiratory activity of leukocytes prior to, 6 and 24 h post infection (hpi). During 15 days, fish were fed with diets containing 0.1% of two β-glucans (β-G 1 and β-G 2, with 71 and 62% of purity, respectively) and then submitted to challenge. Results were compared with a positive control group fed with a β-glucan-free diet. A negative control group, also fed with β-glucan-free diet but inoculated with PBS, was established to evaluate the effect of handling during injection. Our results showed that different β-glucans affected differently the biological responses of matrinxã. The βG 2 modulated the cortisol profile prior to and after the acute infection with A. hydrophila, and increased the mobilization and activity of leukocytes. The infection promoted lymphopenia at 6 hpi and both β-glucans increased the circulating lymphocyte population 24 hpi. Moreover, the β-G 2 prevented the infection-induced neutrophilia at 6 and 24 hpi. Finally, the β-G 2 caused a marked increase in the circulating monocytes prior to infection, and a reduction at 6 hpi that was reversed at 24 hpi. In summary, our study demonstrates that β-G 2 was more efficient on the induction of the cell-mediate immunity in matrinxã.

Topics: Aeromonas hydrophila; Animals; beta-Glucans; Characiformes; Fish Diseases; Gram-Negative Bacterial Infections; Hydrocortisone; Immunity, Innate; Leukocytes

Four diets supplemented with three different levels of β-1,3-glucan (0.5, 1, and 2 g kg

Topics: Aeromonas hydrophila; Animals; beta-Glucans; Cypriniformes; Diet; Dietary Supplements; Fish Diseases; Gram-Negative Bacterial Infections; Intestines; Malondialdehyde; Peroxidases; Superoxide Dismutase

This study attempts to describe the immunostimulatory effects of three fungal glucans on innate immunity responses in an in vitro assays using Pacific red snapper leukocytes. First, the yield glucans obtained was higher in Aspergillus niger, follow by Aspergillus ochraceus and Alternaria botrytis (40, 20 and 10%, respectively). Structural characterization of these fungal glucans by proton nuclear magnetic resonance (NMR) indicated structures containing (1-6)-branched (1-3)-β-D-glucan. The immunostimulatory activity of fungal glucans were assessed in head-kidney leukocytes at 24 h using colorimetric assays and molecular gene expression. In addition, the response against bacterial infection using Aeromonas hydrophila was evaluated by flow cytometry with annexin V/propidium iodide. Leukocytes responded positively to fungal glucans where the viability was higher than 80%. Interestingly, A. niger β-glucans enhanced the phagocytic ability and capacity in head-kidney leukocytes. Immunological assays reveled an increased in nitric oxide production, myeloperoxidase, superoxide dismutase and catalase activities, in fish stimulated with A. niger β-glucans. Induction of cytokines (IL-1β, TNF-α, IL-6, IL-8 and IL-12) were more pronounced in A. niger β-glucans leukocytes stimulated compared to other group. Finally, flow cytometry assay showed that A. botrytis and A. niger β-glucans were able to inhibit apoptosis caused by Aeromonas hydrophila in the Pacific red snapper leukocytes indicating an immunostimulant potent response by fungi derived-glucans. These results strongly support the idea that fungal β-glucans can stimulate the immune mechanism in head-kidney leukocytes and that Aspergillus niger β-glucan possess immunostimulatory properties cell increasing viability, and reducing necrotic cell death caused by Aeromonas hydrophila.

Topics: Aeromonas hydrophila; Alternaria; Animals; Apoptosis; Aspergillus niger; Aspergillus ochraceus; beta-Glucans; Cells, Cultured; Cytokines; Fish Diseases; Gram-Negative Bacterial Infections; Head Kidney; Immunity, Innate; Immunization; Inflammation Mediators; Leukocytes; Lymphocyte Activation; Mycoses; Nitric Oxide; Perciformes

This study investigated the role of endogenous cortisol on the innate immune response in matrinxã (Brycon amazonicus) fed with β-glucan, prior to and after stressor exposure and bacterial challenge. For this, we evaluated the serum cortisol and plasma glucose levels, the serum lysozyme levels, the hemolytic activity of the complement system, and the respiratory activity of leukocytes, as well as the number of circulating erythrocytes and leukocytes of fish fed during 15 days with diets containing β-glucan 0.1% (β-G) or β-glucan 0.1% + metyrapone 30 mg kg

Topics: Aeromonas hydrophila; Animal Feed; Animals; beta-Glucans; Characidae; Diet; Dietary Supplements; Fish Diseases; Gram-Negative Bacterial Infections; Hydrocortisone; Immunity, Innate; Random Allocation

