Effect of Long‐term Oral Administration of an Immunostimulant Diet on Innate Immunity in Sea Bass (Dicentrarchus labrax)

Immunostimulants represent a modern and promising tool in aquaculture, enhancing the resistance of cultured fish to disease and stress. This study investigated the effect of a combination of dietary glucans, α‐tocopherol and ascorbic acid on the innate immune response of cultured sea bass (Dicentrarchus labrax). After 5 weeks of adaptation on a commercial diet containing 100 p.p.m. ascorbic acid and 200 p.p.m. α‐tocopherol, sea bass were switched to a diet supplemented with 2% β?1.3/β?1.6 glucans and ascorbic acid and α‐tocopherol at 500 p.p.m. The supplemented diet was given at 2% of body weight per day over a 2‐week period, every 3 months. Plasma lysozyme concentration, content and distribution of major plasma proteins and complement activity were measured prior to feeding the supplemented diet and after 40 weeks. Alternative pathways of complement activation and lysozyme activity were both significantly enhanced in fish fed on glucans and elevated doses of vitamins. No significant differences were observed in protein content or in albumin/globulin ratio. Compared to lysozyme activity, which showed marked individual variation, complement‐mediated haemolytic activity has been shown to be a more reliable indicator of sea bass immunocompetence. Further studies are in progress to clarify the effect of each dietary component on the innate immune response and disease resistance.     

Effect of oral administration of high vitamin C and E dosages on the gilthead seabream (Sparus aurata L.) innate immune system

The effect of the oral administration of high dosages of vitamins C and E on the innate immune system of the seabream was investigated. Gilthead seabream (Sparus aurata L.) were fed four different diets for 45 days: a commercial diet as control, a 3g/kg vitamin C-supplemented diet, a 1.2g/kg vitamin E-supplemented diet or a diet containing both vitamin supplements. After 15, 30 and 45 days, serum ascorbic acid and alpha-tocopherol levels, growth, complement titers and head-kidney leucocyte phagocytic and respiratory burst activities were evaluated. The results showed that serum vitamin levels reflected dietary input. Fish fed the vitamin C-supplemented diet showed an enhanced respiratory burst activity, while fish fed the vitamin E-supplemented diet exhibited increased complement and phagocytic activities. All of these positive effects were found in fish fed the joint diet, as well as a synergistically enhanced respiratory burst activity at day 30. The results demonstrate that in vivo vitamins C and E exert a synergistic effect enhancing the respiratory burst activity of seabream phagocytes.     

饲料中添加肌肉生长抑制素抑制肽对海鲈生长性能、体组成、血清生化指标及肝脏与血清免疫指标的影响

本试验旨在研究饲料中添加肌肉生长抑制素抑制肽对海鲈生长性能、体组成、血清生化指标及肝脏与血清免疫指标的影响。以初始均重为(9.05±0.05)g的海鲈幼鱼为试验动物,暂养1周后,挑选规格一致的健康试验鱼480尾,随机分为4组,每组4个重复(水族箱),每个重复放养30尾鱼。4组试验鱼分别投喂肌肉生长抑制素抑制肽添加水平为0(对照)、0.25%、0.50%、0.75%的试验饲料135 d。结果显示:1)0.50%组的增重率(WGR)、特定生长率(SGR)和成活率(SR)均显著高于对照组(P0.05),各添加组的饲料系数(FCR)均显著低于对照组(P0.05),且以0.50%组的FCR最低;2)各添加组的背肌粗脂肪含量均显著低于对照组(P0.05),0.50%组的背肌粗蛋白质含量显著高于对照组(P0.05),全鱼的水分、粗蛋白质、粗脂肪、粗灰分和背肌的水分、粗灰分含量在各组之间没有显著差异(P0.05);3)0.25%组的血清总蛋白(TP)、白蛋白(ALB)和总胆固醇(TC)含量显著低于其他各组(P0.05),血清甘油三酯(TG)含量各组间不存在显著差异(P0.05);4)0.50%和0.75%组的血清碱性磷酸酶(ALP)活性显著高于对照组(P0.05),0.25%和0.50%组的肝脏溶菌酶(LZM)活性显著高于对照组(P0.05),肝脏与血清的总抗氧化能力(T-AOC)和肝脏超氧化物歧化酶(SOD)活性各组间不存在显著差异(P0.05)。综上所述,在饲料中添加肌肉生长抑制素抑制肽能够促进海鲈的生长并提高免疫能力,本试验条件下最适添加水平为0.50%。     

