In vitro assay of plant protease inhibitors from four different sources on digestive proteases of rohu, Labeo rohita (Hamilton), fingerlings

The in vitro inhibitory effect of protease inhibitors from four seed extracts (soybean, grasspea, black gram and horse gram) on digestive proteases of rohu was assessed by enzyme inhibition assay and substrate sodium dodecyl sulphate‐polyacrylamide gel electrophoresis. High proteolytic activity was detected in the intestinal extract of rohu (Labeo rohita) fingerlings at two different pH ranges (8–8.5 and 10–11). That protein digestion occurs mainly in the alkaline condition in this fish without a stomach is evident from very high trypsin activity (0.95±0.04 benzoyl‐dl ‐arginine‐p‐nitroanilide U mg protein⁻¹) in the intestine. In case of grass pea seed, more than 50% inhibition of alkaline protease activity was recorded when the ratio of inhibitor to enzyme was 9.41 μg U⁻¹. More than 40% inhibition of protease activity was recorded in case of horse gram seed when the ratio of inhibitor to enzyme was 5.51 μg U⁻¹. Black gram at 11.0 μg U⁻¹ and soybean seed proteins at 62.75 μg U⁻¹ resulted in 50% and more than 30% inhibition of digestive protease activity in rohu fingerlings respectively. A plot of the inhibition values obtained by changing the relative concentrations of enzyme/inhibitor resulted in different dose–response curves for different protein sources. The use of substrate gel electrophoresis allowed the visualization of the aforementioned differences in inhibition. Each seed extract produced a characteristic profile of protease inhibition. It is concluded that protease inhibitors present in plant protein sources adversely affect the digestive proteases in fish and hence there is a need to eliminate/reduce the amount of such inhibitors through proper processing before incorporation into aquafeeds.     

Antioxidant Properties and the Preservative Effect of Whole Radish Extract on Quality of Deccan Mahseer (Tor khudree) Steaks during Chilled Storage

ABSTRACT

The effect of dip treatment with solvent-free whole radish extract (WRE) on quality characteristics of Deccan mahseer steaks was evaluated during the chilled storage. In-vitro antioxidant activities of the extract were evaluated based on total phenolic and flavonoid contents, DPPH radical scavenging activity, and metal chelating ability prior to the dip treatment. The quality parameters, including total volatile base nitrogen (TVB-N), trimethylamine nitrogen (TMA-N), and free fatty acid (FFA), were significantly (p < .05) reduced in WRE treated steaks compared to the control (CT) steaks. WRE treatment of fish steaks significantly controlled the lipid oxidation parameters of thiobarbituric acid reactive substances (TBARS) and peroxide value (PV). Total plate count (TPC) of CT sample was higher than the treated sample and remained within the limit of 7 log₁₀ cfu/g at the end of storage. As per the sensory analysis, shelf life of Deccan mahseer was determined to be 15 days for WRE treated steaks and 9 days for control steaks during chill storage. Based on the results, it can be concluded that WRE treatment can limit the progress of fish spoilage bacteria in fish steaks and can be used as a substitute for synthetic additives.     

Strategies for maximizing utilization of de-oiled rice bran (DORB) in the fish feed

De-oiled rice bran (DORB) is obtained after oil is removed from the rice bran. This is the commonly used ingredient in the feeding of Indian major carps due to its cheaper cost. DORB is used as a feed ingredient in cattle, poultry, and fish diet. Along with this, they are also used in making fiber-rich biscuits and extraction of an antioxidant compound like oryzanol and other industrial uses. Due to the increased competition, there will be a huge imbalance in the demand and supply for this ingredient in the years to come. DORB is associated with high fiber content, anti-nutrients, and non-starch polysaccharides which lead to its poor utilization resulting in a higher feed conversion ratio (FCR) in fish. Hence, the strategic utilization of this ingredient by solid-state fermentation, exogenous enzyme supplementation, and supplementing it with deficient nutrients like amino acids and fatty acids will help in improving the utilization of this ingredient.

