Evaluation of solid‐state fermented protein concentrates as a fish meal replacer in the diets of juvenile rainbow trout,Oncorhynchus mykiss

We evaluated four fermented protein concentrates (FPCs) as a fish meal replacer in juvenile rainbow trout, Oncorhynchus mykiss. Ten diets were formulated to contain low‐temperature fish meal (LT‐FM), Vietnam fish meal (VT‐FM) and four types of FPCs as fish meal replacers (FPC‐A, B, C & D) at 30% and 50% FM replacement levels. FPC‐A was a mixture of solid‐state fermented soybean meal (SBM) and corn gluten meal (CGM) with Bacillus subtilis; FPC‐B was pretreated acid‐hydrolysed FPC‐A; FPC‐C and FPC‐D were FPC‐A + 2% shrimp soluble extract (SSE) and FPC‐B + 2% SSE, respectively. Triplicate groups of fish (average 15.4 g) were fed one of the experimental diets for 8 weeks. At the end of the feeding trial, fish fed the LT‐FM, FPC‐B, FPC‐C and FPC‐D diets showed significantly higher growth performance at 30% FM replacement than those of fish fed the FPC diets at 50% FM replacement. Fish fed 30FPC‐B, 30FPC‐C and 30FPC‐D diets showed higher weight gain (WG) than fish fed 30FPC‐A diet. Haematological parameters showed no clear trends among the experimental groups. Superoxide dismutase, lysozyme and myeloperoxidase activities were found to be higher in fish fed the LT‐FM, FPC‐A, FPC‐B, FPC‐C and FPC‐D diets at 30% FM replacement than in fish fed the FPC diets at 50% FM replacement. Antinutritional factors such as α‐ and β‐conglycinin, glycinin, trypsin inhibitors or zein were absent in FPC compared with SBM and CGM. Intestinal villi length and muscular thickness were significantly reduced in fish fed the VT‐FM and FPC diets at 50% FM replacement compared to fish fed the LT‐FM and FPC diets at 30% FM replacement. The results show that FPC‐B, FPC‐C, FPC‐D could replace up to 30% of LT‐FM in juvenile rainbow trout.     

Increase in the plant protein ratio in the diet of white shrimp,Litopenaeus vannamei (Boone), using Bacillus subtilis E20‐fermented soybean meal as a replacement

This study aimed to improve the nutritional value and utilization of soybean meal (SBM) used as feed ingredient for shrimp by an approach of solid‐state fermentation with Bacillus subtilis E20. The protein content and B. subtilis E20 proliferation significantly increased, as the initial moisture increased from 30% to 50% during fermentation. Compared with SBM, the protein content of FSBM increased by 19% after fermentation, accompanied by an increase of 18.75% in the total hydrolyzed amino acids. The free amino acid profile and content in FSBM also obviously increased by 374.9% compared with SBM. The FSBM is a good substitute for fish meal (FM) in a diet with 37% protein and 7% lipid content. The maximal replacement levels of FM in shrimp diet with SBM and FSBM were 37.42% and 61.67%, respectively, based on the feed efficiency estimated by a broken‐line analysis. The B. subtilis E20‐FSBM can be a potential protein source used as a replacement for FM in shrimp diet.     

Effect of dietary hydrolysate supplementation on growth performance,non‐specific immune response and disease resistance of olive flounder (Paralichthys olivaceus) challenged with Edwardsiella tarda

