Feeding is the main item of fish production, since it represents between 67 and 70 % of the operational cost (Perea et al. 2022 Perea, C., Garcés, Y., Morales, Y., Jiménez, M., Hoyos, J.L. & Vivas, N. 2022. "Digestibility of enzymatic hydrolyzates from animal origin viscera in Piaractusbrachypomus, Cuvier 1818". Biotecnología en el Sector Agropecuario y Agroindustrial, 20(1): 54-67, ISSN: 1909-9959. https://doi.org/10.18684/rbsaa.v20.n1.2022.1606. ). Traditionally, fishmeal and soybeans are the main protein sources for the manufacture of aquaculture feed in Cuba. They are generally imported at high prices and, sometimes, there is little availability in the market, issues that have negative implications for the sustainability of feed and fish production. That is why there are studies on the evaluation of the by-products generated by the fishing sector that may constitute feeding sources for the development of intensive cultivation of Clarias gariepinus.
Llanes et al. (2000) Llanes, J., Toledo, J. & Lazo de la Vega, J. 2000. "Utilización del desecho de pescado en la alimentación del bagre africano Clarias gariepinus". AcuaCUBA, 2(2): 26-31. ISSN: 1608-0467. reported that by-products generated from fish processing (heads, viscera, bones, bones) have high nutritional value for C. gariepinus, which is why they have been established as the main protein source of the diet of this species. The industrial processing of spiny lobster (Panulirus argus) is also among the activities of the fishing sector. The wastes from this processing represent between 20 and 40 %. They are usually discarded and become environmental pollutants (Ramírez et al. 2022 Ramírez, M.A., Peniche, C., Rodríguez, T. & Llanes, J.E. 2022. Caracterización fisicoquímica y microbiológica del concentrado proteico de langosta, como aditivo en la alimentación animal. Taller Alimentos, Aditivos y Bioinsumos. Memorias Convención Producción Animal y Agrodesarrollo. Del 10 al 14 de octubre 2022. Centro de Convenciones Plaza América. Varadero, Matanzas, Cuba. ISBN: 978-959-7171-86-7. ).
Obtaining pulp from lobster cephalothorax generates a by-product that consists of a pink meat mass, but with many exoskeleton microparticles. Additionally, it has a seafood smell, which can be attractive to fish. The objective of this study was to evaluate the productive performance of C. gariepinus young fish, fed with by-products from the extraction of pulp from the spiny lobster cephalothorax.
The research was carried out in the Fish Nutrition Laboratory of the Aquaculture Technologies Development Company (EDTA) in Havana, Cuba. This facility has circular cement tanks, with a capacity of 68 L and a constant water flow 24 hours a day.
C. gariepinus young fish were used, which were acclimatized in a 4.5 m2 pool for a week, where they received commercial feed (29.71 % crude protein and 11.40 MJ/kg of digestible energy). After this time, 240 animals, with an initial mean weight of 10.16 ± 0.07 g, were selected. The treatments were: control (100 % fish by-products), TI (75 % fish by-products and 25 % lobster by-products), TII (50 % fish by-products and 50 % lobster by-products) and TIII (100 % lobster by-products), with three repetitions each and 20 fish per tank, which was considered the experimental unit.
By-products from tilapia filleting and the lobster cephalothorax pulp extraction process were used, which were ground in a meat mill (JAVAR 32, Colombia) until reaching a particle size of 3 mm. The mixing of rations TI and TII was carried out in a mixer (HOBART M-600, Canada) for 5 min. All foods were stored in plastic containers with lids, at -10 oC. Bromatological analyzes were performed in triplicate on fish and lobster by-products (AOAC 2016 AOAC. 2016. Official methods of analysis of AOAC International. 20th ed., Rockville, MD: AOAC International, ISBN: 978-0-935584-87-5, Available: <http://www.directtextbook.com/isbn/9780935584875>, [Consulted: September 22, 2016]. ).