Topics: Adjuvants, Immunologic; Aeromonas hydrophila; Animal Feed; Animals; beta-Glucans; Diet; Dietary Supplements; Fish Diseases; Fish Proteins; Gene Expression Regulation; Gram-Negative Bacterial Infections; Immunity, Innate; Oncorhynchus mykiss; Real-Time Polymerase Chain Reaction; Tissue Distribution

IL17s are pro-inflammatory cytokines that play important roles in host fighting against extracellular bacteria and auto-immune and allergic diseases. IL17D is believed to be the most ancient IL17 member and its functions are far from clarity. Although it has been found in invertebrates, jawless fish, teleosts, and tetrapods, it has not been described in chondrostean fish. Moreover, there are discrepancies concerning its expression pattern in these animals. In this study, we cloned and characterized the cDNA of il17d in Siberia sturgeon (Acipenser baerii), a chondrostean fish and commercially important species in aquaculture. The sturgeon il17d cDNA encodes a deduced protein of 210aa. The classical characteristics of IL17, such as IL17 domain, cysteine and serine residues importantly for cystine-knot formation, and signal peptide, were observed in sturgeon IL17D. Phylogenetic analysis and multiple alignment suggest it is a counterpart of mammalian IL17D. However, in vivo studies demonstrated that the expression pattern of sturgeon il17d mRNA is different from that of other teleosts and jawless fish, and in most cases its expression was down-regulated at the early time points and gradually increasing at late time points when sturgeon were challenged with bacteria (Aernomas hydrophila or Staphylococcus aureus). The In vitro study by using primary spleen cells stimulated with polyI:C revealed a similar expression pattern to that in vivo studies, while the stimulation with β-glucan or LPS, which normally induced expression of il17d mRNA in target cells in vitro in other animals, did not show apparent changes in the expression of il17d mRNA. The results of present study indicated sturgeon IL17D may possess some different characteristics from its counterparts of other fish and invertebrates in the immune response, and may contribute to the understanding of IL17D functions in evolution as well as the potential use in sturgeon aquaculture.

Topics: Aeromonas hydrophila; Amino Acid Sequence; Animals; Base Sequence; beta-Glucans; DNA, Complementary; Fish Diseases; Fish Proteins; Fishes; Gene Expression Regulation; Gram-Negative Bacterial Infections; Interleukin-27; Lipopolysaccharides; Phylogeny; Poly I-C; Random Allocation; RNA, Messenger; Sequence Alignment; Staphylococcal Infections; Staphylococcus aureus

The present study evaluated the effects of dietary β-glucan (0, 0.05%, 0.1%, and 0.2%) on growth performance after 42 days of feeding. Thereafter, rainbow trout (Oncorhynchus mykiss) were infected with Aeromonas salmonicida, and survival rates as well as the regulating processes of stress- and immune-related factors were analyzed. In general, higher dietary β-glucan levels obviously improved specific growth rate (SGR), weight gain (WG) and feed efficiency (FE) (P ≤ 0.05). Survival rates in β-glucan groups increased significantly compared with the control group after A. salmonicida infection (P ≤ 0.05). Serum total superoxide dimutase (T-SOD), peroxidase (POD) as well as catalase (CAT) activities, and their mRNA expressions in the head kidney of fish in the β-glucan groups generally increased to higher levels after infection, and more quickly, compared with in the control group. Serum lysozyme (LSZ) and its expression in the head kidney in β-glucan groups reached a higher peak earlier than in the control group. Serum glutamic oxalacetic transaminase (GOT) and glutamic pyruvic transaminase (GPT) levels in the β-glucan groups were significantly lower than in the control group (P ≤ 0.05). The peak of heat shock protein 70 (HSP70) expression in the 0.2% β-glucan group was higher and occurred earlier than in other groups (P ≤ 0.05). These results confirm that 0.1% and 0.2% dietary β-glucan are beneficial for promoting growth in rainbow trout and enhancing resistance against A. salmonicida. Furthermore, β-glucan could play an important role in regulating stress- and immune-related factors in rainbow trout to more quickly fight against bacterial infection.

