1. Growth Response and Resistance to Streptococcus iniae of Nile Tilapia Oreochromis nilotucus Fed Diets Containing Distiller’s Dried Grains with Solubles Chhorn Lim *, Julio C. Garcia, Mediha Yildirim-Aksoy and Phillip H. Klesius and Phillip H. Klesius USDA-ARS, MSA Aquatic Animal health Research Laboratory P.O. Box 952 Auburn, Alabama 36831 (USA)

2. OBJECTIVES To evaluate the effect of dietary levels of distiller’s dried grains with solubles (DDGS) on: Growth performance Growth performance Body composition Body composition Hematology Hematology Immune response Immune response and resistance of Nile tilapia to Streptococcus iniae challenge and resistance of Nile tilapia to Streptococcus iniae challenge

3. METHODS

4. EXPERIMENTAL FISH Species: Nile tilapia (Oreochromis niloticus) Species: Nile tilapia (Oreochromis niloticus) Av. initial weight:9.41 ± 0.14 g Av. initial weight:9.41 ± 0.14 g Stocking rate: 30 fish/aquarium Stocking rate: 30 fish/aquarium No. of replicate: 4 rep./treatment No. of replicate: 4 rep./treatment No. of treatment:5 (CRD) No. of treatment:5 (CRD)

5. CULTURE FACILITIES AND WATER Tank: 57-L aquarium equipped with flow-through and aeration Tank: 57-L aquarium equipped with flow-through and aeration Water source:Heated, Dechlorinated water Water source:Heated, Dechlorinated water Water flow rate:0.6-1.0 L/min Water flow rate:0.6-1.0 L/min Water temp.: 27.1 ± 0.10  C Water temp.: 27.1 ± 0.10  C D.O.: 4.91 ± 0.07 mg/L D.O.: 4.91 ± 0.07 mg/L Tank cleaning:Once weekly Tank cleaning:Once weekly

6. COMPOSITION OF EXPERIMENTAL DIETS IngredientsExperimental diets (%) 12345 Menhaden fish meal88888 Soybean meal45403524.5 Corn meal3026.723.316.6 Wheat middlings55555 Distiller’s dried grains with solubles–102040 Corn Oil3.62.820.5 CMC33333 Dicalcium phosphate11111 Vitamin premix0.5 Mineral premix0.5 Lysine-HCL––––0.4 Celufil3.42.51.70.4–

7. DIETARY DDGS LEVELS USED AS REPLACEMENTS OF SOYBEAN MEAL AND CORN MEAL MIX Diets 1 2 3 4 5 DDGS (%) 0 10 20 40 40 + 0.4% lysine

8. PROXIMATE COMPOSITION OF EXPERIMENTAL DIETS Proximate composition (%)12Diets345 Moisture90.5590.891.390.9691.36 Protein30.8731.5832.233.1433.73 Lipid5.465.565.455.485.35 Ash6.845.876.856.86.94 Analyzed β-glucan (g/kg diet)< 3.0 3.43.6

9. FEED AND FEEDING Feed preparation:Hobart meat grinder Feed preparation:Hobart meat grinder Type of pellet:3-mm pellet (dried at R.T.) Type of pellet:3-mm pellet (dried at R.T.) Feed storage: Plastic bag kept at -20  C Feed storage: Plastic bag kept at -20  C Feeding schedule: 2 times daily to apparent satiation Feeding schedule: 2 times daily to apparent satiation Feeding duration: 10 weeks Feeding duration: 10 weeks

10. SAMPLING Weight gain: At 2-week intervals. Weight gain: At 2-week intervals. Body composition: 4 pooled whole fish/tank, 2 det./sample. Body composition: 4 pooled whole fish/tank, 2 det./sample. Hematology: Blood of 4 fish/tank, 2 det./fish. Hematology: Blood of 4 fish/tank, 2 det./fish. Immune responses: Serum of 4 fish/tank, 2 det./fish. Immune responses: Serum of 4 fish/tank, 2 det./fish. Bacterial challenge: 20 fish/tank by IP. injection 8.77x10 5 cfu/fish of S. iniae Bacterial challenge: 20 fish/tank by IP. injection 8.77x10 5 cfu/fish of S. iniae (wk 10). (wk 10). Antibody titer: 4 fish/tank prior to and 15 days after challenge. Antibody titer: 4 fish/tank prior to and 15 days after challenge.

11. STATISTICAL ANALYSES All data were analyzed by one-way analysis of variance using the General Linear Model (GLM) of SAS. All data were analyzed by one-way analysis of variance using the General Linear Model (GLM) of SAS. Duncan’s multiple range test was used to compare treatment means. Duncan’s multiple range test was used to compare treatment means. Differences were considered significant at the 0.05 probability level. Differences were considered significant at the 0.05 probability level.