A 9‐week feeding trial was conducted to investigate the effects of dietary supplementation with protein hydrolysates on growth, innate immunity and disease resistance of olive flounder. A fishmeal (FM)‐based diet was regarded as a control, and three diets were prepared by partial replacement of FM with krill hydrolysate, shrimp hydrolysate or tilapia hydrolysate (designated as Con, KH, SH and TH, respectively). Triplicate groups of fish (24.5 ± 0.3 g) were fed one of the diets to apparent satiation twice daily for 9 weeks and then challenged by Edwarsellia tarda. Fish‐fed KH diet showed significantly (P < 0.05) higher growth performance and feed utilization compared with the Con diet. Dry matter digestibility of the diets was significantly increased by KH and TH supplementation. All the examined innate immune responses were significantly increased in fish fed KH diet. Significantly, higher respiratory burst and superoxide dismutase (SOD) activities were found in fish‐fed SH diet. Lysozyme and SOD activities were significantly increased in fish‐fed TH diet. However, no significant effect was found on fish disease resistance. This study indicates that dietary supplementation of the hydrolysates, particularly KH, can improve growth performance, feed utilization and innate immunity of olive flounder.     

Supplementation of Protein Hydrolysates to a Low‐fishmeal Diet Improves Growth and Health Status of Juvenile Olive Flounder,Paralichthys olivaceus

An 11‐wk feeding trial was conducted to evaluate three different protein hydrolysates as feed ingredients in high‐plant‐protein diets for juvenile olive flounder. Five experimental diets were fed to juvenile olive flounder to examine the effect of three different protein hydrolysates on growth performance, innate immunity, and disease resistance against bacterial infection. A basal fishmeal (FM)‐based diet was regarded as a high‐FM diet (HFM) and a diet containing soy protein concentrate (SPC) as a substitute for 50% FM protein was considered as a low‐FM diet (LFM). Three other diets contained three different sources of protein hydrolysates, including shrimp, tilapia, and krill hydrolysates (designated as SH, TH, and KH), replacing 12% of FM protein. All diets were formulated to be isonitrogenous and isocaloric. Triplicate groups of fish (15.1 ± 0.1 g) were handfed one of the diets to apparent satiation twice daily for 11 wk and subsequently challenged against Edwardsiella tarda. Growth performance and feed utilization of fish fed hydrolysate‐supplemented diets were significantly improved compared to those of fish fed the LFM diet. Dietary inclusion of the protein hydrolysates significantly enhanced apparent digestibility of dry matter and protein of the diets. In the proximal intestine, histological alterations were observed in the fish fed the LFM diet. The fish fed the hydrolysate diets showed significantly longer mucosal fold and enterocytes and greater number of goblet cells compared to fish fed the LFM diet. Respiratory burst activity was significantly higher in fish fed the TH and KH diets than fish fed the LFM diet. Significantly higher immunoglobulin levels were found in fish fed SH and KH diets compared to those of fish fed the LFM diet. Dietary inclusion of the protein hydrolysates in SPC‐based diets exhibited the highest lysozyme activity. Significantly higher superoxide activity was observed in groups of fish fed the KH diet. Fish offered the protein hydrolysates were more resistant to bacterial infection caused by E. tarda. The results of this study suggest that the tested protein hydrolysates can be used as potential dietary supplements to improve growth performance and health status of juvenile olive flounder when they were fed a LFM diet.     

Using Bacillus subtilis E20‐fermented soybean meal as replacement for fish meal in the diet of orange‐spotted grouper (Epinephelus coioides,Hamilton)

The potential of Bacillus subtilis E20‐fermented soybean meal (FSBM) as a partial alternative component of fish meal (FM) in fed diets of orange‐spotted grouper (Epinephelus coioides) was evaluated in this study. An FM‐based diet and seven diets containing 10%, 20% and 30% and 10%, 20%, 30% and 40% of FM replaced by soybean meal (SBM) and FSBM, respectively, were fed to grouper for 84 days to evaluate possible substitution levels of FM by tracking growth performance, digestive enzyme activity and morphological changes in the liver and distal intestine. No significant differences in survival and muscle composition of grouper were found between controls and treatments. Growth performance and feed efficiency of fish fed diets with FM replaced by FSBM up to 30% were not significantly different from controls, whereas significantly decreased growth performance and feed efficiency occurred with diets containing >20% of SBM. Based on the feed efficiency, the maximum substituted levels of FM by SBM and FSBM in grouper diets were 18.36% and 29.32%, respectively, based on broken‐line analyses. Histopathological changes in the liver and distal intestine, and significantly lower activity levels of digestive enzymes, including pepsin in the stomach and trypsin, chymotrypsin, amylase and lipase in the distal intestine, were found in fish fed a diet containing 30% of FM replaced by SBM. However, these parameters were improved by the substitution of FSBM. It is therefore believed that FSBM has great potential to be used as a protein source in grouper diets in partial replacement of FM.     

