Effect of Coriolus versicolor supplemented diet on innate immune response and disease resistance in kelp grouper Epinephelus bruneus against Listonella anguillarum

The effect of Coriolus versicolor extract supplemented diets on innate immune response and disease resistance in kelp grouper, Epinephelus bruneus against Listonella anguillarum, is reported. Kelp grouper were divided into four groups of 25 each and fed with C. versicolor enriched diets at 0% (control), 0.01%, 0.1%, and 1.0% level. After 30 days of feeding, all fish were injected interaperitoneally (i.p.) with 50 μl of L. anguillarum (4.7 × 10⁷ CFU) to investigate the immune parameters at weeks 1, 2, and 4. The reactive oxygen species and reactive nitrogen species production were significantly enhanced in fish fed with 0.1% and 1.0% supplementation diets from weeks 1-4 when compared to the non enriched diet fed and infected control. The phagocytic activity significantly increased with 0.1% and 1.0% diets on weeks 2 and 4. The leucocyte myeloperoxidase content, lysozyme activity, and total protein level significantly increased when fed with 0.1% and 1.0% supplementation diets from weeks 1-4. The cumulative mortality was 35% and 45% in 1.0% and 0.1% enriched diet fed groups whereas it was 55% and 80% in 0.01% and 0% groups respectively. The present results suggest that diets enriched with C. versicolor at 0.1% or 1.0% level positively enhance the innate immune system and affords protection from L. anguillarum.     

Influence of diet enriched with green tea on innate humoral and cellular immune response of kelp grouper (Epinephelus bruneus) to Vibrio carchariae infection

Effect of diet enriched with green tea at 0, 0.01, 0.1 or 1.0% levels on immune responses such as non-specific humoral (lysozyme, antiprotease and complement) and cellular (myeloperoxidase content, production of reactive oxygen, and nitrogen species) and disease resistance on week 1, 2 or 4 in kelp grouper Epinephelus bruneus challenged with Vibrio carchariae (2.47 × 10(8) CFU ml(-1)) was quantified. At all doses green tea supplementation significantly enhanced the serum lysozyme activity from weeks 1 to 4. On the other hand, after week 2 the serum hemolytic complement activity, leucocyte myeloperoxidase content and reactive nitrogen species protection significantly increased in groups fed with 0.01 and 0.1% green tea supplementation diets. The serum antiprotease activity significantly increased in group fed with at 1.0% green tea from week 1 to 4. However, all diets except at 0.01% level resulted in a significant decrease in reactive oxygen species protection during the experimental period. Challenged groups fed with green tea enriched diet at 0.01 and 0.1% level had a higher relative percent survival than with 1.0% diet on week 1, 2 or 4. The results suggest that dietary administration of green tea supplementation at a concentration of 0.01 and 0.1% level positively enhances the non-specific humoral and cellular immune responses and disease resistance of kelp grouper E. bruneus to V. carchariae.     

Non-specific immune response and disease resistance induced by Siegesbeckia glabrescens against Vibrio parahaemolyticus in Epinephelus bruneus

The immunomodulatory effect of Siegesbeckia glabrescens extract-supplementation diets on innate immune response and disease resistance of kelp grouper, Epinephelus bruneus against Vibrio parahaemolyticus at weeks 1, 2, and 4 is reported. The serum lysozyme activity was significantly enhanced with any enriched diet from weeks 1-4 when compared to control diet (0%). The alternative complement haemolytic activities significantly were enhanced with all enriched diets on weeks 2 and 4 whereas the cellular reactive oxygen species (ROS) was significantly enhanced only with 1.0% and 2.0% diets. The reactive nitrogen intermediate (RNI) value was significantly enhanced with any enriched diet on weeks 2 and 4, but on first week it did not differ from control. The myeloperoxidase (MPO) production significantly increased with 1.0% and 2.0% diets from second week onwards; with other enriched diets the increase manifested on fourth week; but during first week it did not vary from that of the control with any enriched diet. The protection in terms of cumulative mortality was the least being 25% and 20% when fed with 1.0% and 2.0% diets. The present results indicate that feeding kelp grouper with S. glabrescens extract enriched diet at 1.0% and 2.0% levels significantly enhance the immunological parameters, increase the disease resistance and minimize the cumulative mortality in E. bruneus against V. parahaemolyticus.     