     

Anti-viral activity of methyl 1-chloro-7-methyl-2-propyl-1h-benzo[d] imidazole-5-carboxylate against white spot syndrome virus in freshwater crab (Paratelphusa hydrodromous)

Shrimp farming industries are subjected to severe economic loss due to a disease called white spot syndrome, a viral disease caused by white spot syndrome virus (WSSV) in penaeid shrimp. Numerous active compounds in the market possess anti-viral activity against the white spot syndrome virus, yet the issue remains unsolved. The present study was carried out to determine the anti-viral activity of methyl 1-chloro-7-methyl-2-propyl-1h-benzo[d] imidazole-5-carboxylate (C₁₃H₁₅N₂O₂Cl) against WSSV. The anti-viral activity of the synthetic compound was determined in freshwater crabs. Crabs were divided into three different experimental groups: healthy control groups (N.C.) received NTE buffer, positive control group (P.C.) crabs received WSSV, and treatment group crabs received WSSV along with synthetic weight compound. Experimental groups were observed for 30 days post-infection. Three different organs (gills, muscles, and head soft tissue (HST)) were dissected from all three groups and analyzed using molecular-based techniques, including polymerase chain reaction (PCR), Western blot, and histopathology. Clinical signs of WSSV were observed in the positive and N.C. groups; however, the treatment group showed a 100% survival rate. Confirmation was done using PCR, Western blot, and histopathology. These results demonstrated that the given synthetic compound has significant anti-viral activity against WSSV.

     

Growth and body composition of rohu (Labeo rohita) fed diets containing oilseed meals: partial or total replacement of fish meal with soybean meal

A 56‐day feeding trial was conducted to evaluate growth, feed utilization and body composition of fingerling rohu, Labeo rohita (4.5 ± 0.2 cm; 2.58 ± 0.04 g), fed five isonitrogenous (350 g kg^?1 crude protein) and isoenergetic (15.7 kJ g^?1 metabolizable energy) diets in which the main protein contributors were: diet I, fish meal (FM); diet II, soybean meal (SBM); diet III, groundnut meal (GNM); diet IV, equal amounts of SBM, GNM and canola meal (CM); diet V, equal amounts of FM, SBM, GNM and CM. Fish were stocked in triplicate groups of 25 fish held in 70 L flow‐through tanks (water volume 55 L) and fed twice daily (0900 and 1700 hrs) to apparent satiation. Fish fed diet I exhibited the highest (P < 0.05) values for live weight gain and specific growth rate (% per day). Feed conversion and protein efficiency ratio were better (P < 0.05) in fish‐fed diets I and II. Fish‐fed diet II exhibited lower (P < 0.05) fat and ash contents in carcass. Based on these findings, a 70‐day trial (Experiment II) was conducted with six diets fed under the same conditions where FM was incrementally replaced with SBM. Fingerling L. rohita (6.7 ± 0.4 cm; 4.07 ± 0.02 g) were used for the study. Crystalline l ‐methionine was added to diets containing more than 250 g kg^?1 SBM. This second study showed no significant (P > 0.05) variation in growth, feed utilization and carcass composition for fish fed the different diets. In conclusion, SBM was more effective than GNM or CM, as FM was replaced in feeds for fingerling L. rohita and could completely replace FM when supplemented with methionine and fortified with minerals under the conditions of this study.     

Genetic diversity in common carp stocks assayed by random-amplified polymorphic DNA markers

The common carp (Cyprinus carpio L.) is one of the major aquaculture species, contributing nearly 35% to the inland fish production in Karnataka, India. Stocks collected from Hungary (2), Indonesia and Vietnam were assessed alongside two local stocks in a series of culture performance trials with the objective of setting up a base population for developing a breeding programme. The present study deals with the genetic divergence and polymorphism in these six stocks using random‐amplified polymorphic DNA (RAPD) markers. A total of 180 decamer random primers were screened for polymerase chain reaction (PCR) amplification (OPA 1‐20, OPB1‐20, OPC1‐20, OPD1‐20, OPE1‐20, OPF1‐20, OPG1‐20, OPP1‐20 and OPM1‐20). Eight primers were selected for analysis of common carp genotypes (OPA‐7, OPA‐20, OPB‐17, OPF‐10, OP F‐9, OPG‐4, OPG‐9 and OPP‐16). Out of 492 bands recorded, 57.1% were polymorphic. Stepwise regression analysis was carried out to find best combination markers affecting body weight (P<0.001). The results demonstrate major differences in the genetic structures between different stocks. Dendrogram data showed grouping of individuals according to stocks and corresponding data variables revealed the per cent homology within the stock and also found markers correlating to the body weight.     