Probiotic applications for rainbow trout (Oncorhynchus mykiss Walbaum) I. Effects on growth performance,feed utilization,intestinal microbiota and related health criteria

The effect of dietary probiotics (Bacillus subtilis, Bacillus licheniformis and Enterococcus faecium) used singularly and synergistically on the growth performance, intestinal microbiota and health status of rainbow trout (Oncorhynchus mykiss Walbaum) were assessed after 10 weeks feeding on supplemented diets. No significant improvements of weight gain or specific growth rate were observed in the probiotic fed groups. However, a significant improvement of feed conversion ratio was observed in the group fed E. faecium. High levels of probiotic species were observed in the posterior gastrointestinal tract as transient digesta‐associated populations and potentially resident mucosal populations. Bacillus subtilis and B. licheniformis levels accounted for 36% of the total culturable microbial population adhered to the mucosa and 62% in the digesta. E. faecium levels accounted for 45% of the mucosal population and 89% of the population in the digesta. An increase of serum lysozyme activity was observed in the fish fed diets containing the Bacillus probionts and elevated leukocyte levels were observed in fish fed diets containing Bacillus + E. faecium synergistically. The results of the current study demonstrate potential for B. subtilis, B. licheniformis and E. faecium to improve feed utilization, modulate intestinal microbiota and the health status of rainbow trout.     

Using a fermented mixture of soybean meal and earthworm meal to replace fish meal in the diet of white shrimp,Penaeus vannamei (Boone)

An experiment was conducted to evaluate the potential of a Bacillus subtilis E20‐fermented mixture (FSFEM) containing soybean meal (SBM) and Eisenia fetida earthworm meal (EM) at a ratio of 4:1 to increase the methionine level in order to satisfy the methionine requirement of white shrimp, Penaeus vannamei in a diet with fish meal (FM) completely replaced by mixtures. B. subtilis E20 fermentation improved the mixture's palatability and utilization based on better growth performance in comparison to shrimp fed FSEM (contains fermented SBM and EM at a ratio of 4:1) diets. FSFEM is a good substitute for FM. Maximal replacement levels of FM with FSFEM were 80% in a shrimp diet with 37% of crude protein and 7% of crude lipid based on weight gain and 100% based on feeding efficiency. In addition, shrimp fed experimental diets had no significant differences in survival after being challenged by Vibrio alginolyticus. It is suggested that B. subtilis E20‐FSFEM has the potential to replace FM in cultured shrimp diets.     

Dietary administration of Bacillus (B. licheniformis and B. subtilis) and isomaltooligosaccharide influences the intestinal microflora,immunological parameters and resistance against Vibrio alginolyticus in shrimp,Penaeus japonicus (Decapoda: Penaeidae)

The experiment was conducted to investigate the effects of oral administration of probiotics (Bacillus licheniformis and Bacillus subtilis) in combination with prebiotic isomaltooligosaccharide (IMO) on the intestinal microflora and immunological parameters of Penaeus japonicus and its resistence against Vibrio alginolyticus. A basal diet was supplemented with 0% probiotics and prebiotic (control), 10⁸ colony forming unit (CFU) g^?1B. licheniformis and 0.2% IMO (T1), 10⁸ CFU g^?1B. subtilis and 0.2% IMO (T2), 10⁸ CFU g^?1B. licheniformis in combination with 10⁸ CFU g^?1B. subtilis and 0.2% IMO (T3). The results showed that the total bacterial counts significantly increased (P<0.05) and Vibrio counts significantly decreased (P<0.05) in the intestine of shrimp with supplementation of dietary synbiotics. Shrimp fed the diet with both Bacillus probiotics and IMO (T3) produced significantly higher immune parameters (phenoloxidase activity, lysozyme activity and nitric oxide synthase activity and superoxide dismutase activity) than the control group (P<0.05). Significant lower cumulative mortality after challenge with V. alginolyticus was also observed in shrimp fed diet with both Bacillus probiotics and IMO (T3) than that in the control group (P<0.05). The results indicated that the oral administration of probiotics (B. licheniformis and B. subtilis) and prebiotic IMO has positive effects on bacterial flora in P. japonicus gut, and can activate non‐specific immunity of shrimp.     