Effect of Low Dietary Magnesium on Immune Response and Osmoregulation of Atlantic Salmon

Abstract

The influences of dietary magnesium on immune response and on osmoregulation in parr of Atlantic salmon Salmo salar were determined. Groups of fish were fed a casein–gelatin diet unsupplemented (containing about 200 mg Mg/kg) or supplemented with either 300 or 500 mg Mg/kg dry diet (as MgSO₄) for 12 weeks before vaccination to produce fish with different Mg levels, and the feeding regime was continued throughout the study. No differences were observed between the treatment groups in serum-specific antibody levels every second week for 8 weeks after vaccination against Vibrio anguillarum serotypes O1 and O2. Both lysozyme levels and spontaneous hemolytic activities in serum were elevated in vaccinated fish compared with unvaccinated fish. Neither lysozyme activity, complement hemolytic activity, total protein in serum nor blood hemoglobin were affected by dietary Mg. The spontaneous hemolytic activity in serum was lower in fish fed the unsupplemented diet and highest in fish fed the diet supplemented with 500 mg Mg/kg. After 28 weeks on the diets supplemented with graded levels of Mg, a salinity tolerance test (32.5 g/L, 24 h) was performed. High mortality and elevated serum chloride concentrations in all groups after 24 h reflected a general salinity intolerance, but the highest serum chloride level was observed in fish fed the unsupplemented diet. This indicates that low dietary Mg affects the osmoregulation of Atlantic salmon.     

大口黑鲈维生素A需求量

为了评定大口黑鲈饲料中维生素A的需求量,本研究在基础饲料中添加不同量(0、1 000、2 000、3 000和4 000 IU/kg)的维生素A醋酸酯,配制5种不同维生素A含量(实测维生素A含量分别为415、1 406、2 755、3 501和4 646 IU/kg)的等氮等能饲料(粗蛋白质含量48.47%;总能18.15 M J/kg)。用上述饲料饲喂初始体重为(52.75±0.12)g的大口黑鲈49 d,每个处理设3个重复,每重复30尾鱼。结果表明:随着饲料中维生素A水平从415 IU/kg升高到2 755 IU/kg,试验鱼的增重率、特定生长率、饲料效率和蛋白质效率均显著升高(P0.05);而随饲料维生素A水平的进一步增高,上述指标则呈平稳的趋势。随着饲料中维生素A水平从415 IU/kg升高到3 501 IU/kg,鱼的肝脏中维生素A含量、肝脏总超氧化物歧化酶活性、血清溶菌酶活性,头肾白细胞呼吸爆发活性、血清补体活性及血红蛋白含量显著升高(P0.05),而肝脏丙二醛含量显著降低(P0.05);随饲料维生素A水平的进一步增高,上述指标变化幅度较小。饲料中不同维生素A水平对鱼的血清总蛋白含量和红细胞压积的影响不显著(P0.05)。分别以增重率和肝脏维生素A含量为评价指标,采用折线模型求得,大口黑鲈对饲料中维生素A的最适需求量分别为2 600和3 550 IU/kg;而以非特异性免疫为评价指标,通过方差分析得出的最适需求量与以肝脏维生素A含量为评价指标所得出的需求量相近。本研究得出,大口黑鲈对饲料中维生素A水平的最适需求量为2 600~3 550 IU/kg。     

Influence of Dietary Ascorbic Acid on the Immune Responses of Juvenile Korean Rockfish Sebastes schlegelii

Juvenile Korean Rockfish Sebastes schlegelii (length, 13.6 ± 1.4 cm [mean ± SD]; weight, 53.6 ± 4.2 g) were fed twice daily with diets containing varying levels of ascorbic acid (0, 50, 100, 200, and 400 mg/kg) for 4 weeks. Significant increases in daily weight and length occurred in fish fed more than 50 mg/kg ascorbic acid. The lysozyme activity of fish fed diets containing ascorbic acid was considerably increased in rockfish plasma at 400 mg/kg ascorbic acid and in kidney at over 50 mg/kg ascorbic acid. Total plasma immunoglobulin M (IgM) levels were markedly elevated in fish fed 400 mg/kg ascorbic acid. The results suggest that dietary ascorbic acid supplementation in juvenile rockfish can induce a significant increase in growth and in immunological features such as lysozyme activity and plasma IgM levels.

