Effects of Bacillus subtilis as a single strain probiotic on growth, disease resistance and immune response of striped catfish (Pangasius hypophthalmus)

Abstract

The present study investigated the potential role of Bacillus subtilis as probiotic in striped catfish (Pangasius hypophthalmus). Fish (initial weight = 150.00±2.63g n = 180) were stocked in circular tanks. Four isonitrogenous (30%) and isolipidic (3.29%) diets were formulated having supplementation of B. subtilis at four different levels (P0; 0, P1: 1×106, P2: 1×108 and P3: 1×1010 CFU/g). Each treatment had three replicates, while each replicate had fifteen fish. The trial started on second week of July and continued for eight weeks. Growth, feed conversion ratio, crude protein content, the concentration of amylase and protease, the profile of both dispensable and non-dispensable amino acids in all four dietary groups increased with a gradual increase of B. subtilis in the diet. At the end of growth experiment, fish in all four groups were exposed to Staphylococcus aureus (5×105 CFU/ml). After S. aureus challenge, fish fed with B. subtilis responded better to damage caused by reactive oxygen species and lipid peroxidation and better survival rate. The catalase and superoxide dismutase level also increased in response to bacterial challenge in B. subtilis fed groups. On the other hand, the concentration of malondialdehyde gradually decreased in these groups (+ve P0 >P1>P2>P3). It is concluded that supplementation of B. subtilis as a probiotic improved the growth, protein content, antioxidant response and immunocompetency against S. aureus in striped catfish. The optimum dosage of B. subtilis, at a concentration of 1×1010 CFU/g, resulted in the most favorable outcomes in striped catfish. This single bacterial strain can be used as an effective probiotic in large scale production of aquafeed for striped catfish. Future studies can investigate this probiotic's impact in the intensive culture of the same species.

Fig 1. Step wise process of experimental work performed during the growth experiment and bacterial challenge test.

Fig 2. Histological changes in gut.

Light micrographs of a paraffin section stained with eosin (40x). a; gut in -veP0, b; gut in P1, c; gut in P2, d; gut in P3, e; gut in +ve P0. GC; Goblet cells, LP; Laminar propria, N; Nucleus, L; Lumen, CE; Columnar epithelium, FV; Fusion of villi, LL; Large lumen, FLV; Flattened villi, DCML; Damaged circular muscle layer, DL; Distended lumen, DLML; Damaged longitudinal muscle layer, VF; Vacuole formation, SLP; Swelling of lamina propria, CCA; Cracked clay appearance of the tissues, SLML; Swelling of longitudinal muscle layer, DGC; Damaged goblet cells, DMM; Disarrangement of muscularis mucosa.

Fig 3. Histological changes in gills and liver.

Light micrographs of a paraffin section stained with eosin (40x). a; gills in -veP0, b; gills in P1, c; gills in P2, d; gills in P3, e; gills in +veP0, f; liver in -veP0, g; liver in P1, h; liver in P2, i; liver in P3, j; liver in +veP0. PL; Primary lamellae, SL; Secondary lamellae, FSL; Fusion of secondary lamellae, DSL; Degeneration of secondary lamellae, HT; Hypertrophy, DPL; Degeneration of primary lamellae, TD; Tissue debris, A; Aneurism, H; Hypertrophy, NH; Normal hepatocytes, GC; Granular cytoplasm, BC; Blood congestion HD; Hepatocyte generation, CSN; Central spheroidal hepatocyte nucleus, N; Cell necrosis, PN; Pyknotic nuclei, CD; Cytoplasmic degeneration, IEF; Infiltration of oedematous fluid, rCV; Rupturing of the central vein, V; Vacuolization of hepatocytes.

Fig 4. Histological changes in muscles and kidney.

Light micrographs of a paraffin section stained with eosin (40x). a; muscles in -ve P0, b; muscles in P1, c; muscles in P2, d; muscles in P3, e; muscles in +veP0, f; kidney in -ve P0, g; kidney in P1, h; kidney in P2, i; kidney in P3, j; kidney in +ve P0. MF; Myofibrils; GFMF; Gap formation in myofibril, IMFS; Inter myofibrillar space, DMF; Disintegrated myofibrils, EMF; Oedema between muscle fibre, MD; Muscle degradation, ME; Muscle oedema, G; Glomerulus, CD; Collecting duct, DG; Degenerative glomerulus, IBS; Increased bowman space, FRT; Fusion of renal tubule, DRT; Degenerative renal tubule, CG; Congestion of glomerulus, N; Necrosis, H; Hemorrhage, A; Atrophy.