Evaluation of shrimp or tilapia protein hydrolysate at graded dosages in low fish meal diet for olive flounder (Paralichthys olivaceus)

A study was carried out to examine and optimize the inclusion levels of shrimp hydrolysate (SH) or tilapia hydrolysate (TH) in low fishmeal diets for olive flounder (Paralichthys olivaceus). A fishmeal (FM)‐based diet was considered as a high FM (HFM) diet, and a diet containing soy protein concentrate (SPC) as a FM replacer at 50% substitution level was regarded as a low FM (LFM) diet. Six other experimental diets were prepared by dietary supplementation of SH or TH to LFM diet at different inclusion levels of 15 g/kg, 30 or 45 g/kg in the expense of FM (designated as SH‐1.5, SH‐3.0, SH‐4.5, TH‐1.5, TH‐3.0 and TH‐4.5, respectively). After 10 weeks of a feeding trial, growth performance and feed utilization efficiency of fish were significantly higher in fish fed HFM, SH‐3.0, SH‐4.5, TH‐1.5 and TH‐3.0 diets compared to those of fish fed LFM diet. Intestine diameter, villus height and goblet cell counts of fish were significantly increased by dietary inclusion of SH or TH into LFM diet. Dry matter and protein digestibility of diets were significantly improved by SH or TH incorporation. Innate immunity of fish was significantly enhanced by dietary SH or TH supplementation into LFM diet. Disease resistance of fish was significantly increased against Edwardsiella tarda by dietary inclusion of SH and TH at the highest inclusion level (45 g/kg). The optimum inclusion level of SH or TH in a SPC‐based LFM diet could be ~30 g/kg and 15–30 g/kg, for olive flounder.     

Probiotic applications for rainbow trout (Oncorhynchus mykiss Walbaum) II. Effects on growth performance,feed utilization,intestinal microbiota and related health criteria postantibiotic treatment

The effect of dietary probiotics (Bacillus subtilis, Bacillus licheniformis and Enterococcus faecium) was assessed on rainbow trout (Oncorhynchus mykiss Walbaum) previously treated with oxolinic acid. After feeding on supplemented diets for 10 weeks growth performance, feed utilization, gastrointestinal colonization and health status were assessed. B. subtilis + B. licheniformis fed fish displayed a significant improvement of feed conversation ration (FCR), specific growth rate (SGR) and protein efficiency ratio (PER). High levels of probiotic species were observed in the posterior gastrointestinal tract as transient digesta associated populations and potentially resident mucosal populations. Levels of Bacillus spp. reached log 3.74 CFU g^?1 on the mucosal epithelium and log 7.41 CFU g^?1 in the digesta of fish fed diets supplemented with B. subtilis and B. licheniformis. Enterococci levels reached log 2.84 CFU g^?1 on the mucosa and log 7.78 CFU g^?1 in the digesta of fish fed E. faecium supplemented diets. Feeding trout the Bacillus probionts alone or synergistically with E. faecium resulted in elevated leucocyte levels. The results of the current study demonstrate a potential role of probiotics for stabilizing/reinforcing the gastrointestinal microbiota after antibiotic treatment. This could reinvigorate the intestinal defensive barrier mechanism and provide protection against secondary potential pathogens.     