Topics: Adjuvants, Immunologic; Aeromonas salmonicida; Animal Feed; Animals; beta-Glucans; Diet; Dietary Supplements; Disease Resistance; Dose-Response Relationship, Drug; Fish Diseases; Fish Proteins; Gram-Negative Bacterial Infections; Head Kidney; Immunity, Innate; Oncorhynchus mykiss; Random Allocation

Topics: Administration, Oral; Aeromonas hydrophila; Animals; Anti-Infective Agents; beta-Glucans; Carps; Claudins; Gene Expression Regulation; Gram-Negative Bacterial Infections; Inflammation; Interleukin-1beta; Intestines; Peptide Fragments; RNA, Ribosomal, 16S

The formation of neutrophil extracellular traps (NETs) has been characterised as a novel antimicrobial host defence strategy of neutrophils besides phagocytosis and degranulation, which may lead to entrapment and subsequent immobilisation and/or killing of bacterial pathogens. Here we studied the effect of the feed additive β-glucan, namely MacroGard(®), on the formation and functionality of NETs in carp. Therefore, common carp (Cyprinus carpio) head kidney and kidney cells were isolated and treated with or without β-glucan over time. The formation of NETs was analysed by immunofluorescence microscopy and revealed a distinct increase of NET-formation with β-glucan. Furthermore the subsequent entrapment of Aeromonas hydrophila, an important fish pathogen, was increased after stimulating the cells with β-glucan. However, β-glucan did not lead to a stimulation of antimicrobial activity of neutrophils against A. hydrophila. In conclusion, the data underline the fact that the feed additive β-glucan is able to modulate carp neutrophil functions.

Topics: Aeromonas hydrophila; Animals; beta-Glucans; Carps; Dietary Supplements; Extracellular Space; Gene Expression Profiling; Gram-Negative Bacterial Infections; Head Kidney; Immunity, Innate; Neutrophils

This study attempts to describe the effects of different administration periods of dietary β-glucan and Vit C on the non-specific immune response, physiological parameters and disease resistance of Nile tilapia against Aeromonas hydrophila infection. Therefore, a feeding trial (288 fish) was conducted to determine the best administration period (7, 15, 30 and 45 days) for a Nile tilapia diet supplemented with 0.1% β-glucan and 600 mg Vit C/kg diet. After the administration period, three different groups of 96 fish were exposed to one of the following three stresses: cold-induced stress, transport-induced stress, and A. hydrophila challenge. Hematological, biochemical and immunological responses were analyzed before and/or after stress. Cold-induced stress increased cortisol levels and reduced the leukocyte count in fish fed the test diet for seven days compared with the other periods. After transport-induced stress, fish fed the test diet for seven days required more hours to return to the baseline levels of cortisol and neutrophils. Moreover, independently of the administration period, fish needed 24 h for leukocyte and glucose levels to return to the initial values. The lowest survival after bacterial infection was observed in fish test diet for seven days. Based on fish hematological and biochemical responses, diet supplemented with 0.1% of β-glucan and 600 mg of Vit C/kg fed for at least 15 days is recommended for Nile tilapia especially when fish are likely to encounter transport-induced stress, and this stress was more severe than cold-induced stress or bacterial challenge.

Topics: Aeromonas hydrophila; Analysis of Variance; Animal Husbandry; Animals; Aquaculture; Ascorbic Acid; beta-Glucans; Brazil; Cichlids; Cold Temperature; Dietary Supplements; Disease Resistance; Fish Diseases; Gram-Negative Bacterial Infections; Stress, Physiological

Effects of β-glucan on innate immune responses and survival were studied in pacu experimentally infected with Aeromonas hydrophila. Fish fed diets containing 0, 0.1% and 1% β-glucan were injected with A. hydrophila. β-glucan enhanced fish survival in both treated groups (26.7% and 21.2% of the control, respectively). Leukocyte respiratory burst and alternative complement pathway activities were elevated after bacterial challenge regardless the β-glucan concentration. Lysozyme activity was higher after infection and showed a gradual increase as β-glucan concentration increased. A significant elevation in WBC count was observed either after bacterial challenge or by influence of β-glucan separately. The same response was observed in the number of thrombocytes, lymphocytes, eosinophils, LG-PAS positive cell and monocytes. It can be concluded that feeding pacu with β-glucan can increase protection against A. hydrophila, due to changes in non-specific immune responses.