Diet enriched with mushroom Phellinus linteus extract enhances the growth, innate immune response, and disease resistance of kelp grouper, Epinephelus bruneus against vibriosis

The effect of diet supplemented with Phellinus linteus fed for 30 days was investigated in grouper Epinephelus bruneus challenged with Vibrio anguillarum, Vibrio harveyi, Vibrio alginolyticus, and Vibrio carchariae; infected and treated fish had a significantly higher percent weight gain and feed efficiency. In groups fed with enriched diet and challenged with V. anguillarum and V. harveyi the mortality rate declined with a consequent rise in survival rate than with other pathogens. On the other hand, in groups fed with P. linteus enriched diet and challenged with V. anguillarum, V. harveyi, and V. alginolyticus the cellular and humoral immune responses, such as the alternative complement activity (ACH(50)), serum lysozyme activity, phagocytic activity (PA), phagocytic index (PI) significantly higher than in the control group. The respiratory bursts (RB), superoxide dismutase (SOD), and glutathione peroxidase (GPx) activities were found significantly enhanced when the groups fed with enriched diet against V. anguillarum and V. harveyi. The results reveal that kelp grouper fed for 30 days with P. linteus enriched diet had higher cellular and humoral immune response and disease protection from vibriosis than the group fed on basal diet with the protection linked to stimulation of immune system.     

Styrax japonica supplementation diet enhances the innate immune response in Epinephelus bruneus against bacterial and protozoan infections

Kelp grouper, Epinephelus bruneus, fed for 30 days with 0% (control), 0.1%, 1.0%, and 2.0% of Styrax japonica supplementation diets, led to reductions in mortality after being challenged with a bacterium (Vibrio harveyi) and a ciliate protozoan (Uronema marinum). The enriched diets significantly increased the survival rate as compared to the controls. The phagocytic and respiratory activities were significantly increased in kelp groupers given 1.0% and 2.0% enriched diets. The complement activity, lysozyme activity, serum bactericidal activity, and total protein level significantly increased with any enriched diet against the pathogens; however antiprotease activity and myeloperoxidase levels significantly increased only with 1.0% and 2.0% enriched diets while the α2-macroglobulin level was significantly enhanced with 1.0% enriched diet. The study suggests that incorporation of S. japonica at 1.0% and 2.0% level in the diet significantly enhances the immune responses in the kelp grouper E. bruneus against V. harveyi and U. marinum.     

Kalopanax pictus as feed additive controls bacterial and parasitic infections in kelp grouper, Epinephelus bruneus

Feeding kelp grouper, Epinephelus bruneus (26.1 ± 1.4), with 0%, 0.1%, 1.0%, and 2.0% Kalopanax pictus extract-supplementation diets, for 30 days reduced mortality on being challenged intraperitoneally 100 μl with bacterium Vibrio alginolyticus (2.1 × 10(7) cfu ml(-1)) and ciliate parasite Philasterides dicentrarchi (2.3 × 10(7) ciliates ml(-1)). The red blood cells (RBC), white blood cells (WBC), haemoglobin, haematocrit, lymphocytes, and monocytes levels significantly increased in kelp grouper fed with all doses of K. pictus-supplementation diets and challenged with bacterium and parasite when compared to control. However, the levels of mean corpuscular volume (MCV), mean corpuscular haemoglobin (MCH), mean corpuscular haemoglobin concentration (MCHC), neutrophils, and thrombocytes did not significantly. The phagocytic activity, complement activity, and antiprotease activity did not significantly change in kelp grouper fed with 0.1% K. pictus-supplementation diets and challenged with bacterium and parasite. The respiratory activity, lysozyme activity, bactericidal activity, total protein level, and myeloperoxidase levels significantly increased in kelp grouper fed with all the doses of K. pictus-supplementation diet and challenged with bacterium and parasite. However, α2-macroglobulin level significantly increased with 1.0% diet, but not with 0.1% and 2.0% diets. Therefore this study suggests that 1.0% and 2.0% K. pictus-supplementation diets positively protected and enhanced the immune system in kelp grouper E. bruneus against V. alginolyticus and P. dicentrarchi infection.     

Long-term effects of passive integrated transponder (PIT) tagging on the kelp grouper Epinephelus bruneus

The effects of passive integrated transponder (PIT) tagging on the growth and survival of kelp grouper Epinephelus bruneus was examined over 1 year. The overall average mass gain for the 1 year period of the abdominal cavity tagged group was higher than the dorsal muscle tagged group. No significant differences were found in overall specific growth rate and food conversion between control and treatment groups.     