Temperature and dissolved oxygen values were taken every day with a portable digital oximeter (HANNA®, Romania). The feeding rate was 15 % of the biomass, supplied in two daily rations for 45 d. Group samplings were carried out every 10 days, to adjust the rations. At the end of the bioassay, every fish was individually weighed to calculate productive indicators:
Final mean weight
Statistical analysis. To analyze the results, one-way classification analysis of variance was carried out, according to a completely randomized design. The mean values were compared using Duncan (1955) Duncan, D.B. 1955. "Multiple Range and Multiple F Tests". Biometrics, 11(1): 1-42, ISSN: 0006-341X. https://dx.doi.org/10.2307/3001478. test, in the necessary cases. The theoretical assumptions of the analysis of variance were verified for the variables food supplied per fish, daily weight gain and food conversion based on the tests of Shapiro and Wilk (1965) Shapiro, S.S. & Wilk, M.B. 1965. "An Analysis of Variance Test for Normality (Complete Samples)". Biometrika, 52(3/4): 591–611, ISSN: 0006-3444. https://doi.org/10.1093/biomet/52.3-4.591. for the normality of the errors and according to the test of Levene (1960) Levene, H. 1960. Robust tests for the equality of variance In: Olkin, I., Contributions to Probability and Statistics: Essays in Honor of Harold Hotelling, Stanford University Press, pp. 278–292, ISBN: 978-0-8047-0596-7. for homogeneity of variance. The variables met the theoretical assumptions of the ANOVA. The InfoStat statistical package, version 2012, was used (Di Rienzo et al. 2012 Di Rienzo, J. A., Casanoves, F., Balzarini, M. G., González, L., Tablada, M. & Robledo, C. W. 2012. Infostat versión 2012. Grupo Infostat. Universidad Nacional de Córdoba, Argentina. Available: http://www.infostat.com.ar. [Consulted: October 9, 2019] ).
Water temperature in the tanks fluctuated between 26.3 and 27.5 °C, the dissolved oxygen concentration between 5.3 and 6.8 mg/L and the pH was maintained between 7.6 and 7.9. These values are considered comfort for the good performance of the species (Toledo et al. 2015 Toledo, J., Llanes, J.E. & Romero, C. 2015. "Nutrición y alimentación de peces de aguas cálidas". AcuaCUBA, 17 (1): 5-22, ISSN: 1608-0467. ).
The chemical composition of lobster by-products showed values of 51.6 % humidity, 17.8 % crude protein, 1.23 % lipids, 52.6 % ash, 14.91 % calcium and 1.31 % phosphorus. The resulting mass of lobster by-products continued to have very small exoskeleton particles despite milling. Fish were very attracted to lobster by-products, which may be evidence of the high concentration of free amino acids. However, their intake was not complete on most occasions in TIII (100 % lobster by-products), which can be related to the presence of particles from the exoskeleton. This did not happen in the rest of treatments that were mixed with fish by-products.
Survival was 100 % for all treatments, which shows its safety and its use does not compromise animal health. However, the increase of lobster by-products in the rations decreased the amount of food supplied per animal (table 1). This reduction can be attributed to the lower growth of fish that consumed lobster by-products, which is why they received less food, since feed is restricted according to body weight.
| Indicators | Control 100 % fish by-products | T I 75 % fish by-products 25 % lobster by-products | T II 50 % fish by-products 50 % lobster by-products | T III 100 % lobster by-products | SE | P |
|---|---|---|---|---|---|---|
| Provided food/fish, g | 137.15 a | 120.38 b | 109.72 c | 86.53 d | 5.55 | 0.0001 |
| Final weight, g | 74.51a | 52.91b | 43.01c | 20.51d | 1.97 | 0.0001 |
| Daily weight gain, g/d | 1.43 a | 0.95 b | 0.73 c | 0.23 d | 0.05 | 0.0001 |
| Feed conversion | 2.41 a | 3.18 b | 3.78 c | 9.81 d | 0.89 | 0.0001 |
Different letters in the same line differ at p < 0.05 according to Duncan (1955) Duncan, D.B. 1955. "Multiple Range and Multiple F Tests". Biometrics, 11(1): 1-42, ISSN: 0006-341X. https://dx.doi.org/10.2307/3001478.
Growth indicators and feed conversion (table 1) were unfavorable, as lobster by-products levels increased, due to lower feed intake and, therefore, proteins. This indicates the low nutritional value of lobster by-products for feeding C. gariepinus, regardless of the level of proteins and essential amino acids that they may present. The results could also show low activity of chitinolytic enzymes in the digestive tract of the species, which means that crustaceans are not among the main groups for feeding clarias in their natural environment.