Topics: Aeromonas hydrophila; Animal Feed; Animals; beta-Glucans; Disease Resistance; Fish Diseases; Gram-Negative Bacterial Infections; Survival Analysis

The effect of β-glucans as feed additive on the profile of C-reactive protein (CRP) and complement acute phase responses was studied in common carp Cyprinus carpio after exposition to a bacterial infection with Aeromonas salmonicida. Carp were orally administered with β-glucan (MacroGard®) for 14 days with a daily β-glucan intake of 6 mg per kg body weight. Fish were then intraperitoneally injected with either PBS or 1 × 10⁸ bacteria per fish and sampled at time 0, 6, 12, 24, 48, 72, 96 and 120 h post-injection (p.i.) for serum and head kidney, liver and mid-gut tissues. CRP levels and complement activity were determined in the serum samples whilst the gene expression profiles of CRP and complement related genes (crp1, crp2, c1r/s, bf/c2, c3 and masp2) were analysed in the tissues by quantitative PCR. Results obtained showed that oral administration of β-glucan for 14 days significantly increased serum CRP levels up to 2 fold and serum alternative complement activity (ACP) up to 35 fold. The bacterial infection on its own (i.e. not combined with a β-glucan feeding) did have significant effects on complement response whilst CRP was not detectably induced during the carp acute phase reaction. However, the combination of the infection and the β-glucan feeding did show significant effects on both CRP and complement profiles with higher serum CRP levels and serum ACP activity in the β-glucan fed fish than in the control fed fish. In addition, a distinct organ and time dependent expression profile pattern was detected for all the selected genes: a peak of gene expression first occurred in the head kidney tissue (6 h p.i. or 12 h p.i.), then an up-regulation in the liver several hours later (24 h p.i.) and finally up- or down-regulations in the mid-gut at 24 h p.i. and 72 h p.i. In conclusion, the results of this study suggest that MacroGard® stimulated CRP and complement responses to A. salmonicida infection in common carp.

Topics: Acute-Phase Reaction; Aeromonas salmonicida; Animals; beta-Glucans; C-Reactive Protein; Carps; Dietary Supplements; Fish Diseases; Gene Expression Profiling; Gene Expression Regulation; Gram-Negative Bacterial Infections; Immunity, Innate; Injections, Intraperitoneal; Real-Time Polymerase Chain Reaction

The objective of the present study was to determine the action of β-glucans as feed additives on the gene expression profile of some inflammatory-related cytokines from common carp (Cyprinus carpio L.) during the early stages of a non-lethal bacterial infection with Aeromonas salmonicida. β-glucan (MacroGard(®)), was administered daily to carp (6 mg per kg body weight) in the form of supplemented commercial food pellets for 14 days prior to infection. Control and treated fish were then intraperitoneally injected with PBS or 4×10(8) bacteria per fish and were sampled at time 0 and 6h, 12h, 1 day, 3 days and 5 days post-injection. Head kidney and gut were collected and the gene expression patterns for tnfα1, tnfα2, il1β, il6 and il10 were analyzed by quantitative PCR. Results obtained showed that treatment with β-glucans generally down-regulated the expression of all measured genes when compared to their corresponding controls. After injection, highest changes in the gene expression levels were obtained at 6h; particularly, in head kidney there was higher up-regulation of tnfa1 and tnfa2 in infected fish fed β-glucans in comparison to control feed; however, in gut there was a significant down-regulation of tnfα1, tnfα2, il1β and il6 in infected fish fed β-glucans. Analysis of carp specific antibodies against A. salmonicida 30 days after injection revealed their levels were reduced in the infected β-glucan group. In conclusion, a diet supplemented with β-glucan (MacroGard(®)) reduced the gene expression levels of some inflammation-related cytokines in common carp. Such a response appears to be dependent of organ studied and therefore the immunostimulant may be preventing an acute and potential dangerous response in gut, whilst enhancing the inflammatory response in head kidney when exposed to A. salmonicida.

Topics: Adjuvants, Immunologic; Aeromonas salmonicida; Animals; Antibodies, Bacterial; beta-Glucans; Carps; Dietary Supplements; Fish Diseases; Gene Expression Profiling; Gene Expression Regulation; Gram-Negative Bacterial Infections; Head Kidney; Inflammation; Intestines; Time Factors

A novel host innate immune defence mechanism against invading pathogens, namely the formation of neutrophil extracellular traps (NETs), has recently been discovered. These NETs are described as DNA fibres released by dying neutrophils, which are able to entrap and kill various microbes. Here we studied the effect of the feed additive β-glucan, namely MacroGard(®), on the degradation of NETs by the important fish pathogen Aeromonas hydrophila. Therefore, common carp (Cyprinus carpio) head kidney cells consisting of approximately 45% neutrophils were isolated and treated with or without β-glucan. The degradation of NETs after co-incubation with A. hydrophila was analysed by immunofluorescence microscopy. The data show that A. hydrophila is able to degrade NETs and that treatment of cells with β-glucan significantly protects the NETs against bacterial degradation. Control experiments revealed that β-glucan augments nuclease activity of the bacteria at the same time while protecting the NETs against its degradation. In conclusion the data indicate that β-glucan might affect the composition and stabilisation of NETs and thereby protecting them against degradation by A. hydrophila nuclease.