Effects of dietary katuk (Sauropus androgynus L. Merr.) on growth,non-specific immune and diseases resistance against Vibrio alginolyticus infection in grouper Epinephelus coioides

This study was conducted to investigate the effect of katuk (Sauropus androgynus L. Merr) on growth, non-specific immune and diseases resistance against Vibrio alginolyticus in grouper (Epinephelus coioides). Grouper juveniles (mean weight 10.97 ± 1.99 g, and length 9.67 ± 0.33 cm) were separated into four groups and cultivated in 100-L tank. Each group was fed with diets containing 0, 1.0, 2.5 and 5.0 g/kg herbs diet twice daily. Fish were sampled for non-specific immune parameters at 0, 1, 2, 4, 7, 14 and 30 days. Results showed that fish received S. androgynus at 1.0 and 2.5 g/kg diets affected the growth and non-specific immune responses. Weight gain, specific growth rate, respiratory burst activity, phagocytosis and reactive oxygen species significantly increased in fish administered with 1.0 and 2.5 g/kg S. androgynus diets. The mortality rate after V. alginolyticus challenge decreased in fish fed with 1.0 g/kg S. androgynus extract. Thus, this study indicated that administration of grouper with S. androgynus supplemented diets can affect the growth performances, diseases resistance and enhances non-specific immune responses.     

Dietary supplementation with chitin and chitosan on haematology and innate immune response in Epinephelus bruneus against Philasterides dicentrarchi

The present study investigated the effect of 1.0% chitin and chitosan supplementation diets on haematology and immune response in Kelp grouper, Epinephelus bruneus against protozoan parasite, Philasterides dicentrarchi. The red blood cells (RBC), white blood cells (WBC), haemoglobin levels, lymphocytes, monocytes, and neutrophils significantly increased in kelp grouper fed with chitin or chitosan enriched diets against P. dicentrarchi. The mean corpuscular volume (MCV), mean corpuscular haemoglobin (MCH), mean corpuscular haemoglobin concentration (MCHC), and thrombocytes did not significantly change against pathogen. The phagocytic activity, respiratory burst activity, complement activity, antiprotease activity, and α2-macroglobulin were significantly enhanced in fish fed with 1% chitin and chitosan diet on weeks 2 and 4. The lysozyme activity, total protein, and myeloperoxidase activity significantly increased in fish fed with chitin or chitosan supplementation diet from weeks 1 to 4 against pathogen. The cumulative mortality was found low in fish fed with chitin and chitosan enriched diets than those of control against pathogen. The present study suggests that supplementation of 1.0% chitin or chitosan in diets positively enhances immune response and affords disease resistance in kelp grouper, E. bruneus against P. dicentrarchi infection.     

The immunostimulatory effects of sodium alginate and iota-carrageenan on orange-spotted grouper Epinephelus coicoides and its resistance against Vibrio alginolyticus

The lysozyme activity, alternative complement activity (ACH50), respiratory burst, SOD (superoxide dismutase) activity and phagocytic activity of orange-spotted grouper Epinephelus coicoides were examined when the fish were injected intraperitoneally with sodium alginate at 10, 20, 30mgkg-1 and iota-carrageenan at 10, 20, 30mgkg-1, respectively after 24, 72 and 120 h. Serum ACH50 increased directly with dose after 24 and 72 h for both sodium alginate and iota-carrageenan treatments. The fish that received sodium alginate at 20mgkg-1 after 24 and 72 h, and the fish that received iota-carrageenan after 72 and 120 h showed significantly increased respiratory burst, SOD activity and phagocytic activity, respectively. In another experiment, E. coicoides which had been injected individually with sodium alginate and iota-carrageenan at 10, 20, 30mgkg-1, were challenged with Vibrio alginolyticus at 1.8x10(9) colony-forming units (cfu)fish-1 and then placed in seawater of 33 per thousand. The survival of fish that received sodium alginate at 20mgkg-1, and the fish that received iota-carrageenan at 30mgkg-1 was significantly higher than that of fish which received saline and the control fish after 48 h as well as at the termination of the experiment (120 h after the challenge). It is therefore concluded that E. coicoides which received sodium alginate at 20mgkg-1 or iota-carrageenan at 30mgkg-1 increased the non-specific immune response and resistance from V. alginolyticus infection.     