Received October 1, 2014; accepted May 2, 2015     

Effect of vitamin E and fat source in sows' diets on immune response of suckling and weaned piglets

Thirty-six 7-mo-old gilts were used to study the effects of dietary vitamin E and fat source (5% sunflower oil or animal fat) in pregnant and lactating sow diets on serum vitamin E concentration and on cell-mediated and humoral immune response in suckling and weaned piglets. Six gilts each received one of six diets throughout pregnancy and lactation. The basal diets (13 mg alpha-tocopherol/kg diet) were supplemented with dl-alpha-tocopheryl acetate to 48 and 136 mg alpha-tocopherol/kg of feed (average analyzed values). After weaning (at 4 wk of age) all pigs received identical diets (20 mg of alpha-tocopherol/kg feed). One week after weaning, pigs were immunized (i.m. with ovalbumin and tetanus toxoid) and antibody production was measured. Blood samples were taken immediately after birth, at 1 wk after birth, at weaning, and at four weekly intervals after weaning. Samples were analyzed for alpha-tocopherol concentration, total number of leukocytes, T- and B-lymphocytes, lymphocyte stimulation with concanavalin A, lysozyme activity, and immunoglobulin concentrations. It was concluded that a high vitamin E level in the sow's diet increased serum vitamin E concentration of 1-wk-old pigs (P less than .05). Immune response against ovalbumin was increased (P less than .05) at 1 wk of age after immunization for weaned pigs from sows fed the high level of vitamin E. Also, the phagocytic measures of pigs at 1 wk of age were increased by the medium vitamin E level (P less than .05). Fat sources in the sow's diet had no consistent effect on the immunological measures of pigs.     

Effects of dietary supplementation with vitamin C or vitamin E on cardiac lipid peroxidation and growth performance in broilers at risk of developing ascites syndrome

OBJECTIVE: To assess effects of high dietary amounts of vitamin C or vitamin E and oxidative stress on the heart and growth performance of broilers maintained at an altitude of 2,200 m above sea level. ANIMALS: 360 chicks (1-day-old broilers). PROCEDURE: Birds were randomly assigned to 3 groups (120 chicks/group). Each group of birds was fed a specific diet (control group, basal diet containing 12 mg of vitamin E (DL-alpha-tocopherol acetate)/kg of feed without additional ascorbic acid; vitamin E group, basal diet supplemented with 75 mg of vitamin E/kg of feed; and vitamin C group, basal diet supplemented with 400 mg of ascorbic acid/kg of feed) throughout the entire 7 weeks of the study. Feed consumption and body weight of chicks were recorded on a weekly basis. Nine randomly selected birds from each group were euthanatized each week. Remaining birds were euthanatized at the end of the study. Samples of cardiac tissues were obtained to measure thiobarbituric acid reactive substances (TBARS), an indicator of oxidative stress. RESULTS: Vitamin E-supplemented diets resulted in better growth performance, lower rates of feed conversion, and lower TBARS content. Vitamin C-supplemented diets resulted in lower feed consumption and lower rates of feed conversion. When used separately, neither of the vitamins had any effect on mortality attributable to ascites syndrome. CONCLUSION AND CLINICAL RELEVANCE: It is recommended that diets supplemented with vitamin C, vitamin E, or both be fed to broilers maintained at an altitude of 2,200 m above sea level to improve growth performance.     

Oral administration of yeast, Saccharomyces cerevisiae, enhances the cellular innate immune response of gilthead seabream (Sparus aurata L.)

The effects of including lyophilised whole yeast, Saccharomyces cerevisiae, in the diet on the seabream innate immune response were investigated. Gilthead seabream (Sparus aurata L.) specimens were fed four different diets for 4 weeks: a commercial diet as control and the same diet supplemented with 1, 5 or 10 g/kg yeast. After 1, 2 and 4 weeks, serum complement titres, as a humoral parameter, and phagocytic, respiratory burst, myeloperoxidase and natural cytotoxic activities of head-kidney leucocytes, as cellular parameters, were evaluated. The results showed that yeast supplements enhanced all the latter responses, but not the humoral response. This enhancement was dose-dependent except for the cytotoxic activity that was only stimulated by the lower dose of yeast assayed. As yeast cell walls are able to enhance the seabream cellular innate immune response, these results support the possible use of whole yeast as natural inmunostimulants in common fish diets.     