Topics: Aeromonas salmonicida; Animals; beta-Glucans; Carps; Cell Death; Dietary Supplements; Extracellular Space; Fish Diseases; Gene Expression Profiling; Gene Expression Regulation; Gram-Negative Bacterial Infections; Head Kidney; Microscopy, Fluorescence; Neutrophils

The association between β-glucan (MacroGard®) supplemented feed and apoptosis in immune-related organs of common carp (Cyprinus carpio) was studied using fluorescence microscopy and real-time PCR. In addition the effect of Aeromonas salmonicida, LPS and Poly(I:C) injections on this relationship was evaluated. Whilst acridine orange staining revealed that apoptosis levels were independent of MacroGard® and LPS/Poly(I:C) administration or their combination, it was shown that injection with A. salmonicida increased the percentage of apoptotic cells irrespective of the feeding regime. It was apparent that in all the treatments gene expression profiles displayed organ and time dependency. For example no effect was observed at 7 days of MacroGard® administration while 25 days of feeding led to increased iNOS expression and differential up-regulation of anti- or pro-apoptotic genes depending on organ. This may indicate differences in NO sensitivity. MacroGard® also led to an elevation of pro- as well as anti-apoptotic genes in LPS or Poly(I:C) injected fish, while LPS/Poly(I:C) alone had little effect. A. salmonicida caused enhanced iNOS expression and it is possible that the type of apoptosis pathway induced is organ dependent as Caspase 9 is induced in mid-gut but not in pronephros. These results indicate that MacroGard® feeding alone or in combination with other pathogenic factors did not induce significant apoptosis in immune organs.

Topics: Aeromonas salmonicida; Animals; Apoptosis; beta-Glucans; Carps; Dietary Supplements; Fish Diseases; Gene Expression Profiling; Gene Expression Regulation; Gram-Negative Bacterial Infections; Immunity, Innate; Injections, Intraperitoneal; Lipopolysaccharides; Poly I-C; Real-Time Polymerase Chain Reaction; Reverse Transcriptase Polymerase Chain Reaction

The present study was conducted to investigate the effect of beta-glucan (derived from Saccharomyces cerevisiae) on the immune response and its protection against an infection of the bacterial pathogen Aeromonas hydrophila in zebrafish (Danio rerio). Zebrafish received beta-glucan by intraperitoneal injection at three different concentrations (5, 2 and 0.5 mgml(-1)) at 6, 4 and 2 days prior the challenge. On challenge day the control and beta-glucan pretreated zebrafish were intraperitoneally injected with A. hydrophila and mortality was recorded for 4 days. Intraperitoneal injection of 5 mgml(-1) of beta-glucan significantly reduced the mortality. A single injection of 5 mgml(-1) of beta-glucan 6 days before challenge also enhanced significantly the survival against the infection. The treatment with beta-glucan increased the myelomonocytic cell population in the kidney at 6h postchallenge with A. hydrophila. Moreover it enhanced the ability of kidney cells to kill A. hydrophila. beta-glucan did not affect the expression of TNFalpha or IL-1 beta but seemed to modulate IFNgamma and chemokine expression in kidney.

Topics: Adjuvants, Immunologic; Aeromonas hydrophila; Animals; beta-Glucans; Cytokines; Fish Diseases; Gene Expression Regulation; Gram-Negative Bacterial Infections; Immunity, Innate; Kidney; Survival Analysis; Zebrafish

GATA-3 is a T cell-specific transcription factor and is essential for the development of the T cell lineage and differentiation of T helper type 2 cells. We have identified and characterized the full-length Atlantic salmon GATA-3 cDNA (3074bp), having two zinc finger domains which are fully conserved within teleosts and higher vertebrates. RT-PCR analysis revealed that the Atlantic salmon GATA-3 (AsGATA-3) is strongly expressed in gills, thymus, and brain. Moreover, the involvement of GATA-3 in Atlantic salmon immune response was demonstrated by investigating the early time dependent expression profile of GATA-3 in spleen and head kidney following intraperitoneal injection of live Aeromonas salmonicida, LPS, and beta-glucan. Furthermore, we have determined 1.9kb of upstream promoter sequence and found a number of sequence motifs which match those of known transcription factor binding sites and the AsGATA-3 promoter is a TATA-less promoter. Activities of presumptive regulatory regions of this gene were assessed by transfecting different 5' deletion constructs and the result showed the basal promoter and positive transcriptional regulator activity of AsGATA-3 gene is determined by sequences located between +58 and -199bp upstream of the transcriptional start site (TSS). This study provides further insights into the transcriptional regulation of AsGATA-3.