Immunomodulatory effect of sodium alginate enriched diet in kelp grouper Epinephelus brneus against Streptococcus iniae

The effect of diets containing sodium alginate at 0, 0.5, 1.0, and 2.0 g kg^?1 following challenge with Streptococcus iniae in kelp grouper Epinephelus bruneus were assessed with reference to survival rate and innate immune parameters such as alternative complement, lysozyme, natural haemagglutination, respiratory burst, superoxide dismutase, and phagocytic activities on week 1, 2, and 4. Fish fed with sodium alginate containing diet at 1.0 and 2.0 g kg^?1 after being challenged with S. iniae had higher survival rates of 75% and 60%, respectively than those fed with control diet (0 g kg^?1). With any enriched diet the percentage of macrophages significantly decreased from week 1–4, while the percentage of neutrophils and lymphocytes significantly increased. The alternate complement activity, natural haemagglutination, and phagocytic activities of infected fish fed with sodium alginate containing diet at 1.0 g kg^?1 on week 2 and 1.0 and 2.0 g kg^?1 diets on week 4 were significantly higher when compared to the control. The lysozyme, respiratory bursts, and superoxide dismutase activities of fish fed with enriched diets at 1.0 and 2.0 g kg^?1 were significantly increased on week 2 and 4. We therefore recommend that at 1.0 or 2.0 g kg^?1 dietary administration of sodium alginate can enhance innate immunity and disease resistance in kelp grouper against S. iniae.     

Propolis and Herba Epimedii extracts enhance the non-specific immune response and disease resistance of Chinese sucker,Myxocyprinus asiaticus

The effect of traditional Chinese medicine (TCM) formulated from propolis and Herba Epimedii extracts at the ratio of 3:1 (w/w) on non-specific immune response of Chinese sucker (Myxocyprinus asiaticus) was investigated. Fish were fed diets containing 0 (control), 0.1%, 0.5% or 1.0% TCM extracts for five weeks. The respiratory burst and phagocytic activities of blood leukocytes, lysozyme and natural haemolytic complement activities in plasma were measured weekly. After five weeks of feeding, fish were infected with Aeromonas hydrophila and mortalities were recorded. Results of this study showed that feeding Chinese sucker with different dosage of TCM extracts stimulated respiratory burst activity, phagocytosis of phagocytic cells in blood and lysozyme activity in plasma. They had no effect on plasma natural haemolytic complement activity. All dosage of treated groups showed reduced mortality following A. hydrophila infection. Feed containing 0.5% TCM extracts was the most effective with the mortality of the fish significantly reduced by 35% compared to the control. The results indicate that propolis and Herba Epimedii extracts in combination enhances the non-specific immune response and disease resistance of Chinese sucker against A. hydrophila.     

Poly D,L-lactide-co-glycolic acid (PLGA)-encapsulated vaccine on immune system in Epinephelus bruneus against Uronema marinum

We investigate the efficacy of poly D,L-lactide-co-glycolic acid (PLGA)-encapsulated vaccine on innate and adaptive immune response in kelp grouper (Epinephelus bruneus) against Uronema marinum at weeks 1, 2, and 4. The respiratory burst (RB) activity, complement activity, and α2-macroglobulin were significantly enhanced in fish immunization with vaccine on week 4 whereas vaccine and PLGA-encapsulated vaccine from weeks 1 to 4. The serum lysozyme activity, antiprotease activity, and antibody level were significantly enhanced in fish immunized with vaccine and PLGA-encapsulated vaccine on weeks 2 and 4. The cumulative mortality was low in PLGA-encapsulated vaccine with 20% whereas high in PLGA and vaccine with 40% and 30%. The results from the present study suggest that PLGA-encapsulated vaccine is useful for further design of immunoprophylatic nano formulation against scuticociliatosis.     

Effect of multiple injections of beta-glucan on non-specific immune response and disease resistance in Labeo rohita fingerlings

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.     

