Growth promoting and immunomodulatory effects of free and encapsulated dietary probiotics on Labeo rohita

Abstract

Several microorganisms used as probiotic for one aquatic species could be harmful to other species. For maximal benefit, selection of specific probiotic strain /species for the target fish species in a particular culture condition/environment is prerequisite. The aim of the study was to evaluate the comparative efficiency of two potential probiotics, yeast Geotrichum candidum QAUGC-01 and bacteria Bacillus cereus locally isolated from yogurt, a dairy product obtained from local store and gastrointestinal tract of adult rohu (Labeo rohita) respectively, and select the one for introducing into the practical culture system of the extensively culturable indigenous fish species L. rohita. The study was conducted in four phases. In the first phase, two sets of 11 weeks feeding experiments on two life stages (i.e. swim up fry and advanced fry of L. rohita) were conducted for evaluating the comparative effects of dietary B. cereus and G. candidum (at similar rate 1×109 CFU/g feed) on survival, growth rate, intestinal bacterial load, activities of intestinal enzymes (cellulase, amylase, protease), muscle composition, aspartate aminotransferase (AST) activity, hemato-immunological indices (RBCs, Hb, HCT, WBCs, MCHC, respiratory bursts and phagocytic activity, total protein, lysozyme, IgM) and resistance to the pathogen Aeromonas hydrophila. On the basis of its most significant positive impact on all studied parameters, G. candidum in contrast to B. cereus was selected for evaluating its practical application in commercial culture of L. rohita. In the second phase, G. candidum was encapsulated by using alginate alone (Alg) and in combination with starch (Alg-S) and xanthan (Alg-X) and coated with chitosan (Alg-C, Alg-S-C, Alg-X-C) and nano-chitosan (Alg-CN, Alg-S-CN, Alg-X-CN) for improving the stability, viability and target release of probiotic. The comparative structural and in vitro functional characteristics of prepared microcapsules revealed that microcapsules of G. candidum prepared with Ag-CN as compared to other formulations were more spherical in shape with smooth surface, and had highest viable count after exposure to three tested pH (2, 4 and 8), higher temperature (60ºC) for an hour and 60 days storage at ambient temperature and 4ºC. In the third phase of the study, compared the in vivo efficiency of the G. candidum encapsulated with alginate alone (Alg) or in combination with starch (Alg-S) and xanthan(Alg-X) and coated with chitosan (Alg-C, Alg-S-C, Alg-X-C) and nano-chitosan (Alg-CN, Alg-S-CN, Alg-X-CN) by feeding as a dietary supplement to the fingerlings of L. rohita for 11weeks under semi-control conditions (reared in fiberglass tanks in the outdoor facility on prepared diet). Results indicated the most significant positive effect of Alg-CN encapsulated G. candidum supplemented diet on growth, feed conversion ratio, intestinal enzyme activities, hematology, AST, ALT, ALP, hemato-immunological indices, lipid profile; high density lipoprotein (HDL), low density lipoprotein (LDL), total cholesterol and triglyceride content of fish as compared to basal diet supplemented with G. candidum encapsulated with other encapsulating formulations. In the fourth phase, commercial application of free /un-encapsulated and Alginate-chitosan encapsulated G. candidum was evaluated by feeding dietary probiotic supplemented diet to fingerlings of L. rohita reared in earthen ponds under semi-intensive culture condition. Results indicated significantly improved growth rate, intestinal enzymes activities and hemato-immunological indices. Additionally, up regulation of heat shock protein HSP 70 gene in muscle, intestine and liver tissues, increased muscle PUFA (especially Eicosatrienoic acid, linolenic, Eicosapentaenoic acid, Docosapentaenoic acid , Docosahexaenoic acid), ratio of ω3 to ω6 fatty acids, essential amino acids (particularly phenylalanine, valine and arginine) and the reduction in serum AST and ALT activities, total cholesterol and triglycerides were also observed in a group of fingerlings fed diet supplemented with Alginate-chitosan encapsulated G. candidum followed by un-encapsulated probiotic. Overall results indicated the more pronounced effect of G. candidum QAU-001 especially Alg-CN encapsulated G. candidum on survival, growth, health status, immunity, muscle proximate composition /nutritive value of L. rohita reared in semi control (reared in fiberglass tank on prepared feed) and semi intensive culture system (reared in earthen pond on live feed plus formulated feed). Thus, suggests its application as feed additive for early rearing of swim up fry and practical/commercial production of L. rohita.