Topics: Aeromonas salmonicida; Amino Acid Sequence; Animals; Base Sequence; beta-Glucans; Cloning, Molecular; GATA3 Transcription Factor; Genes, Reporter; Gram-Negative Bacterial Infections; HeLa Cells; Humans; Lipopolysaccharides; Molecular Sequence Data; Phylogeny; Promoter Regions, Genetic; Salmo salar; Sequence Alignment; Transfection

The present experiment was carried out to study the effect of different dosages of beta-glucan suspension derived from barley on the innate immune response and disease resistance of Anabas testudineus spawns against infection caused by Aeromonas hydrophila. Four different dosages of beta-glucan suspension in phosphate buffered saline at the rate of 0, 5, 10, 15 mg l(-1) were taken and 8 days old spawn were exposed for 2 h and 3 h. The cell suspension of spawn was assayed for total protein, acid phosphatase activity, lysozyme activity, bactericidal and NBT. Further, the spawns were challenged with 3 x 10(5) cells ml(-1) of A. hydrophila and survivability percentage and immunological parameters were assayed upto day 7. On day 7, most of the immunological parameters such as lysozyme activity, bactericidal activity and NBT activity were significantly enhanced after exposing the fish to all the concentrations of beta-glucan. Challenge study indicated least mortality in the group of spawns immersed in 15 mg l(-1) beta-glucan suspension for 3 h. Thus, 3 h exposure to beta-glucan suspension could reduce the mortality and increase the immunity of A. testudineus spawns.

Topics: Acid Phosphatase; Aeromonas hydrophila; Animals; beta-Glucans; Fish Diseases; Gram-Negative Bacterial Infections; Immunity, Innate; Muramidase; Perches; Superoxides; Survival Analysis

The purpose of this study was to determine if multiple injections of different dosages of beta-glucan derived from barley would enhance the immune response and disease resistance against infections due to opportunistic pathogens Aeromonas hydrophila and Edwardsiella tarda in Labeo rohita fingerlings. Hence, four different dosages of beta-glucan suspension in phosphate-buffered saline at the rate of 0, 5, 10, 15 mg kg(-1) body weight of fish were injected intraperitoneally to the fingerlings of Labeo rohita at two-week intervals for four times. After every two-week interval different serum biochemical, haematological and immunological parameters of fish were evaluated. At the end of immunostimulation trial of 56 days, fish were divided into four subgroups under each major treatment group for challenge through i.p injection and bath immersion with two pathogens, A. hydrophila and E. tarda. The mortality (%) and agglutinating antibody titre was recoded on 28th day post challenge. Most of the immune parameters such as leucocyte count, phagocytic ratio, phagocytic index, lysozyme activity, complement activity, serum bactericidal activity were significantly (P < 0.05) enhanced on 42 days after three i.p injections of 10 mg of beta-glucan kg(-1) body wt. Challenge study indicated least mortality in the group of fishes injected with medium dose of 10 mg of beta-glucan kg(-1) body wt. four times. Multiple injections of beta-glucan might have maintained the activation of phagocytic cells for a long period which in turn would lead to long-term protection in fishes. Thus, injections of 10 mg of beta-glucan kg(-1) body wt. for three times can be advocated to enhance the immune response of fish species under aquaculture.

Topics: Aeromonas hydrophila; Animals; beta-Glucans; Blood Chemical Analysis; Cyprinidae; Dose-Response Relationship, Drug; Edwardsiella tarda; Enterobacteriaceae Infections; Fish Diseases; Gram-Negative Bacterial Infections; Immunity, Innate; Injections, Intraperitoneal; Leukocytes; Linear Models; Muramidase; Phagocytosis; Superoxides

This study investigated the effects of short and prolonged administration of a yeast beta-glucan on non-specific immune parameters, growth rate and the disease resistance of Asian catfish, Clarias batrachus. Fish fed with a basal diet (control) and test diet (basal diet supplemented with 0.1% glucan) for 1, 2 and 3 weeks were assayed for superoxide production, serum myeloperoxidase (MPO) content, natural haemagglutinin level, complement and lysozyme activities. Fish were weighed at weekly intervals and specific growth rate (SGR, % increase in body weight per day) was determined. After each week, fish were challenged with Aeromonas hydrophila to measure the level of protection. Results showed that glucan administration at 0.1% in feed, significantly (P<0.05) enhanced MPO and lysozyme levels, superoxide production, haemagglutination titre and level of protection against A. hydrophila challenge, irrespective of length of exposure. The alternative complement activity and SGR were not affected by the dietary supplementation of yeast glucan. As glucan feeding at 0.1% for 1 week is able to enhance the non-specific immunity and disease resistance of catfish efficiently, short-term feeding might be used in farmed catfish diets to enhance disease resistance.

