We used a factorial approach to distinguish maintenance from growth requirements for protein, lysine and methionine in the black tiger shrimp, Penaeus monodon. Juvenile P. monodon (initial weight 2·4 g) were fed during 6 weeks one of ten semi-purified diets based on casein and purified amino acids (AA) as N source. The diets contained four levels of crude protein (CP, from 5 to 54 % DM diet) with two levels (% CP) of lysine or methionine (normal or 30 % deficient). Requirements were determined using linear and non-linear regression models. We could thus obtain the first ever data on maintenance (N equilibrium) requirements for CP and AA in P. monodon. CP requirements for maintenance (4·5 g/kg body weight (BW) per d) represented approximately 19 % of the CP requirement for maximal N gain (23·9 g/kg BW per d). The marginal efficiency of utilisation reached a maximum of 38 % for N, 0·77 for lysine and 1·62 for methionine using N gain as response. Lysine requirements were 0·20 g/kg BW per d for N maintenance and 1·40 g/kg BW per d for maximal N gain. Methionine requirements were 0·11 g/kg BW per d for N maintenance and 0·70 g/kg BW per d for maximal N gain. The lysine (5·8 %) and methionine (2·9 %) requirements for maximal N gain, expressed as percentage of protein requirement, agree with literature data using a dose -response technique with smaller P. monodon. The observed interaction between dietary CP and methionine for N gain demonstrates that requirements for indispensable AA (expressed as % CP) cannot be evaluated separately from CP requirements. Crustaceans: Protein requirement: Indispensable amino acids: Logistic model: Marginal utilisation efficiencyThe black tiger shrimp (Penaeus monodon) is the second most cultured crustacean species worldwide (1) . Due to the importance of protein for shrimp growth, its high cost in formulated feeds and the environmental implications of N losses, it is essential to gain a better understanding of N requirements and N utilisation in P. monodon. Available data on crude protein requirements (CP, % DM diet) of P. monodon show a large degree of variability, i.e. from 36-40 % (2) up to 50 % (3) . Several factors, e.g. differences in protein source, dietary energy level, life stage, rearing conditions and, in particular, differences in feed intake (FI), can explain some of this variation (4) . The confounding effect of FI on dietary protein requirement estimates has also been illustrated with the pacific white shrimp, Litopenaeus vannamei (5,6) . The latter authors (6) demonstrated that maximum weight gain could be obtained by a wide range of dietary CP levels (30-40 % DM diet) at different feed allowances (50, 75 and 100 % of typical daily intakes), underlining the importance of expressing protein requirements on absolute basis rather than as a percentage of the diet. The use of the factorial approach, which allows the distinction between maintenance and growth for estimating protein requirements, has been initially developed for terrestrial animals (7,8) and has also been appl...
This 35-d feeding experiment examined in juvenile shrimp Penaeus monodon (3·3 g initial body weight) the effects of methionine (Met), choline and cystine on protein accretion and the activity of two key enzymes of remethylation (betaine -homocysteine methyltransferase; BHMT) and trans-sulfuration (cystathionine b-synthase; CBS). The interaction between Met and choline was tested using semi-purified diets either adequate or limiting (30 or 50 %) in total sulphur amino acid (SAA) content with a constant cystine:Met ratio. The diets contained either basal or excess choline (3 v. 7 g/kg feed). Cystine was added to two other 30 and 50 % Met-limiting diets to adjust the SAA supply to that of the control diet in order to evaluate the interaction between Met and cystine. As expected, N accretion was significantly lower with the SAA-limiting diets but increased back to control levels by the extra choline or cystine, demonstrating their sparing effect on Met utilisation for protein accretion. We show, for the first time, the activities of BHMT and CBS in shrimp hepatopancreas. Only BHMT responded to the SAA deficiencies, whereas the extra choline and cystine did not stimulate remethylation or down-regulate trans-sulfuration. Our data also suggest the capacity of P. monodon to synthesise taurine, being significantly affected by the cystine level in the 30 % SAA-limiting diets. Further research is warranted to better understand the metabolic regulation of taurine synthesis in shrimp and of the observed Met-sparing effects.Key words: Crustaceans: Sulphur amino acids: Methionine utilisation: Methionine sparing: TaurineThe marine black tiger shrimp Penaeus monodon is the world's second most cultured crustacean species (1) . For crustacean shrimp, as for farmed finfish, plant protein sources are increasingly included in feeds in order to reduce the reliance on wild-caught marine protein sources. However, the replacement of fish or shrimp meal by plant protein sources changes the amino acid (AA) profile of the diet, with methionine (Met) as one of the first-limiting essential AA (2,3) . In P. monodon fed an optimal crude protein level,we have previously noted that a 30 % Met deficiency diminished protein accretion (4) and increased deamination (5) , suggesting a change in AA catabolism in shrimp receiving an imbalanced dietary Met supply. In P. monodon, or crustacean species in general, not much is known on the metabolic utilisation of Met besides its need for protein synthesis. In contrast, the importance of Met as a methyl-group donor for methylation reactions and as a precursor of other sulphur-containing compounds, such as cysteine or taurine, is well recognised in vertebrates (6 -10) . As such, homocysteine (Hcy), at the branch point of the three major pathways of Met metabolism (transmethylation, remethylation and transsulfuration), is often regarded as a regulatory component of Met metabolism since Hcy can be either trans-sulfurated for cysteine production (catalysed by cystathionine b-synthase; CBS) or remethyla...
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