Effect of Alpha-Tocopherol and Ascorbic Acid on Bovine Oocyte in Vitro Maturation

In vitro culture results in higher oxygen concentrations than in vivo environments, leading to an increased level of reactive oxygen species (ROS) that cause lipid peroxidation of cellular membranes. Alpha-tocopherol (active form of vitamin E) is an antioxidant that protects mammalian cells against lipid peroxidation, which is regenerated by ascorbic acid. The aim of this study was to determine the effect of the addition of alpha-tocopherol and/or ascorbic acid to the maturation medium on bovine oocyte in vitro maturation (IVM) and subsequently on in vitro fertilization (IVF) and embryo development. Cumulus-oocyte complexes (COCs) were matured in Medium 199 (control), and with the addition of alpha-tocopherol and/or ascorbic acid. The concentration of alpha-tocopherol in COCs was determined by high-performance liquid chromatography (HPLC). IVF and in vitro culture (IVC) were carried out in modified synthetic oviductal fluid (mSOF). The quantity of alpha-tocopherol naturally present in COCs diminished by half during IVM (p < 0.05), although in the presence of ascorbic acid it remained constant. A greater amount of alpha-tocopherol was detected in COCs matured in medium supplemented with this antioxidant (p < 0.05), but the addition of alpha-tocopherol plus ascorbic acid maintained higher levels of alpha-tocopherol (p < 0.05). Significant differences were not observed in the percentages of nuclear maturation and fertilization among different treatments. The presence of alpha-tocopherol or ascorbic acid in the maturation medium failed to modify the percentage of blastocysts obtained, unlike the addition of both antioxidants when a significant decrease was observed (p < 0.05). Absorbic acid maintained the antioxidant capacity of the alpha-tocopherol incorporated to COC membranes during IVM. The active form of vitamin E during maturation impaired the acquisition of oocyte developmental competence.     

饲料维生素A对美国红鱼生长和免疫的影响

本试验旨在研究饲料维生素A对美国红鱼生长(增重率、特定生长率、存活率和饲料效率)和免疫反应相关指标(血清中溶菌酶活性、超氧化物歧化酶活性和总补体活性)的影响.采用单因子试验设计,将300尾平均体重为(151.46±8.18)g的美国红鱼随机分为5个处理,每个处理设3个重复,每个重复20尾,分别饲喂5种不同VA水平(30、4 166、8 533、18 532、33 263 IU/kg)的等氮等能饲料,进行为期8周的生长试验.结果表明:饲料中VA水平为30 IU/kg时其增重率、饲料效率、特定生长率、成活率均显著低于4 166、8 533、18 532 IU/kg组(P<0.05),VA水平为33 263 IU/kg时各指标有所降低.其中饲料效率和特定生长率显著低于4 166、8 533、18 532 IU/kg组(P<0.05);VA水平为30 IU/kg时血清溶菌酶活性,SOD活性和总补体活性显著低于其他组(P<0.05),VA水平在18 532 IU/kg时血清溶菌酶活性达到最高值,其次为33 263、8 533、4 166 IU/kg组;VA水平在8 533 IU/kg时血清SOD和总补体活性达到最高值,并显著高于4 166、18 532、33 263IU/kg组(P<0.05).综上所述,饲料中适宜VA水平能显著促进美国红鱼生长、提高相关免疫力,且饲料VA水平为8 533 IU/kg时对美国红鱼生长和免疫相关指标影响较显著.     