Topics: Adjuvants, Immunologic; Aeromonas hydrophila; Animal Feed; Animals; beta-Glucans; Catfishes; Complement System Proteins; Dietary Supplements; Fish Diseases; Gram-Negative Bacterial Infections; Hemagglutination; Immunity, Innate; Muramidase; Nitroblue Tetrazolium; Peroxidase; Random Allocation; Respiratory Burst; Time Factors

Previous studies have demonstrated that bacterial lipopolysaccharide (LPS) and heat killed Staphylococcus aureus (SA) activation of inflammatory cells depended in part upon activation of heterotrimeric Gi proteins. It has also been shown that (1 --> 3) beta-D-glucan can suppress inflammatory cell activation by microbial products although the cellular mechanism of the glucan effect remains to be clearly defined. We hypothesized that Gi proteins function as a common convergent signaling pathway for both LPS and SA leading to monocyte mediator production. Additionally, we hypothesized that soluble glucan suppresses LPS and SA induced cytokine production via Gi protein coupled signaling. Human THP-1 promonocytic cells were pretreated with pertussis toxin (PTx, 100 ng/ml or 1 microgram/ml) 6 hours prior to stimulation with LPS (10 microgram/ml) and SA (10 microgram/ml) and/or soluble glucan (10 microgram/ml). Both LPS and SA significantly (p < 0.05) induced cytokine production IL-6 > TNF alpha > IL-1 beta > GM-CSF > IL-10 > IFN gamma. The induction of these cytokines was significantly (p < 0.05) suppressed by PTx. Glucan treatment alone had no effect on cytokine production but suppressed (P < 0.05) LPS and SA induced cytokines. PTx further augmented (p > 0.05) the inhibitory effect of glucan on the LPS and SA induced cytokine expression. The data support the hypothesis that Gi proteins function as a common signaling protein for both LPS and SA induction of pro-and anti-inflammatory cytokines and that soluble glucan effectively suppresses cytokine production to the microbial stimuli. In contrast, the effect of soluble glucan on inhibiting cellular activation by LPS and SA is Gi protein independent.

Topics: beta-Glucans; Cell Culture Techniques; Cytokines; Gram-Negative Bacterial Infections; Gram-Positive Bacterial Infections; GTP-Binding Proteins; Humans; Inflammation; Lipopolysaccharides; Monocytes; Proteoglycans; Shock, Septic; Signal Transduction; Staphylococcal Infections; Staphylococcus aureus

In order to determine the efficacy and immunoreversal effect of the 4 dietary immunomodulators, viz. lactoferrin, beta-1,3 glucan, levamisole and vitamin C, on disease resistance of a commercially important catfish, Clarias batrachus, fish were fed diets supplemented with various levels of these substances in 2 subgroups, healthy and immunocompromised, during a 30 d trial. An artificial immunosuppressive state was induced by giving 3 intraperitoneal (i.p.) injections of cyclophosphamide (CYP) at a dose level of 200 mg kg(-1) body weight at 1 wk intervals in the immunocompromised vaccinated subgroup and 3 consecutive injections 3 d before challenge in the immunocompromised non-vaccinated subgroup. On the first day of the experiment, the fish were vaccinated against a formalin-killed Aeromonas hydrophila bacterin. After 30 d, antibody titre (as measured through bacterial agglutination titre) and disease resistance against A. hydrophila were determined. The results demonstrate that all 4 immunomodulators were capable of significantly (p < 0.05) enhancing the specific immune response; this was evident through raised antibody titre and protection against A. hydrophila in both healthy and immunocompromised vaccinated subgroups compared to their respective controls. Similarly, all 4 substances significantly raised the survival rates in immunocompromised and healthy non-vaccinated fish. Thus, these substances were capable of reducing the immunosuppression induced by CYP injections in both vaccinated and non-vaccinated fish compared to their respective controls. Among the 4 substances studied, beta-1,3 glucan was found to be the most effective immunomodulator, followed by levamisole, lactoferrin and vitamin C in Asian catfish. Therefore, the results support the introduction of these substances into the diet of fish grown in farms under immunosuppressive/stressful conditions in order to enhance protection against infection and offer economic benefits.

