Effect of feed in the development of digestive System during ontogeny of major carp, Labeo rohita

Abstract

Production of good quality fish seed is the major constrain in carps culture industry, which directly depends on the feed/nutrients provided during ontogeny. Due to the unavailability of larval prepared feed, postlarvae (PL) are generally cultured in nursery earthen ponds on live feed, where high mortality and stunted growth of carp’s seed are reported as the major issues. Rohu (Labeo rohita) is the abundantly cultured Indian major carp in most Asian countries including Pakistan. The present study was designed to study the impact of live feed and prepared larval feed on the ontogeny of the digestive system of L. rohita and addresses the major limitations with live feed. This study was conducted in four phases. In the first phase nursery ponds were prepared by conventional methods and live feed was produced. Results indicated month-wise variation in primary and secondary productivity of ponds; green algae dominated during March, April, and May, while Diatoms dominated the rest of the months. Similarly, rotifers, Cladocerans, and copepods dominated the overall zooplankton mass. The proximate composition of both phytoplankton and zooplankton indicated an increasing trend in CP (%), CF (%) and dry matter (%) from March to August. Among Fatty acids, n-3 and n-6 polyunsaturated fatty acids, Docosahexaenoic Acid (DHA) and Eicosapentaenoic Acid (EPA) indicated an increasing trend from March to July and subsequently showed a decreasing trend. In the second phase of the study, based on the proximate composition of live feed, a 60% CP nanoparticulate larval diet (ND) was prepared and a 35-day feeding trial in a replicate of five was conducted to evaluate the impact of partial and total replacement of live feed (LF) with ND on rohu (L. rohita) during early rearing. Larvae, 3 days after hatch (3 DAH) were evenly distributed into 3 groups; T1 reared exclusively on LF, T2 on ND and T3 co-fed with both LF and ND (1:1). For ontogenetic changes, sampling was done at 3,10,15,20,25,30 and 35DAH from the experimental groups. All groups showed feed dependent increase in growth and the expression of genes involved in feed intake and growth with age DAH. The T3 group showed significantly higher WG, SGR, and expression of IGF-1 followed by the T2 group while the highest expression of ghrelin and GHS-R were observed in a T3 followed by a T1 group. Furthermore, leptin showed the highest expression in a T2 followed by the T1 group. The intestinal enzymes showed variable trends with the highest activity of cellulase, amylase and protease in the T1, T2 and T3 groups respectively. The overall result of this trial showed the most significant effect of co-feeding followed by feeding ND, thus indicating the need for standardization of the prepared diet by determining the optimum requirement of CP (%) and xi crude fat (%). Thus, in the third phase of the study, two independent experimental trials were conducted to standardize protein and fat levels in the larval feed of L. rohita. To optimize the dietary protein requirement, four experimental larval feeds having 45, 50, 55 & 60 % CP were prepared and a 35-day feeding trial in replicate of five was conducted with the same experimental set up. Among all the diets, P55 followed by P60 showed the most significant positive effect on all studied indices except amylase activity which did not show any significant difference among groups. Moreover, in all groups, GH showed a positive correlation, while MyoD and MYF5 showed a negative correlation with age DAH. To optimize the dietary lipid level, another experiment was conducted in the same experimental set up and rearing conditions except that 55% CP was fixed in experimental diets while fat (fish oil) was added at graded levels (4, 8, 12, 16 %), using fish oil as the major lipid source. Lipid at 8% followed by 12% inclusion level showed the most significant effect on all studied indices including growth, alkaline phosphatase and lipase activity. The ontogeny variation of protease activity showed a uniform increase up to 25DAH, followed by a surge at 30DAH and then slight but non-significant decrease in activity. The ontogenetic changes in GH gene expression showed a positive correlation with age DAH, i.e., expression uniformly increased with age DAH while MyoD and MYF5 expression showed a negative relation with DAH, i.e., uniformly decreased with age DAH. After understanding the ontogenetic variation in the digestive enzymes trend in response to live feed, ND and various protein and lipid levels a standardized nanoparticulate feed (SNF) with 55% CP and 8% fish oil were formulated and prepared and 35 days feeding trial in a replicate of 5 was conducted to compare the efficiency of SNF as compared live feed (LF). Results indicated a significant effect of both independent variables (feed and age DAH) on all studied indices. The comparative effect of LF and SNF indicated significantly higher survival (%), growth rate, activity of digestive enzymes and expression of MyoD, MYF5, GH, IGF-1 and MEF2A in a group reared on SNF while, myostatin expression was significantly higher in a LF group compared to SNF group. Both lipase and proteases were seen to be enhanced with optimum lipid and protein level in the diet (experiment 3 and 4). Discussing this increase in proteases and lipase as the possible outcome of efficient utilization (in term of early maturation of digestive tissues/organ) of optimum protein and lipids, in experiment 5 although a high protein % was available in LF but the trypsin and chymotrypsin activity was significantly higher in SNF group. This inference may conclude a slight instantaneous impact of diet over direct release of certain digestive enzymes, however the major xii change come from the maturation of that tissue. Further, the only slight change in proteas activity in experiment 5 (compared to experiment 3, P55) may be due to the early shift of larvae to gut peptidases (high trypsin and chymotrypsin activity with SNF). Overall, this study confirms the role of the extrinsic factor, larval feed on the regulation of ontogenetic changes in the release of digestive enzymes and their effect on survival and growth and indicates the most significant effect of SNF compared to live feed. Thus, the replacement of life feed with SNF could be recommended for improving the survival and growth during early rearing and to produce quality seed of L. rohita.