Gutowska et al. (2004) Gutowska, M., Drazen, J. & Robinson, B. 2004. "Digestive chitinolytic activity in marine fishes of Monterey Bay, California". Comparative Biochemistry and Physiology, Part A.139: 351–358, ISSN: 1531-4332. https://doi.org/10.1016/j.cbpb.2004.09.020. measured the activities of quinolytic enzymes in the stomachs and intestines of 13 species of marine fish and found, in the majority, chitinous material and the highest enzyme activity in the stomach, which showed that crustaceans are part of the feeding habits of these fish. The cited authors conjectured that the function of chitinase is to assist in the breakdown of the exoskeleton of prey, to allow other digestive enzymes access to soft internal tissues, while that of chitobiase is to break down chitin dimers into absorbable monomers of chitin β N-acetyl-glucosamine.
The negative results of this study with lobster by-products could be related to the high contents of natural chitin present in these by-products. Chitin constitutes an important part of the exoskeleton of crustaceans (Borić et al. 2020 Borić, M., Vicente, F. A., Jurković, D. L., Novak, U. & Likozar, B. 2020. "Chitin isolation from crustacean waste using a hybrid demineralization/DBD plasma process". Carbohydrate Polymers, 246: 1–8, ISSN: 1879-1344. https://doi.org/10.1016/j.carbpol.2020.116648. ) and it is a natural biopolymer composed of a mixture of polymers, mainly unbranched N-acetyl-D-glucosamine and a small amount of D-glucosamine (Soetemans et al. 2020 Soetemans, L., Uyttebroek, M. & Bastiaens, L. 2020. "Characteristics of chitin extracted from black soldier fly in different life stages". International Journal of Biological Macromolecules, 165: 3206-3214, ISSN: 1879-0003. https://doi.org/10.1016/j.ijbiomac.2020.11.041 ). This complex carbohydrate, although commonly found in the natural diet of many fish, is considered a non-digestible fiber that reduces digestibility of proteins and lipids and, consequently, affects the efficiency of nutrient absorption in the intestinal tract (Toledo et al. 2015 Toledo, J., Llanes, J.E. & Romero, C. 2015. "Nutrición y alimentación de peces de aguas cálidas". AcuaCUBA, 17 (1): 5-22, ISSN: 1608-0467. ).
Mergelsberg et al. (2019) Mergelsberg, S.T., Ulrich, R.N., Xiao, S. & Dove, P.M. 2019. "Composition Systematics in the Exoskeleton of the American Lobster, Homarusamericanus and Implications for Malacostraca". Frontiers in Earth Science, 7: 69, ISSN: 2296-6463. https://doi.org/10.3389/feart.2019.00069. reported that chitin naturally found in lobster by-products is associated with proteins, forming a glycoprotein complex with a complex structure that makes its degradation difficult. Previously, Lu and Ku (2013) Lu, C.H. & Ku, C.C. 2013. "Effects shrimp waste meal on growth performance and chitinas activity in juvenile cobia (Rachycentron canadum) ". Aquaculture, 44(8): 1190-1195, ISSN: 0044-8486. https://doi.org/10.1111/j.1365-2109.2012.03097.x. studied the effects of replacing 0, 10, 20 and 25 % of fishmeal with shrimp waste meal (HRC, in Spanish) on juvenile cobia Rachycentron canadum for six weeks and reported decreased protein efficiency and low lipid content in muscle, with 20 and 25 % of HRC. Furthermore, they reported that chitinolytic activity was high in the pyloric caeca, but there was only a slight increase in the 10 % HRC diet in the intestines, a level they recommended to replace fish meal.
Karlsen et al. (2017) Karlsen, O., Amlund, H., Berg, A. & Olsen, R. 2017. "The effect of dietetic chitin on growth and nutrient digestibility in farmed Atlantic cod, Atlantic salmon and Atlantic halibut". Aquaculture Research, 48: 123-133, ISSN: 1365-2109. https://doi.org/10.1111/are.12867. noted a reduced growth rate in Atlantic salmon (Salmo salar), fed with diets rich in chitin, and considered the hypothesis that chitin acts as an energy trap when fish are not able to digest and use correctly this polysaccharide. In contrast, Elsefary et al. (2021) Elserafy, S., Abdel-Hameid, N. & Abdel-Salam, H. 2021. "Effect of shrimp waste extracted chitin on growth and some biochemical parameters of the Nile tilapia". Egyptian Journal of Aquatic Biology and Fisheries, 25(1): 313 – 329, ISSN: 2536-9814. https://doi.org/10.21608/EJABF.2021.143244. evaluated chitin extracted from shrimp exoskeletons in the growth of Nile tilapia (Oreochromis niloticus) young fish at 0, 2, 5 and 10 % inclusion levels and reported the best growth and feed conversion with 5 %. When the same diets were supplemented with 1 g of PRO-PAC probiotic/kg of diet, the results with 10 % were superior.