Effects of the Dietary Fermented Tuna By‐product Meal on Growth,Blood Parameters,Nonspecific Immune Response,and Disease Resistance in Juvenile Olive Flounder,Paralichthys olivaceus

This study evaluated the effects of dietary fermented tuna by‐product meal (FTBM) in juvenile olive flounder, Paralichthys olivaceus. Five diets were formulated to replace fishmeal (FM) with FTBM at 0% (FTBM₀), 12.5% (FTBM_12.5), 25.0% (FTBM₂₅), 37.5% (FTBM_37.5), or 50% (FTBM₅₀). After 8 wk, weight gain, specific growth rate, and feed efficiency of fish fed FTBM₀ and FTBM_12.5 diets were significantly higher than fish fed the other diets (P < 0.05). Also, mean cumulative survival rates (%) of fish fed the FTBM₀ and FTBM_12.5 diets were significantly higher than those fed FTBM₅₀ diet at Day 9 postchallenge with Edwardsiella tarda (P < 0.05). Protein efficiency ratio of fish fed FTBM₀ and FTBM_12.5 diets was significantly higher (P < 0.05) than fish fed diets FTBM_37.5 and FTBM₅₀. Broken‐line regression analysis of weight gain showed an optimal FM replacement level of 10.65% with FTBM. Therefore, the optimal dietary inclusion of FTBM in juvenile olive flounder diets could be greater than 10.65% but less than 12.5% without any adverse physiological effects on fish health.     

Effects of two dietary probiotics (Bacillus subtilis or licheniformis) with two prebiotics (mannan or fructo oligosaccharide) in Japanese eel,Anguilla japonica

A 12‐week feeding trial was conducted to investigate the effects of two dietary probiotics; Bacillus subtilis KCTC 2217 or Bacillus licheniformis KCCM 11775 with two prebiotics; mannan oligosaccharide (MOS) or fructooligosaccharide (FOS) in Japanese eel. Fish averaging 12.8 ± 0.47 g (mean ± SD) were randomly distributed into five treatments with triplicate tanks. A basal control diet (CON) and four synbiotic diets supplementing B. subtilis + MOS (BSM), B. subtilis + FOS (BSF), B. licheniformis + MOS (BLM), and B. licheniformis + FOS (BLF). Weight gain and specific growth rate of fish fed all synbiotic diets were higher than those of fish fed CON. Immune‐related gene expression of heat shock protein 70 and immunoglobulin M of fish fed BSF and BLM were significantly higher than those of fish fed CON. Fish fed BSF and BLM had significantly higher intestinal villi length than those of fish fed BLF and CON (p < .05). Disease resistance against Aeromonas hydrophila of fish fed all synbiotic diets were significantly higher than those of fish fed CON (p < .05). Therefore, these results indicated that dietary B. subtilis with FOS (BSF) and B. licheniformis with MOS (BLM) could have beneficial effects on intestinal morphology, and immune‐related gene expression in Japanese eel.     

Supplemental iron and phosphorus increase dietary inclusion of cottonseed and soybean meal in olive flounder (Paralichthys olivaceus)

A long‐term feeding experiment was conducted to investigate the use of cottonseed and soybean meal (CS) with iron and phosphorus supplements in diets for olive flounder (Paralichthys olivaceus). Olive flounder with an initial average size of 28.5 ± 0.35 g (mean ± SD) were divided into 15 groups (three tanks per dietary treatment) and fed 480 g kg^?1 crude protein diets in which each of five isonitrogenous diets was formulated to contain different levels of cottonseed/soybean meal (1 : 1) to replace fish meal (FM) with iron and phosphorus supplementations. The five experimental diets were as follows: diet 1 (control), 0%CS; diet 2, 20%CS; diet 3, 30%CS; diet 4, 30%CS + Fe&P; and diet 5, 40%CS + Fe&P. After 26 weeks of feeding trial, no significant differences were observed in weight gain, feed utilization and survival among all the treatments. The total gossypol accumulation in liver of fish fed diets supplemented with iron was significantly lower than that of fish fed diets without supplementation of iron. The results indicate that the addition of iron in diets could prevent the absorption of free gossypol. The findings in this study suggest that dietary supplements of iron and phosphorus could increase the inclusion of cottonseed and soybean meal for FM replacement in diets for marine fish species.     

