Application of natural stable isotopes in aquaculture nutrition

The use of stable isotopes as nutritional tools in aquaculture represents a powerful analytical technique that allows direct assessment of the origin, flow and fate of nutrients. Aquaculture systems can be considered as simplified ecological systems and the present study aimed to apply carbon and nitrogen stable isotopes (�13C and �15N) as analytical tools in determining nutrient incorporation from different experimental diets, ingredients and feeding regimes. Due to their high growth and commercial value, Pacific white shrimp (Litopenaeus vannamei) and Senegalese sole (Solea senegalensis) were used as model organisms. In a preliminary experiment, a range of dietary ingredients and larval feeds were analyzed for elemental and isotopic compositions in order to formulate practical diets and also to design experimental larval feeding protocols. The nutritional contribution of fish meal and soy protein isolate to the growth of shrimp was estimated using an isotopic mixing model. In all cases, analysis of muscle tissue indicated a biased nutritional contribution from fish meal, so that in postlarval shrimp fed on a 46% protein diet containing a 50:50 ratio of the two ingredients, 69% of nitrogen in muscle tissue originated from fish meal, while juvenile shrimps fed the same diet, incorporated 73% of nitrogen from fish meal and 27% from soy protein. Nutritional contributions from soy protein isolate to growth were high, which highlights the suitability of soy derivatives as alternative protein sources for penaeid shrimp. Senegalese sole larvae were co-fed live and inert diets as part of two feeding regimes designed to supply varying proportions of both elements on a dry weight-basis. The relative incorporation of nutrients provided by both dietary components was successfully assessed through the analysis of stable isotopes in diets and consumers and after integration of data in an isotopic mixing model. At 23 days after hatching, the relative contributions of live and inert diets to tissue growth in larvae were 88 and 12% for a co-feeding regime consisting of 70% Artemia and 30% inert diet, respectively, and 73 and 27% for a co-feeding regime having the opposite dietary proportions. Results confirmed that partial replacement of live food with inert diets promotes growth and survival rates similar to those observed on animals fed only live food. Nutritional contributions from live food to tissue growth were significantly higher than nutrient proportions established in the co-feeding regimes. Carbon and nitrogen turnover rates and their half times in tissue were also estimated by integrating isotopic changes occurring over time in an exponential model. A second larval experiment was conducted on Senegalese sole in order to assess the isotopic influence of different larval feeding regimes on larval tissue and to assess nutrient turnover rates. Dietary effects on the digestive physiology of fish were also studied by determining digestive enzyme activity of trypsin and chymotrypsin and trypsinogen gene expression. Changes in isotopic values and digestive enzyme activities were clearly influenced by the different feeding regimes and specific dietary changes. The nutritional contribution from co-fed Artemia nauplii and inert diets to growth in mysis and early postlarval Pacific white shrimp was assessed by analyzing the �13C values in diets and shrimp tissue. Artemia nauplii and inert diets showed significantly different isotopic values and were supplied as single control diets and also in three co-feeding regimes in which 25, 50 and 75% of Artemia was replaced by inert diet on a carbon-weight basis. There was no significant difference in growth between shrimp fed Artemia only and those co-fed Artemia with inert diet, although the variability was high. Results indicated that observed nutrient contributions from Artemia nauplii were significantly higher than proportions established in the co-feeding regimes. From mysis 3 to postlarvae 5, nutrient contributions to growth in a dietary regime providing equal carbon amounts of each diet type (50:50) ranged from 73 to 87% for Artemia and from 13 to 27% for inert diet. The results demonstrated the effectiveness of up to 50% replacement of Artemia with inert diet for L. vannamei mysis and early postlarval stages, indicating also that the inert diet may provide specific nutrients not found in Artemia and that promote survival, while digestibility may limit its contribution to tissue growth. The small sample size required for isotopic analysis allowed assessing the influence of maternal and dietary sources on �13C and �15N values in shrimp eggs and early larval stages. Results from these experiments have practical applications in nutritional evaluations of larval diets, and given that the technique is useful in determining periods when organisms are more physiologically able to ingest, digest and assimilate nutrients from specific sources, the results are also of assistance in optimizing feeding protocols for the larval and postlarval rearing of Pacific white shrimp and Senegalese sole. Results support previous observations suggesting that isotope discrimination factors are potential indicators of the nutritional quality of a diet or ingredient in relation to the consuming organisms.