Identification and expression profiles of multiple genes in Nile tilapia in response to bacterial infections

To understand the molecular mechanisms involved in response of Nile tilapia (Oreochromis niloticus) to bacterial infection, suppression subtractive cDNA hybridization technique was used to identify upregulated genes in the posterior kidney of Nile tilapia at 6h post infection with Aeromonas hydrophila. A total of 31 unique expressed sequence tags (ESTs) were identified from 192 clones of the subtractive cDNA library. Quantitative PCR revealed that nine of the 31 ESTs were significantly (p<0.05) upregulated in Nile tilapia at 6h post infection with A. hydrophila at an injection dose of 10(5)CFU per fish (≈ 20% mortality). Of the nine upregulated genes, four were also significantly (p<0.05) induced in Nile tilapia at 6h post infection with A. hydrophila at an injection dose of 10(6)CFU per fish (≈ 60% mortality). Of the four genes induced by A. hydrophila at both injection doses, three were also significantly (p<0.05) upregulated in Nile tilapia at 6h post infection with Streptococcus iniae at doses of 10(6) and at 10(5)CFU per fish (≈ 70% and ≈ 30% mortality, respectively). The three genes induced by both bacteria included EST 2A05 (similar to adenylate kinase domain containing protein 1), EST 2G11 (unknown protein, shared similarity with Salmo salar IgH locus B genomic sequence with e value of 0.02), and EST 2H04 (unknown protein). Significant upregulation of these genes in Nile tilapia following bacterial infections suggested that they might play important roles in host response to infections of A. hydrophila and S. iniae.     

Thiamin Requirement of Nile Tilapia,Oreochromis niloticus

Juvenile Nile tilapia, Oreochromis niloticus, were fed to apparent satiation twice daily with purified diets containing 0, 1.0, 2.0, 4.0, 8.0, and 16.0 mg/kg and 0, 2.0, 4.0, 8.0, 16.0, and 32.0 mg/kg of thiamin in separate 14‐ and 8‐wk trials (Experiments 1 and 2, respectively). Fish fed the diet devoid of thiamin developed neurological disorders, anorexia, reduced growth, and feed efficiency and increased mortality (Experiment 2 only) within 4–6 and 8–10 wk for Experiments 2 and 1, respectively. Low red blood cell count (RBC) and hematocrit (Ht) were observed in fish fed the thiamin‐deficient diet. Serum pyruvate was elevated in fish fed the thiamin unsupplemented diet. Serum lactate was not affected by dietary thiamin levels. Whole body protein was unaffected by dietary levels of thiamin. Body moisture and ash increased whereas body lipid decreased in fish fed the thiamin unsupplemented diets. None of these abnormalities were observed in fish fed the thiamin‐supplemented diets. Using the response curves determined by PROC NLMIXED to estimate dietary thiamin levels required for various response variables, a dietary thiamin level of 3.5 mg/kg diet was adequate for optimum growth, feed intake and efficiency, survival, prevention of neurological symptoms, and maintaining normal levels of RBC, Ht, serum pyruvate, and proximate body composition.     

Effects of dietary supplementation of a purified nucleotide mixture on immune function and disease and stress resistance in channel catfish,Ictalurus punctatus

Juvenile channel catfish (14.4 g average initial weight) were fed diets supplemented with a purified nucleotide mixture for 8 weeks. The mixture consisted of five nucleotides supplied on an equal basis as disodium salts at combined concentrations of 0 (control), 0.1%, 0.3%, 0.9% or 2.7% of diet. Addition of nucleotides to diet produced a dose‐dependent reduction in survival of channel catfish to Edwardsiella ictaluri. Although the reasons are unclear, the high levels of nucleotides supplemented in this study may have contributed to the decrease in disease resistance, and this decrease appeared to be unrelated to the innate immune (unaffected) and specific antibody (enhanced) responses. Stress resistance increased with a corresponding amelioration of the immunosuppressive effects of the stress response on non‐specific immunity (lysozyme and bactericidal activity) as nucleotides increased in diet. Use of exogenous nucleotides as a prophylactic treatment before culture‐related stress exposure may prove beneficial by decreasing the immunosuppressive effects of stress but not in prevention of ESC.     

Growth performance,vitamin E status,and proximate and fatty acid composition of channel catfish, <Emphasis Type="Italic">Ictalurus punctatus</Emphasis>, fed diets containing various levels of fish oil and vitamin E