Topics: Adjuvants, Immunologic; Aeromonas hydrophila; Animals; Antibodies; Ascorbic Acid; Bacterial Vaccines; beta-Glucans; Catfishes; Cyclophosphamide; Dietary Supplements; Fish Diseases; Gram-Negative Bacterial Infections; Immunity, Innate; Immunocompromised Host; Lactoferrin; Levamisole; Survival Analysis; Time Factors

Combined effects of beta-glucan and lipopolysaccharide (LPS) on survival and immune response were studied in Cyprinus carpio that were challenged with the pathogen Aeromonas hydrophila. beta-Glucan from Saccharomyces cervisiae and LPS from a virulent strain of A. hydrophila were used in this study. Different concentrations of beta-glucan+LPS mixture were administered on days 1, 7, and 14 through different routes (intraperitoneal injection, bathing, and oral administration). Control and test fish were challenged by intraperitoneal injection of LD50 concentration of A. hydrophila on day 16 and subsequently, mortality and relative percent survival (RPS) were recorded. Intraperitoneal injection elicited 100% RPS even at the lowest concentration (100 microg beta-glucan+10 microg LPS); whereas, oral administration improved RPS rate of carps at higher concentration (1% beta-glucan+0.25% LPS). Bathing did not improve the RPS. Test animals injected with even the minimum dose of the immunomodulators (100 microg beta-glucan+10 microg LPS/fish) had a significant increase in total blood leukocyte counts and an increase in the proportion of neutrophils and monocytes. Superoxide anion production by macrophages was also elevated, which presumably aided the efficient killing of bacterial pathogen. Lower concentration of beta-glucan+LPS had an adjuvant effect on antibody production as pretreatment by injection of 100 microg beta-glucan+10 microg LPS/fish resulted in higher antibody titer against A. hydrophila following vaccination. RT-PCR analyses showed that the expression of interleukin-1beta mRNA did not increase in test fish when compared with the control. Classical and alternative complement pathways were not affected by either the dose or the route of administration of the compounds. It may be concluded that intraperitoneal injection and oral administration, and not the bathing, of beta-glucan+LPS mixture in carp could enhance resistance to challenge by A. hydrophila through changes in several non-specific and specific immune responses.

Topics: Adjuvants, Immunologic; Aeromonas hydrophila; Animals; Antibodies, Bacterial; Aquaculture; beta-Glucans; Carps; Complement Pathway, Alternative; Complement Pathway, Classical; Fish Diseases; Gram-Negative Bacterial Infections; Interleukin-1beta; Leukocyte Count; Lipopolysaccharides; Reverse Transcriptase Polymerase Chain Reaction; Superoxides

Effects of beta-glucan administration on survival and immune modulations were studied in Cyprinus carpio against the bacterial pathogen, Aeromonas hydrophila. Beta-glucan was extracted from Saccharomyces cervisiae and purified. A virulent strain of the pathogen A. hydrophila was collected from infected fish. Different concentrations of beta-glucan were administered to test animals on day 1, 3 and 5 through different routes (intraperitoneal injection (ip), bathing and oral administration). Control and test animals were challenged by ip injection of LD50 concentration of A. hydrophila on day 7 and mortality was observed and Relative Percent Survival (RPS) was calculated. Intraperitoneal injection of 500 microg of glucan significantly enhanced the RPS; bathing and oral administration of glucan did not influence the RPS. On day 7, test animals injected with 100, 500 and 1000 microg of glucan had a significant increase in total blood leucocyte counts and an increase in the proportion of neutrophils and monocytes. Superoxide anion production by kidney macrophages was also elevated. RT-PCR and northern blot analysis of interleukin-1 mRNA showed elevated expression in kidney on day 7 in fish injected with glucan. Glucan had an adjuvant effect on antibody production as pretreatment by injection of 100-1000 microg glucan/fish resulted in the highest antibody titer against A. hydrophila following vaccination. Classical and alternative complement pathways were not affected by glucan administration by any of the three routes.

Topics: Aeromonas hydrophila; Animals; beta-Glucans; Blotting, Northern; Carps; Complement Pathway, Alternative; DNA Primers; DNA, Complementary; Dose-Response Relationship, Drug; Fish Diseases; Gram-Negative Bacterial Infections; Interleukin-1; Kidney; Leukocyte Count; Macrophages; Reverse Transcriptase Polymerase Chain Reaction; Saccharomyces cerevisiae; Superoxides; Survival Analysis