Immunomodulation by dietary vitamin C in healthy and aflatoxin B1-induced immunocompromised rohu (Labeo rohita)

The aim of this study was to examine the immunomodulatory effect of high levels of dietary vitamin C in healthy and immunocompromised rohu (Labeo rohita) treated with aflatoxin B1 (AFB1). Four groups of rohu were fed experimental diets containing either no vitamin C or supplemented with vitamin C at 500 ppm for 60 days. On the first day of feeding, one group fed the high vitamin C diet and one fed the vitamin C deficient diet, were injected intraperitoneally with a single doses of AFB1 at 1.25 mg kg(-1) body weight. The effect of AFB1 and high dietary vitamin C on specific and non-specific immunity, and disease resistance against Aeromonas hydrophila were examined in the rohu. The ability of vitamin C to counteract immunosuppression induced by AFB1 was also examined. Specific immunity indicated by haemagglutination and haemolysin titres against sheep red blood cells (SRBC), and bacterial agglutination appeared to be unaffected by either the AFB1 treatment or the vitamin C enriched diet. A significant reduction was observed in the non-specific immunity of AFB1-treated fish, however, indicated by lowered bactericidal and lysozyme activities. High dietary vitamin C, on the other hand, enhanced the non-specific immunity of fish, including an enhanced phagocytic ratio and increased serum lysozyme activity. Feeding a high level of dietary vitamin C to AFB1-treated fish increased these parameters to levels similar to those found in control fish. High dietary vitamin C significantly (p < 0.05) enhanced protection against Aeromonas hydrophila infection in both healthy and immunocompromised fish. Results from this study help to establish the beneficial effect of dietary vitamin C on AFB1-induced immunosuppression, as well as confirming the immunostimulatory effect of vitamin C in rohu.     

Effect of a mixed herb-enriched diet on the innate immune response and disease resistance of Paralichthys olivaceus against Philasterides dicentrarchi infection

We investigated the effect of a mixed herb-enriched diet obtained from pomegranate Punica granatum, Dalmatian chrysanthemum Chrysanthemum cinerariaefolium, and mastic-leaved prickly-ash Zanthoxylum schinifolium on innate immune mechanisms (e.g., phagocytosis activity, respiratory burst activity, alternative complement activity, lysozyme activity, and disease resistance) of olive flounder Paralichthys olivaceus against the scuticociliate Philasterides dicentrarchi. All experimental groups were challenged with P. dicentrarchi (1 x 10(5) ciliates/mL) through intraperitoneal administration of the pathogen (50 microL) on day 1. On day 7, the infected groups were fed 0, 5, 50, and 100 mg/kg of the enriched diets. The innate immune parameters, cumulative mortality, and relative percent survival (RPS) were assessed at weeks 1, 2, 3, and 4. Administration of 50 or 100 mg/kg of the herbal-enriched diet enhanced immunity throughout the experimental period. However, at the 5-mg/kg dosage, the enriched diet did not enhance the innate immune estimates at any time. At doses of 50 and 100 mg/kg, administration of the diet preceding the challenge with P. dicentrarchi decreased the percentage cumulative mortality in the experimental groups and thereby increased RPS values. This study reports that administration of 50 or 100 mg/kg mixed herbal-enriched diet could positively influence the innate immune response to P. dicentrarchi and enhance the health status of olive flounder with respect to this microbe.     

The partial substitution of digestible protein with gelatinized starch as an energy source reduces susceptibility to lipid oxidation in rainbow trout (Oncorhynchus mykiss) and sea bass (Dicentrarchus labrax) muscle

We evaluated the influence of dietary gelatinized starch and protein on the fatty acid composition of muscle in rainbow trout and European sea bass and on the susceptibility of flesh to lipid peroxidation. The possibility that flesh peroxidation could be accounted for by lipogenesis and the deposition of fat was also explored. The inclusion of gelatinized starch in the diet of rainbow trout improved growth with respect to that observed in fish fed crude starch (P<.001). This was especially noticeable at the lowest concentration of dietary protein tested (P = .037); suggesting that gelatinized starch may partially replace protein in the production of energy without inducing a negative effect on growth. However, in European sea bass, the gelatinization of starch and dietary protein concentration showed no significant effect on final body weight. The intramuscular neutral lipid concentration of the sea bass was reduced by the gelatinization of dietary starch (P = .034). The highest dietary protein concentration increased the proportion of saturated fatty acids in the neutral (P = .0742) and polar (P = .0033) lipid fractions. The dietary inclusion of high levels of protein in rainbow trout led to a lower concentration of total (n-3) (P = .0457) and (n-6) (P = .0522) fatty acids and a higher concentration of total monounsaturated fatty acids (P = .0006). The inclusion of gelatinized starch led to a lower concentration of (n-3) fatty acids (P = .0034) and a higher concentration of saturated fatty acids (P = .0007). The polar fraction was hardly affected by the same treatment. A significantly lower susceptibility of the dorsal muscle to oxidation was observed in groups of European sea bass fed gelatinized starch (P<.01). A similar trend was observed in rainbow trout, although differences were not significant. The findings suggest that the digestible protein concentration of nutrient-dense diets for rainbow trout and European sea bass can be reduced with a beneficial effect on tissue lipid oxidation and no negative effects on growth and muscle composition.     