A study was conducted to determine the effect of increasing dietary levels of fish oil on vitamin E requirement and their effect on growth performance, liver vitamin E status, and tissue proximate and fatty acid compositions of channel catfish. Basal purified diets (42% protein and 3,800 kcal DE/kg) supplemented with 6, 10, and 14% menhaden fish oil were each supplemented with 50, 100, and 200 mg vitamin E/kg (3 × 3 factorial experiment). Each diet was fed to juvenile channel catfish in three random aquaria to apparent satiation twice daily for 12 weeks. Weight gain, feed intake, and feed efficiency ratio were not affected by dietary levels of fish oil, vitamin E, or their interaction. Survival rate at the end of week 12 was significantly lower for fish fed diets containing 14% fish oil, regardless of vitamin E content. Whole-body moisture significantly decreased and lipid increased when dietary lipid levels were increased to 10 or 14%. Dietary vitamin E levels had no effect on body proximate composition. Lipid content of liver was not influenced by dietary levels of fish oil and vitamin E or their interaction. Hepatosomatic index significantly decreased with increasing lipid levels but was not affected by dietary levels of vitamin E. Liver vitamin E increased with increasing dietary vitamin E but decreased with increasing fish oil levels. Fatty acid composition of whole body and liver reflected that of dietary lipid but was not influenced by dietary levels of vitamin E. Whole-body saturates increased, whereas MUFA decreased with increasing dietary levels of fish oil. Liver saturates were not affected by fish oil levels, but MUFA and n-6 decreased and increased, respectively, with increasing fish oil levels. Total n-3 and n-3 HUFA in both tissues increased with increasing fish oil levels in diets, but liver stored much higher levels of these fatty acids.     

Susceptibility of Nile tilapia (Oreochromis niloticus) fed with dietary sodium chloride to nitrite toxicity

Juvenile Nile tilapia (Oreochromis niloticus) were fed with diets supplemented with 0 or 6% NaCl for 10 weeks. Tilapia were exposed to approximately 21 mg/l nitrite-N after 5 and 10 weeks of feeding to determine the effect of dietary NaCl supplementation on resistance to nitrite toxicity. Fish were sampled before (baseline, pre-exposure) and after 24 h nitrite exposure to determine the effects of dietary NaCl on mortality, hematology (hematocrit, hemoglobin, and methemoglobin), and plasma electrolyte dynamics (nitrite, chloride, sodium, and potassium). After 10 weeks of feeding, tilapia were also challenged with Streptococcus iniae to determine the effect of sodium chloride on immunity. Tilapia fed with the NaCl-supplemented diet had significantly higher weight gain compared with the control group, which was associated with a significant increase in feed intake in the NaCl group. Mortality from nitrite exposure was lower in tilapia fed with the NaCl-supplemented diet compared with the control group at 5 and 10 weeks, but the differences were not significant. However, dietary NaCl supplementation caused a significant decrease in plasma nitrite levels after nitrite exposure. The dietary reduction in nitrite may be related to the increase in plasma chloride in the 6% NaCl-supplemented fish. A direct link between the effects of dietary NaCl supplementation on methemoglobin (MetHb) could not be established. Tilapia in this study were subjected to acute nitrite toxicity. Dietary sodium chloride may be more effective in protecting against nitrite toxicity at lower levels of nitrite, but the conditions at which it proves to be effective may be limited and requires further investigation. Feeding NaCl to tilapia did not affect susceptibility to S. iniae or immune function, but nitrite exposure cause a stress-related reduction in non-specific immune function. This is the first study to examine the effects of dietary salt on nitrite toxicity in tilapia.     

Antimicrobial activity of the biopolymer chitosan against Streptococcus iniae

The antimicrobial activity and mode of action of chitosan were evaluated against Streptococcus iniae, a pathogenic Gram‐positive bacterium of fish worldwide. Cell proliferation kinetics were examined following exposure to varying concentrations of chitosan. The action of chitosan on S. iniae was also investigated by measuring agglutination activity, conductivity, and extracellular and intracellular bacterial adenosine triphosphate (ATP) levels. Chitosan exhibited antibacterial activity against S. iniae at concentrations of 0.1% and above and was lethal at a concentration of 0.4% and higher. The mechanism of antibacterial activity of chitosan at the inhibitory level of bacterial growth appears to hinge upon the interaction between chitosan and the oppositely charged bacterial surface. This interplay causes agglutination, which was readily observed grossly and microscopically. After interacting with the cell surface via adsorption, an efflux of intracellular ATP was documented, which suggests that chitosan disrupts the bacterial cell causing leakage of cytosolic contents and ultimately cell death. Results suggest chitosan may be worth evaluating as a natural alternative to antibiotic against S. iniae infection of fish.     

Growth Response and Resistance to Edwardsiella ictaluri of Channel Catfish,Ictalurus punctatus,Fed Diets Containing Distiller’s Dried Grains with Solubles

A study was conducted to examine the effect of dietary levels of distiller’s dried grains with solubles (DDGS) on growth, body composition, hematology, immune response, and resistance of channel catfish, Ictalurus punctatus, to Edwardsiella ictaluri challenge. Five diets containing 0, 10, 20, 30, and 40% DDGS with supplemental lysine (Diets 1–5) as partial replacements of a combination of soybean meal and cornmeal on an equal protein basis were fed to juvenile catfish (13.33 ± 0.25 g) for 12 wk. Growth performance and feed utilization efficiency were similar for fish in all treatments. Body lipid and moisture increased and decreased, respectively, in fish feed DDGS‐containing diets relative to the control group. Dietary treatment had no effect on red and white blood cell counts. Hemoglobin and hematocrit were significantly higher in fish fed diets containing DDGS than in those fed the control diet. Fish fed 20–40% DDGS diets had increased serum total immunoglobulin, and those fed the 30% DDGS diet had significantly increased antibody titers 21 d following E. ictaluri challenge. Other immune variables evaluated were not affected by dietary treatments. Preliminary results on bacterial challenge showed an increased resistance against E. ictaluri in fish fed DDGS‐containing diets (Diets 2–5).     