Evaluation of Antarctic krill (Euphausia superba) meal supplementation in diets for olive flounder (Paralichthys olivaceus)

The supplemental effect of Antarctic krill meal (KM) into a low fish meal (FM) diet was evaluated for olive flounder (Paralichthys olivaceus). A 56% FM‐based diet was regarded as a high FM inclusion diet (HFM), and a low‐FM diet (LFM) was prepared by replacing 50% FM from the HFM. Four other diets were prepared by supplementing 3%, 6%, 9% and 12% KM into the LFM diet gradually replacing soy protein concentrate and tankage meal (designated as KM3, KM6, KM9 and KM12 respectively). Quadruplicate groups of fish were fed one of the diets for 12 weeks. The growth performance and feed utilization efficiency were improved by the dietary KM supplementation. Digestibility of dietary protein and dry matter was increased by the KM3‐9 diets. Haematocrit and haemoglobin were increased by KM supplementation. The innate immunity and antioxidant capacity assessed by Ig, antiprotease, lysozyme, GPx and SOD and the condition factor of fish were significantly increased by KM3‐9 diets. Moreover, goblet cell counts, villi length and fillet yield of fish were significantly improved by all the KM‐containing diets (KM3‐12). A 25‐day‐long challenge test with the Edwardsiella tarda pathogen showed that the cumulative mortality was higher in fish fed the LFM diet than in fish fed the HFM or KM‐supplemented diets. The results indicate that dietary KM supplementation in a LFM diet can increase growth performance and feed utilization efficiency, diet digestibility, intestinal development and functions, innate immunity and disease resistance of olive flounder. The recommended level of KM inclusion in a LFM diet seems to be 6.6% according to quadratic regression analysis.     

Preliminary evaluation of growth performance and immune response of Nile tilapia Oreochromis niloticus supplemented with two putative probiotic bacteria

This study evaluated the effect of two bacteria, Bacillus amyloliquefaciens and the dairy yogurt (DY) Lactobacillus sp., on improving the growth performance, feed conversion ratio (FCR) and some immunological and haematological parameters. Nile tilapia (19.1 g) were stocked in 0.42 m³ tanks at 67 fish m^?3 and fed with two probiotic‐incorporated diets and a control diet for 99 days, followed by a normal diet for another 61 days. At the end of the probiotic feeding, the mean weight, FCR and production rate showed no significant difference among the treatments. However, after 61 days of feeding of the normal diet, the B. amyloliquefaciens fish showed significantly superior growth and better FCR than the control. The DY group had the lowest growth and the poorest FCR. Significantly higher serum lysozyme activity, head‐kidney superoxide dismutase, total immunoglobulin and serum bacterial agglutination titres were recorded in the probiotic groups than the control. Haematological parameters showed no difference between treatments. The B. amyloliquefaciens fed fish showed that the gut microflora was dominated by B. amyloliquefaciens even after the withdrawal of the probiotic. On the contrary, the probiotic bacterial species isolated from the dairy yoghurt did not persist longer in the gut. These results indicate the beneficial effect of administering the bacteria isolated from fish gut microflora, B. amyloliquefaciens, in improving growth, FCR and immunological parameters. The high persistence of B. amyloliquefaciens indicates that the probiotic colonization in the gut is essential for the best responses and economics.     

