Introduction
Grey mullet (Mugil cephalus) is a promising omnivorous species for the sustainable diversification of aquaculture due to its low dietary nutrients requirements and the broad market for its roe (bottarga). Bacterial single cell ingredients produced by fermenting industry by-products represent a sustainable alternative of fishmeal and soybean meal (Glencross et al., 2020). The application of bacterial single cell protein (SCP) ingredient was assessed on growth, digestive enzyme activity and on the gene expression of PepT-1 at three different time T0 (0h before feeding), T1 (0/6h after feeding), T2 (6/12h after feeding). To the best of our knowledge, this is the first characterisation of digestive enzymes at different times in grey mullet under captive conditions and different diets.
Materials and Methods
Three isonitrogenous, isolipidic and isoenergetic experimental diets were formulated to contain increasing inclusion levels of bacterial SCP (SCP0, 0%; SCP10, 10%; SCP20, 20%). Diets were tested on triplicate fish groups of 45 individuals (initial weight: 67.9 g) over a period of 113 days. During the trial, feed was provided to apparent satiation during a 6 hours meal once a day, temperature was kept at 26.8 ± 2.5 °C and oxygen above 7.5 mg L-1. At the end of the trial final body weight (FBW), specific growth rate (SGR), feed intake (FI) and feed conversion rate (FCR) were calculated. Digestive enzyme activity (total alkaline protease, trypsin, chymotrypsin, leucine aminopeptidase, α-amylase, bile salt-activated lipase), anterior and posterior intestine of brush border (aminopeptidase and alkaline phosphatase) and gene expression of PepT-1 were performed in three different times T0 (0h before feeding), T1 (0/6h after feeding), T2 (6/12h after feeding). Differences among treatments were considered significant at P < 0.05.
Results
At the end of the trial, fish fed SCP0 showed a significantly higher body weight and SGR and a significantly lower FI and FCR compared to SCP10 and SCP20 (P < 0.05). Temporal variation of digestive enzymes activity in different diet is reported in Fig.1. In fish fed SCP0, total alkaline proteases activities, trypsin and chymotrypsin increased significantly at T1 in comparison to T0, followed by a decreasing pattern at T2. No significant temporal variation for total alkaline protease, trypsin and chymotrypsin was detected in SCP10 and SCP20. On the other hand, in T0 and T1 the SCP0 diet was statistically different from SCP10 and SCP20. No significant differences were found in T2. In leucine aminopeptidase fish fed SCP0 showed an increase significantly in T2 compered to T0 and T1 which gave the same results. The same trend has been found for other diets. Fish fed with SCP10 and SCP20 in T2 exhibited significant differences from T1 and T0 that followed the same values. Comparison between time with diet, it turned out that in T1 the diet values SCP0 were different from SCP10 and SCP20 values. No differences in T0 and T2. In fish fed SCP0 diet, α-amylase was significantly higher in T1 than T0. No differences were found in the comparison between time with diet. In bile salt-activated lipase, fish fed SCP0 diet in T2 showed a significantly high increase compared with fish fed SCP0 in the other two times. No differences in the comparison between time with diet. The results measured in the intestinal brush border (anterior and posterior intestine) were very similar to each other. Significantly higher differences of aminopeptidase were encountered in fish fed SCP0 diet in the anterior intestine at T1 than fish fed SCP0 at T0 and T2. No significant temporal variation for aminopeptidase was detected in SCP10 and SCP20. No differences in comparison of time with diet. No significant temporal variation for alkaline phosphatase were found. Instead there has been observed a significant difference in the time-diet interaction. In the posterior intestine, alkaline phosphatase displayed an increase in fish fed SCP10 diet at T1 from SCP0 diet at T1. In the results of gene expression of PepT-1 as shown in Fig. 2, there was a significant reduction in the anterior intestine in fish fed SCP0 diet at T0 than T1 and T2. No significant temporal variation was detected in SCP10 and SCP20. No differences in the comparison time with diet. There was a significant reduction in fish fed SCP0 diet in posterior intestine at T1 than T0 and T2. In the comparison time with diet, it turned out that in T2 in posterior intestine the diet values for SCP0 were higher from the SCP10 and SCP20 values.
Discussion and Conclusion
The lower performance of C. glutamicum SCP as a protein source for grey mullet is probably due to a particular organisation of the digestive system because there was incomplete protein utilisation of SCP due to poor lysis of cell-wall components and also associated with the captive conditions (failure in the development of a functional gizzard). Analyses of enzyme activity showed the important key role in the physiology of the grey mullet digestive system. Proteases, in particular, played a key role but were negatively affected by diet. The temporal variation of grey mullet is similar to that found in the literature for other Mediterranean species of commercial interest. These are the first results concerning the enzymatic activity of the grey mullet. This highlights the extreme importance of analyses of the physiology of the species and the most adaptable diet.
References
Glencross, B., Huyben, D., Schrama, J., 2020. The Application of Single-Cell Ingredients in Aquaculture Feeds—A Review. Fishes 5, 22. https://doi.org/10.3390/fishes5030022