Interaction between dietary levels of vitamins C and E on growth and immune responses in channel catfish, Ictalurus punctatus (Rafinesque)

This study was conducted to evaluate the effect of dietary levels of vitamins C (0, 100 and 2000 mg kg⁻¹), E (0, 50 and 500 mg kg⁻¹) and their interaction on the growth performance, liver contents of ascorbic acid and α-tocopherol, haematology and immune response of channel catfish, Ictalurus punctatus . Each diet was fed to catfish in triplicate aquaria to apparent satiation twice daily for 12 weeks. The results indicate that the amount of vitamin E contained in the basal diet (23.1 mg kg⁻¹) was sufficient to promote good growth, feed efficiency and survival, but its supplementation was needed to maintain high haematological values and liver vitamin E. Supplementation of vitamin C (100 mg kg⁻¹) to the basal diet containing 10.5 mg kg⁻¹ was required for good growth, feed efficiency, survival and prevention of vertebral deformity and optimum haematological indices. Liver storage of ascorbic acid and α-tocopherol increased with increasing dietary levels of each vitamin. Dietary vitamin E levels had no effect on liver ascorbic acid content, but increasing dietary vitamin C increased liver α-tocopherol. Some measured immune parameters (serum protein and superoxide anion production) were enhanced by supplementation of vitamin C or E. Chemotaxis ratio and phagocytosis were not affected by treatments.     

Nutritional value of frass from black soldier fly larvae,Hermetia illucens,in a channel catfish,Ictalurus punctatus,diet

Frass is the by‐product of the larval meal industry and includes larval waste, exoskeleton sheds and residual feed ingredients. Experimental frass was derived from the larvae of black solder flies fed distillers' dried grains with solubles and had a protein and fat content of 216 and 60 g/kg, respectively. A 10‐week study was conducted to evaluate the effect of dietary levels of frass on growth, feed utilization, and body proximate and mineral composition of channel catfish, Ictalurus punctatus. Five diets containing 0, 50, 100, 200 and 300 g frass per kg diet were fed to channel catfish (5.24 ± 0.04 g) in quadruplicate aquaria to apparent satiation twice daily. Final weight gain was significantly increased in fish fed diets containing frass at levels from 100 to 300 g/kg. Fish fed diets without frass, and with 300 g/kg frass, showed the lowest and highest feed intake, respectively. Feed and protein efficiencies, however, were significantly lower in fish fed frass at levels of 200 g/kg and higher compared to the control diet. Survival, whole‐body composition and mineral content were not affected by frass. In summary, black soldier fly larval frass has potential as a protein source or just an ingredient for enhancing palatability of catfish diets.     

Immune Response and Resistance to Stress and Edwardsiella ictaluri Challenge in Channel Catfish, Ictalurus punctatus, Fed Diets Containing Commercial Whole-Cell Yeast or Yeast Subcomponents

Dietary supplementation of yeast or yeast subcomponents (YYS) as commercial preparations of β‐glucan (MacroGard^®; Biotec‐Mackzymal, Tromsø, Norway; and Betagard A^®; Aqua‐In‐Tech, Inc., Seattle, WA, USA), mannan oligosaccharide (Bio‐Mos? Aqua Grade; Alltech, Nicholasville, KY, USA), or whole‐cell Saccharomyces cerevisiae (Levucell SB20^®; Lallemand Animal Nutrition, Milwaukee, WI, USA) at the manufacturer’s recommended levels was evaluated on the physiological performance of juvenile channel catfish, Ictalurus punctatus. Fish were fed YYS diets for 4 wk, followed by 2 wk of control diet. Fish were sampled at the end of each feeding period (4 and 6 wk) to measure hematological and immune parameters and growth and to determine the effects of dietary β‐glucan on resistance to Edwardsiella ictaluri infection and to low‐water stress (6 wk). Supplementation of YYS in diets did not affect growth performance, hematology, or immune function. Survival from E. ictaluri infection was from 5 to 17.5% higher in fish fed YYS diets than in the control group, but the increases were not significant. Some improvement in stress resistance was observed in YYS‐fed catfish after exposure to low‐water stress. Stress reduction in fish fed diets supplemented with yeast subcomponents has been reported previously, but thus far, no explanation has been proposed for this effect. The present study and the previously published research suggest that dietary YYS supplementation does not appear to improve resistance of channel catfish to E. ictaluri.