Effect of Sophora flavescens on non-specific immune response of tilapia (GIFT Oreochromis niloticus) and disease resistance against Streptococcus agalactiae

The paper describes the effect of a diet supplemented with the Chinese traditional herbal medicine Sophora flavescens on the immunity and disease resistance of an Oreochromis niloticus GIFT strain. Experimental diets containing 0.025%, 0.050%, 0.100%, 0.200%, and 0.400% S. flavescens, as well as a control group without S. flavescens were used. We tested the non-specific humoral immune responses (lysozyme, antiprotease, and complement) and cellular immune responses (reactive oxygen species and nitrogen species production and myeloperoxidase), as well as disease resistance against Streptococcus agalactiae. S. flavescens supplementation at all dose significantly enhanced serum lysozyme, antiprotease, and natural hemolytic complement activity. Similarly, all S. flavescens doses enhanced cellular myeloperoxidase activity. The increased production of reactive oxygen species and reactive nitrogen intermediates by peripheral blood leucocytes was observed in most of the treatment groups throughout the test period. The fish fed 0.100% S. flavescens had a percent mortality of 21.1% and a relative percent survival of 73.3% compared with the group fed the basal diet during the S. agalactiae challenge. The results suggest that S. flavescens can be recommended as a tilapia feed supplement to enhance fish immunity and disease resistance against S. agalactiae.     

Isolation and Characterization of Vibrio carchariae,a Causative Agent of Gastroenteritis in the Groupers,Epinephelus coioides

An outbreak of serious mortality among the cultured groupers Epinephelus coioides, characterized by a swollen intestine containing yellow fluid, occurred in the summer of 1993 in Taiwan. A motile strain EmI82KL was isolated from the intestinal yellow fluid of the moribund groupers with tryptic soy agar supplemented with 2% NaCl and/or thiosulfate citrate bile salt sucrose agar. This strain was characterized and identified as Vibrio carchariae and was susceptible to chloramphenicol, doxycycline-HCl, nalidixic acid, oxolinic acid, oxytetracycline, and sulfonamide while resistant to ampicillin and penicillin G. In addition, the strain was neither auto-agglutinating nor hemagglutinating, but it was hemolytic against erythrocytes from sheep, rabbit, tilapia, and grouper. The bacteria could be reisolated from kidney, liver, and the transparent yellow fluid of swollen intestine of moribund groupers after bacterial challenge and re-identified as the same species. The LD₅₀ value was 2.53 × 10⁷ colony forming units/g grouper body weight. Received: 26 December 1996 / Accepted: 20 February 1997     

Dietary sodium alginate administration affects fingerling growth and resistance to Streptococcus sp. and iridovirus, and juvenile non-specific immune responses of the orange-spotted grouper, Epinephelus coioides

The percent weight gain (PWG) and feeding efficiency (FE) of fingerling orange-spotted grouper, Epinephelus coioides, fed diets containing sodium alginate at 1.0 and 2.0 g kg(-1) were calculated on the 2nd, 4th, 6th, and 8th weeks after feeding. Survival rates of the fingerling grouper against Streptococcus sp. and an iridovirus, and non-specific immune parameters such as alternative complement activity (ACH50), lysozyme activity, natural haemagglutination activity, respiratory bursts, superoxide dismutase (SOD) activity, and phagocytic activity of juvenile grouper were also determined when the fish were fed diets containing sodium alginate at 0.5, 1.0, or 2.0 g kg(-1). The PWG and FE of fish were better when the fish were fed diets containing sodium alginate at 1.0, and 1.0 and 2.0 g kg(-1), respectively. The PWG and FE of fish fed the 0, 1.0 and 2.0 g kg(-1) sodium alginate-containing diets after 8 weeks were 271.0%, 454.4% and 327.8%, and 0.61, 0.72 and 0.68, respectively. Fish fed a diet containing sodium alginate at the level of 2.0 g kg(-1) had a significantly higher survival rate than those fed the control diet after challenge with Streptococcus sp. and an iridovirus causing an increase of survival rate by 25.0% and 16.7%, respectively, compared to the control group. The ACH(50) level of fish fed the sodium alginate-containing diets at 2.0 g kg(-1) was significantly higher than those fed the 1.0 g kg(-1) sodium alginate diet and control diet after 12 days, and had increased to 1.9-fold, compared to those fed the control diet. The lysozyme activity, phagocytic activity, respiratory bursts, and SOD level of fish fed the sodium alginate-containing diets at 1.0 and 2.0 g kg(-1) were significantly higher than those fed the control diet after 12 days, and had increased to 1.97- and 1.68-fold, 1.35- and 1.50-fold, 1.63- and 1.81-fold, and 1.23- and 1.31-fold, respectively, compared to those fed the control diet. We therefore recommend dietary sodium alginate administration at 1.0 and 2.0 g kg(-1), respectively, to promote growth and enhance immunity and resistance against Streptococcus sp. and an iridovirus.