12. RESULTS

13. GROWTH PERFORMANCE Levels of DDGS Weight gain Feed intakeFERPERSurvival (%) (g) (%) 0 48.0 ab 75.4 0.66 a 1.93 a 97.5 10 51.1 a 84.0 0.63 ab 1.82 ab 93.3 20 48.9 ab 74.7 0.65 ab 1.85 ab 93.2 40 41.1 c 73.5 0.58 b 1.59 c 98.2 40+0.4% lysine 44.7 bc 73.1 0.62 ab 1.70 bc 96.7

14. PROXIMATE BODY COMPOSITION Levels of MoisturePercent wet weight basis (%) DDGS (%) ProteinLipidAsh 069.016.9 a 6.634.58 1069.8 16.3 ab 6.74.47 2068.8 16.2 ab 7.45.58 4071.515.6 b 6.744.68 40+0.4% lysine71.8 16.1 ab 6.514.83

15. HEMATOLOGY Levels of DDGS RBCWBCHemoglobinHematocrit (%) (x10 6 /µl)(x10 5 /µl)(g/dl)(%) 02.043.399.4728.51 102.033.549.1125.57 201.903.569.1028.53 401.903.749.3427.44 40+0.4% lysine1.913.58.4127.00

16. IMMUNE RESPONSE Levels of DDGS Serum protein Lysozyme activity Antibody titer (%) (mg/ml)(µg/ml)(log 10 ) 031.617.151.20 1032.416.200.77 2032.316.961.32 4032.116.351.37 40+0.4% lysine30.716.471.05

17. BACTERIAL CHALLENGE Levels of DDGS Days to first Cumulative mortality (%) mortality(%) 02.0047.5 101.3338.3 202.0056.3 401.0050.0 40+0.4% lysine1.2542.5

18. SUMMARY Dietary DDGS levels had no effect on feed intake and survival. Dietary DDGS levels had no effect on feed intake and survival. WG, FER and PER of fish fed the diet containing the 40% DDGS-diet without lysine supplementation were significantly lower than those of the control. WG, FER and PER of fish fed the diet containing the 40% DDGS-diet without lysine supplementation were significantly lower than those of the control. Supplementation of lysine to the 40% DDGS-diet improved the growth and FER to levels comparable to those of the control. Supplementation of lysine to the 40% DDGS-diet improved the growth and FER to levels comparable to those of the control. Body protein content of fish fed the 40% DDGS diet without lysine supplementation was significantly lower than that of the control. Body protein content of fish fed the 40% DDGS diet without lysine supplementation was significantly lower than that of the control. Body moisture, lipid and ash were not significantly effected by dietary levels of DDGS Body moisture, lipid and ash were not significantly effected by dietary levels of DDGS

19. SUMMARY RBC, WBC, hemoglobin and hematocrit were not significantly effected by dietary levels of DDGS. RBC, WBC, hemoglobin and hematocrit were not significantly effected by dietary levels of DDGS. Serum protein, lysozyme activity and antibody titer against S. iniae were also not effected by dietary levels of DDGS. Serum protein, lysozyme activity and antibody titer against S. iniae were also not effected by dietary levels of DDGS. Dietary DDGS levels had no effect on number of the days at which first mortality occurred and cumulative mortality of fish 15 days post-challenge with S. iniae. Dietary DDGS levels had no effect on number of the days at which first mortality occurred and cumulative mortality of fish 15 days post-challenge with S. iniae.

20. CONCLUSION Our data indicate that at least 20% DDGS can be used in Nile tilapia diets as a replacement of a combination of soybean and corn meal on an equal protein basis. Our data indicate that at least 20% DDGS can be used in Nile tilapia diets as a replacement of a combination of soybean and corn meal on an equal protein basis. Increasing dietary levels of DDGS to 40% significantly decreased growth, and feed and protein efficiency ratios. Increasing dietary levels of DDGS to 40% significantly decreased growth, and feed and protein efficiency ratios. Nile tilapia appeared to be able to utilize supplemental lysine since fish fed the 40% DDGS-diet supplemented with lysine had improved weight gain and FER. Nile tilapia appeared to be able to utilize supplemental lysine since fish fed the 40% DDGS-diet supplemented with lysine had improved weight gain and FER. Lower body protein of fish fed the 40% DDGS without lysine supplementation could be related to the smaller size fish. Lower body protein of fish fed the 40% DDGS without lysine supplementation could be related to the smaller size fish. Dietary levels of DDGS has no effect on hematology, immune responses and resistance of Nile tilapia to S. iniae infection. Dietary levels of DDGS has no effect on hematology, immune responses and resistance of Nile tilapia to S. iniae infection.