Another important aspect to consider is the high content of calcium carbonate present in lobster by-products, which increases dietary calcium level and, therefore, produces a calcium-phosphorus (Ca-P) imbalance in the fish rations. Lall and Kaushil (2021) Lall, S.P. & Kaushik, S.J. 2021. "Nutrition and Metabolism of Minerals in Fish". Animals, 11: 2711, ISSN: 2076-2615. https://doi.org/10.3390/ani11092711. reported that there is still little knowledge of mineral nutrition of fish with respect to their dietary requirements, physiological functions and absorption from the gastrointestinal tract and bioavailability in feed ingredients. However, the amount of calcium (Ca) to include in the fish diet depends on the culture water. In the case of freshwater fish, if the waters are hard as in Cuba, Ca dietary needs are very low.
It is difficult to establish Ca dietary requirements in fish due to its presence in water, since animals can absorb it. Calcium absorption by the environment varies among species, the endocrine system, availability in the diet and Ca concentration in water (Toledo et al. 2015 Toledo, J., Llanes, J.E. & Romero, C. 2015. "Nutrición y alimentación de peces de aguas cálidas". AcuaCUBA, 17 (1): 5-22, ISSN: 1608-0467. ). It is well demonstrated that diets should maintain an ideal Ca:P ratio (1.5:1.0). Otherwise, the excess of both can cause problems in the body development of the fish, making it difficult for them to absorb and remain in the culture environment (Velazco and Gutiérrez 2019 Velazco, J.S. & Gutiérrez, M. 2019. "Aspectos nutricionales de peces ornamentales de agua dulce". Revista Politécnica, 15(30): 82-93, ISSN: 2256-5353. https://doi.org/10.33571/rpolitec.v15n30a8. ). Digestibility and availability of amino acids also decreases when there is excess calcium in fish rations (Perea et al. 2022 Perea, C., Garcés, Y., Morales, Y., Jiménez, M., Hoyos, J.L. & Vivas, N. 2022. "Digestibility of enzymatic hydrolyzates from animal origin viscera in Piaractusbrachypomus, Cuvier 1818". Biotecnología en el Sector Agropecuario y Agroindustrial, 20(1): 54-67, ISSN: 1909-9959. https://doi.org/10.18684/rbsaa.v20.n1.2022.1606. ).
D’Abramo (2021) D'Abramo LR. 2021. "Sustainable aquafeed and aquaculture production systems as impacted by challenges of global food security and climate change". Journal of the World Aquaculture Society, 52 (6): 1162-1167, ISSN: 1749-7345. https://doi.org/10.1111/jwas.12867. reported that excess dietary calcium can have a negative impact on digestion because it joins to fatty acids to form soaps and reduces the availability of energy sources. Porn-Ngam et al. (1993) Porn –Ngam, N., Satoh, S., Takeuchi, T. & Watanabe, T. 1993. "Effect of the ratio of phosphorus to calcium on zinc availability to rainbow trout in high phosphorus diet". Nippon Suisan Gakkaishi, 59(12): 2065-2070, ISSN: 1349-998X. stated that excess dietary Ca decreased growth, feeding efficiency, bone mineralization and low P absorption was found in common carp Ciprinus carpio and rainbow trout Onchorhynchus mykiss. Regarding the latter, when available P in the diet is low, fish increase their absorption in the intestine, reduce P excretion in the kidney and mobilize P from the skeleton to cover vital functions in other compartments of the body. Sustained demineralization of bones over a long period will weaken the skeleton and cause deformities (Velazco and Gutiérrez 2019 Velazco, J.S. & Gutiérrez, M. 2019. "Aspectos nutricionales de peces ornamentales de agua dulce". Revista Politécnica, 15(30): 82-93, ISSN: 2256-5353. https://doi.org/10.33571/rpolitec.v15n30a8. ).
It is concluded that the by-products of the pulp extraction process from the spiny lobster cephalothorax were not feasible for feeding C. gariepinus. Its transformation into products with high added value, such as chitin, chitosans and protein concentrates, is recommended for subsequent evaluation as growth enhancers in this species.