Fish skeletal development is under the continuous influence of biotic and abiotic parameters (1) . Being a valuable welfare index (2), skeletal abnormalities appearance undermines the quality of the experimental results and inevitably, the marketabili ty of the aquaculture products. One of the most critical factors towards normal fish skeletal development is appropriate larval nutrition (3). Protein fraction, being the major component of fishmeal, has been proven advantageous for larval growth, survival and antioxidative response in the form of various sources and hydrolysis levels (4). Simultaneously, an effect of dietary peptides in harmonious skeletal development has been observed (5 ). Despite the well characterized beneficial effect of small dietary protein products on the normal development of fish skeletal elements, the molecular basis of these observations remains scare.
The purpose of the present study is to evaluate the effect of three experimental diets with partial peptide incorporation on a) the skeletal development of sea bass larvae and b) the resistance of their vertebral column against swimming induced lordosis . A possible effect of the diets on gene markers of larval maturation (digestion, ossification, and mineralization) was also examined.
Materials and methods
Three isonitrogenous experimental diets were formulated by partial substitution of the total protein source with 0% (C, Control group), 6% (P6) and 12% (P12) shrimp di and tri-peptides. After a short Artemia inclusion (6-20dph), experimental diets were provided exclusively throughout the trials. Study of larval and juvenile quality was performed through means of whole mount staining, histology analysis and micro-ct scanning. To evaluate bones’ integrity against haemal lordosis induction, early juveniles with normal skeletal development were subjected to swimming challenge test (SCT, 6). Gene expression analysis on selected gene of interest was performed by means of rt-qPCR.
Results and Discussion
Results of the early skeletal development revealed an advantageous peptide-enhanced ossification in the case of P12 individuals (Fig. 1). Similarly, P12 had a beneficial effect in the frequency of exercise-induced lordosis after the SCT reducing the deformities rate in only 15.8% (Fig. 2a). RNA analyses confirmed a peptide-enhanced total larval maturation, in te rms of their digestive function, bone mineralization and myogenesis in P12 . After the exercise, a differential regulation of the genes in the affected haemal area confirmed the scenario about an early peptide nutritional programming of the post-larval musculoskeletal responses.
With the present project, we highlight the beneficial effect of the early peptide diets enhancing the total larval maturation in terms of digestive function and ossification in sea bass larvae. The limitations concerning the free amino acid participation should be taken into consideration . Potential incorporation of these small di- and tripeptides in inert larval but also post-larval diets could enhance larval and post-larval skeletal development.