Antiparasitic and immunomodulatory effect of innovative treatments against Myxobolus sp. infection in Diplodus puntazzo

The potential antiparasitic and immunomodulatory effect of three treatments against myxosporean parasites on the innate immune system of sharpsnout sea bream (Diplodus puntazzo) was investigated. Fish naturally infected with Myxobolus sp. (Bivalvulida/Platysporina), a histozoic parasite mainly affecting the renal interstitial tissue, were treated by oral administration of a combination of salinomycin with amprolium, Origanum essential oil or fumagillin in a small-scale field trial. Various leucocyte functions influenced by myxosporean infection were examined in order to determine treatment effects on leucocyte immunocompetence of treated fish. One month post treatment all drugs caused a significant decrease in prevalence and intensity of infection in comparison to untreated, infected fish. The effect was most prominent in salinomycin with amprolium treated fish, which 1-month post treatment contained either no cysts at all or a few spores free in melanomacrophage centres revealing almost total elimination of the parasite and the antiparasitic action of the treatment. There was no histopathological evidence of drug toxicity. Antiparasitic action was accompanied by a significant enhancement of phagocytic activity demonstrated by ingestion of large numbers of latex beads and the secretion of high levels of reactive nitrogen intermediates by phagocytes in vitro. Complete restoration of the diminished mitogenic responses and serum lysozyme secretion was also detected in salinomycin with amprolium-treated fish compared to untreated, infected fish. These data suggest that salilomycin with amprolium may be a promising treatment for myxosporean infections in intensively cultured warm-water fish, exhibiting action partially via the enhancement of host, innate immune functions and leading to parasite elimination.     

Dietary lutein and fish oil interact to alter atherosclerotic lesions in a Japanese quail model of atherosclerosis

Interactions between concentration of dietary lutein and fish oil in diets on atherosclerosis incidences were studied in a cholesterol-induced-atherosclerosis (CIA) model. CIA Japanese quail were fed a basal diet with three amounts of lutein (0, 25 and 50 mg/kg diet) and two amounts of fish oil (3% and 6%) in a 3 × 2 factorial in five replications. Samples were collected at 24 and 27 weeks of age. Atherosclerosis lesions in the dorsal aorta were measured by histochemistry sectioning. At 27 weeks of age, increasing dietary fish oil content to 6% decreased (p < 0.01) the atherosclerotic lesions only in the 0 mg lutein supplemented groups. At 27 weeks of age, increasing dietary fish oil content to 6% increased the atherosclerotic lesion score when lutein was supplemented at either 25 or 50 mg/kg feed. Aorta and liver lutein content increased (p < 0.01) with increasing dietary lutein content at 27 weeks of age. Increasing dietary fish oil content to 6% increased (p < 0.01) the aorta fat content by twofold and decreased (p < 0.01) the liver fat by 26% at 27 weeks of age. Increasing the dietary fish oil content to 6% increased (p = 0.01) the total PUFA and decreased (p = 0.03) the total mono unsaturated fatty acids content of the aorta at 27 weeks of age. At 27 weeks of age, increasing dietary fish oil content to 6% decreased the amount of TBARS (p = 0.01) and IL-1 mRNA (p < 0.01) only in the 0 mg lutein supplemented groups. Increasing dietary fish oil content to 6% increased the amount of TBARS and IL-1 mRNA of the aorta when lutein was supplemented at either 25 or 50 mg/kg diet. Dietary lutein supplementation decreased atherosclerosis lesions only at low levels of dietary polyunsaturated fatty acids.     