Expression of insulin-like growth factor genes in olive flounder,Paralichthys olivaceus,fed a diet with partial replacement of dietary fish meal

This study analyzed the expression of growth-related genes of olive flounder, Paralichthys olivaceus, fed a low-fish meal (FM) diet to investigate the replacement of the FM diet in the field. The expression of growth hormone (GH)/insulin-like growth factor (IGF) axis genes in the brain/pituitary/liver and plasma hormone concentrations were measured. A basal experimental diet was formulated using 63% FM and 12% soybean meal as the primary protein sources, and then two other diets were made by replacing 20% and 30% of the FM with soy protein concentrate, tankage meal, and poultry by-products meal. Each diet was fed to duplicate groups of juvenile flounder (150 ± 3.0 g) twice a day. After 20 weeks, the plasma growth-related hormone concentrations were similar between the control and FM20 groups. Moreover, GH/IGF axis gene mRNA expression in the brain/pituitary/liver was similar between the control and FM20 groups. Immunoblotting of muscle and liver showed the same pattern. This study showed the utility of replacing 20% of the FM diet and supports the possibility of field application.     

Effects of dietary supplementation of citrus by‐products fermented with a probiotic microbe on growth performance,innate immunity and disease resistance against Edwardsiella tarda in juvenile olive flounder,Paralichthys olivaceus (Temminck & Schlegel)

Two consecutive studies were conducted to evaluate the dietary supplementation of citrus by‐products (CB) fermented with probiotic bacteria on growth performance, feed utilization, innate immune responses and disease resistance of juvenile olive flounder. In Experiment I, five diets were formulated to contain 0% (control) or 3% four different CB fermented with Bacillus subtilis (BS), Enterococcus faecium (EF), Lactobacillus rhamnosus (LR) and L. plantarum (LP) (designated as CON, CBF‐BS, CBF‐EF, CBF‐LR and CBF‐LP, respectively). During 10 weeks of a feeding trial, growth performance and feed efficiency were not significantly different among all the fish groups. However, fish fed CBF containing diets had significantly higher survivals than the CON group. Disease resistance of fish against Edwardsiella tarda was increased by the fermentation of CB. In Experiment II, we chose the BS as a promising probiotic and formulated five diets to contain 0%, 2%, 4%, 6% and 8% CBF‐BS. Growth performance was not significantly affected by the CBF‐BS supplementation during 6 weeks of a feeding trial. Innate immunity of fish was significantly enhanced by CBF‐BS supplementation. Myeloperoxidase and lysozyme activities were increased in a dose‐dependent manner by dietary CBF‐BS inclusions. In a consecutive challenge test against E. tarda, an increased disease resistance was found by CBF‐BS supplementation. These studies indicate that the fermentation process of CB with probiotic has beneficial effects on innate immunity and thereby increases disease resistance of olive flounder against E. tarda. Bacillus subtilis can be used as a promising probiotic microbe for by‐product fermentation in fish feeds.     

Evaluation of fermented fish‐offal in the formulated diet of the freshwater catfish Heteropneustes fossilis

A 60‐day feeding trial was conducted to test the effect of partial replacement of fishmeal by fish‐offal (FO) in the diet for the freshwater catfish Heteropneustes fossilis. Three isonitrogenous (31.4% CP) diets were formulated to include a reference diet (T1) with 40% fishmeal (FM) and 0% FO and two supplementary diets: one (T2) containing 25% FM and 25% FO and another (T3) containing 20% FM and 30% FO. The FO was fermented along with mustard oil cake and rice bran before using it as an ingredient in the preparation of feed. Two separate trials were conducted with these three diets: a growth trial and a digestibility trial. H. fossilis fed the diets containing FO showed better growth and proximate composition of carcass than those fed the reference diet. Fish fed T3 diet showed maximum feed conversion, protein utilization and growth. Apparent protein digestibility (APD) was also significantly higher in the T3 diet as compared with the T1 diet. The results of the trial indicated that using microbial fermentation, FO could be included up to a 30% level as a partial (50%) replacement of fishmeal in the formulation of diet for H. fossilis.     