The effects of five different glycans on innate immune responses by phagocytes of hybrid tilapia and Japanese eels Anguilla japonica

The aim of this study was to evaluate the immune responses in hybrid tilapia (Nile tilapia Oreochromis niloticus x Mozambique tilapia O. mossambicus) and Japanese eels Anguilla japonica after treatment with five glycans: barley, krestin, MacroGard, scleroglucan, and zymosan. The effects of the glycans on the innate immune responses of the fish were investigated using the phagocytic index (PI), lysozyme activity, complement opsonization, and activation assay. The results of the lysozyme assay demonstrated that the lysozyme activities increased after treatment with glycans. Moreover, based on the PI, treatment with each of the five glycans resulted in increased phagocytic activities in anterior kidney and peripheral blood phagocytes in both tilapia and Japanese eels. The opsonic effect of complement on phagocytosis in tilapia and Japanese eels were investigated using baker's yeast, which served as the activator in the classical complement pathway (CCP) and in the alternative complement pathway (ACP). Tilapia and Japanese eel sera that were treated with glycans greatly enhanced phagocytosis. The classical pathway--hemolytic complement titer (CH50) of Japanese eels treated with glycans was slightly increased in vitro and in vivo. While glycan treatment enhanced the CCP of both species in vitro and in vivo, the alternative pathway-hemolytic complement titer (ACH50) was only increased in vitro and in vivo in glycan-treated tilapia. Thus, it follows that the ACP must have been activated in tilapia treated with glycans. However, in Japanese eels, the ACH50 of the ACP activation assay was undetected in vitro or in vivo due to possible unknown factors in the Japanese eel serum that caused lysis of the rabbit red blood cells. Our study investigated the effects of glycans used to enhance phagocytosis and activate both of the complement pathways involved in stimulating the innate immune responses of Japanese eels and tilapia.     

Immunostimulant properties of a cell wall-modified whole Saccharomyces cerevisiae strain administered by diet to seabream (Sparus aurata L.)

The possible differences in the immunostimulant effects between whole wild Saccharomyces cerevisiae and a mutant strain (fks-1) administered in the diet to gilthead seabream (Sparus aurata L.) were studied. Fish were fed diets containing 0 (control) or 10 g whole wild yeast or fks-1 strain per kilogram feed for 2, 4 or 6 weeks. After each sampling, the innate humoral (complement, lysozyme and peroxidases) and cellular (intracellular peroxidases, phagocytosis, respiratory burst activity and cytotoxicity) responses were determined. Among the humoral responses, serum peroxidases and complement activity were significantly decreased after 6 weeks of feeding with the fks-1 strain-supplemented diet, while lysozyme activity increased after 2 and 4 weeks of feeding the fks-1 strain. Of the cellular responses studied, phagocytosis was increased to a significant degree at all the assayed times but only by the fks-1 strain-supplemented diet, while respiratory burst activity (after 4 weeks) and natural cytotoxicity (after 4 and 6 weeks) increased with either yeast strain. The intracellular peroxidase content was not affected by the dietary supplementation of either yeast strain. These results are discussed with a view to assessing the possible use of wild and/or mutant yeast strains for immunostimulant purposes in aquaculture.     

Korean mistletoe enriched diet enhances innate immune response in kelp grouper, Epinephelus bruneus against Philasterides dicentrarchi

The present study investigated the immunostimulatory effect of Korean mistletoe Viscum album extract (KM-E) on innate immune response in kelp grouper Epinephelus bruneus against Philasterides dicentrarchi. Kelp grouper were divided into four groups of 25 each and fed with 0 (control), 0.5, 1.0, and 2.0% enriched diets with Korean mistletoe extract (KM-E). After feeding for 30 days, the fish were injected intraperitoneally (i.p.) with 100 μl of P. dicentrarchi (4.2 × 10(7)ciliates/ml) to study the immune responses at weeks 1, 2, and 4. The respiratory burst activity did not significantly enhance when fed with 0.5% and 1.0% supplementation diets on week 1 when compared to control diet. On weeks 2 and 4, the respiratory burst activity significantly increased with 1.0% and 2.0% diets. The phagocytic activity significantly enhanced with 1.0% and 2.0% diets, but not with 0.5% diet at any time. When fed with 1.0% KM-E-diet the lysozyme activity did not significantly vary at any week whereas with 1.0% and 2.0% diets it was significantly enhanced. The total protein level significantly increased with 1.0% and 2.0% KM-E-diets from weeks 1 to 4 as compared to control. The present study suggests that 1.0% or 2.0% KM-E-supplementation diet positively enhances the innate immune response in E. bruneus against P. dicentrarchi infection.