Evaluation of carbohydrate‐to‐lipid ratio in diets supplemented with Bacillus subtilis probiotic strain on growth performance,body composition and digestibility in juvenile white seabass (Atractoscion nobilis,Ayres 1860)

This study aimed to evaluate the effect of increasing the carbohydrate to lipid ratio in diets supplemented with Bacillus subtilis in white seabass (Atractoscion nobilis) juveniles on digestibility, growth performance, feed utilization efficiency and tissue composition. Four experimental isoenergetic and isoproteic diets supplemented with B. subtilis were formulated with increasing starch to lipid ratios (g/g) of 10:19; 14:17; 18:15 and 22:13 (diets PD10, PD14, PD18 and PD22 respectively). A negative control diet for B. subtilis supplementation was also formulated to be similar to diet PD10 (diet CD10). Each experimental diet was fed to triplicate groups of 20 fish per tank (IBW: 9.5 g), three times a day, to apparent satiation, for 60 days. Supplementation with B. subtilis, of the low starch‐high lipid diet (10:19), significantly enhanced the overall growth performance, digestibility and feed efficiency. Moreover, the increase in starch to lipid ratio, in supplemented probiotic diets, significantly increased growth performance, feed intake, feed utilization efficiency and protein and energy digestibility, being maximum for the highest dietary starch to lipid level. Lipid content of whole body, muscle and liver and hepatosomatic index significantly increased with the increase in starch to lipid ratio. The results of this study indicate that B. subtilis supplementation significantly enhanced growth performance, digestibility and feed utilization of low starch‐high lipid diet. However, further improvement of growth performance and feed utilization efficiency may be achieved combining B. subtilis supplementation with increased dietary starch to lipid ratio to 22:13.     

Single or combined effects of fructo‐ and mannan oligosaccharide supplements and Bacillus clausii on the growth,feed utilization,body composition,digestive enzyme activity,innate immune response and lipid metabolism of the Japanese flounder Paralichthys olivaceus

In this study, we examined the effects of the following eight experimental diets, which varied in fructo oligosaccharides (FOS), mannan oligosaccharides (MOS) and Bacillus clausii concentrations, on the Japanese flounder: control diet (no FOS, MOS and B. clausii), diet F (5 g kg⁻¹ FOS), diet M (5 g kg⁻¹ MOS), diet FM (2.5 g kg⁻¹ FOS + 2.5 g kg⁻¹ MOS), diet B (10⁷ cells g⁻¹B. clausii), diet FB (5 g kg⁻¹ FOS + 10⁷ cells g⁻¹B. clausii), diet MB (5 g kg⁻¹ MOS + 10⁷ cells g⁻¹B. clausii) and diet FMB (2.5 g kg⁻¹ FOS + 2.5 g kg⁻¹ MOS + 10⁷ cells g⁻¹B. clausii). Japanese flounder, initially weighing an average of 21 g, were distributed into 24 net cages at a stocking density of 20 fish per cage. Each diet was hand‐fed to three groups of fish twice daily for 56 days. The weight gain rate (WGR) in fish fed diets B, MB and FMB were significantly higher than in fish fed the control diet, where the fish fed diet FMB had the highest WGR. Fish fed any of the diets, except diets F and B, exhibited better feed conversion ratio than those fed the control diet. Diets MB and FMB significantly elevated intestinal protease activity compared with the control diet, but only the diet FMB promoted amylase activity. Feeding diets FB and FMB increased body protein deposition; additionally, feeding diets B, MB and FMB significantly reduced body lipid deposition. Lysozyme (LSZ) activity was significantly higher in fish fed diets B, FB, MB and FMB than in fish fed the control diet. All diets, except diet M, decreased triglyceride (TG) levels compared to the control diet. Low‐density lipoprotein cholesterol levels in fish fed diets F, FB and FMB were significantly lower than in fish fed the control diet. Without exception, no diets affected feeding rate, condition factor, body moisture, ash contents, phagocytic activity of leucocytes or cholesterol or high‐density lipoprotein cholesterol levels. Our results suggest that diets supplemented with FOS, MOS and B. clausii improved growth performance and health benefits of the Japanese flounder more than other diets or the control diet.