Studies on digestibility and digestive enzyme activities in Labeo rohita (Hamilton) juveniles: effect of microbial α-amylase supplementation in non-gelatinized or gelatinized corn-based diet at two protein levels

A 60-days feeding trial was conducted to delineate the effect of both gelatinized and non-gelatinized corn with or without supplementation with exogenous α-amylase at two dietary protein levels (35% and 28%) on dry matter digestibility, digestive enzymes and tissue glycogen content of Labeo rohita juveniles. Three hundred and sixty juveniles (average weight 10±0.15 g] were randomly distributed into 12 treatment groups with each of two replicates. Twelve semi-purified diets containing either 35% or 28% crude protein were prepared by including gelatinized (G) or non-gelatinized (NG) corn as carbohydrate source with different level of microbial α-amylase (0, 50, 100 and 150 mg kg⁻¹). The dry matter digestibility of G corn fed groups was significantly higher (P < 0.05) than that of the NG corn fed groups. Hepatosomatic index (HSI), liver glycogen and intestinal amylase activity of G starch fed groups were significantly higher (P < 0.05) than those of the NG corn fed groups. However, the reverse trend was found for gastrosomatic index (GSI), muscle glycogen and intestinal protease activity. Addition of 50 mg α-amylase kg⁻¹ feed improved the dry matter digestibility of NG starch fed groups, which was similar to that of the G corn fed groups or NG corn supplemented with 100/150 mg α-amylase kg⁻¹ feed. HSI, liver glycogen and intestinal amylase activity were significantly increased (P < 0.05) at minimum level of α-amylase in the feed (50 mg kg⁻¹) and did not increase due to further inclusion of amylase in the diet. Supplementation with α-amylase at 50 mg kg⁻¹ increased the intestinal amylase activity beyond which no significant changes were observed. Protease activity of liver and intestine was highest (P < 0.05) in higher crude protein (CP) fed groups, but protease activity of the intestine was significantly higher in the α-amylase supplemented groups. Hence, it was concluded that feed with 28% CP containing either G corn without α-amylase or NG corn with 50 mg α-amylase kg⁻¹ may be used as the alternative carbohydrate source for L. rohita juveniles.     

Non-gelatinized corn supplemented with α-amylase at sub-optimum protein level enhances the growth of Labeo rohita (Hamilton) fingerlings

A feeding trial was conducted for 60 days to delineate the effect of both gelatinized and non‐gelatinized corn with or without supplementation of exogenous α‐amylase at two level of protein 35% (optimum) and 27% (sub‐optimum) on growth, tissue composition and protein sparing effect of carbohydrate in the diet of Labeo rohita fingerlings. Three hundred and sixty fingerlings (average weight 10.00±0.15 g) were randomly distributed in 12 treatment groups with two replicates for each. Twelve semi‐purified diets containing either 35% or 27% CP were prepared with different levels of α‐amylase (0, 50, 100 and 150 mg kg^?1) and starch type [gelatinized (G)/non‐gelatinized, (NG)] viz. T₁ (NG, 35% CP, 0 mg kg^?1α‐amylase), T₂ (G, 35% CP, 0 mg kg^?1α‐amylase), T3 (NG, 27% CP, 50 mg kg^?1α‐amylase), T₄ (NG, 35% CP, 50 mg kg^?1α‐amylase), T₅ (G, 27% CP, 50 mg kg^?1α‐amylase), T₆ (G, 35% CP, 50 mg kg^?1α‐amylase), T₇ (NG, 27% CP, 100 mg kg^?1α‐amylase), T₈ (NG, 35% CP, 100 mg kg^?1α‐amylase), T₉ (G, 27% CP, 100 mg kg^?1α‐amylase), T₁₀ (G, 35% CP, 100 mg kg^?1α‐amylase), T₁₁ (NG, 27% CP, 150 mg kg^?1α‐amylase) and T₁₂ (NG, 35% CP, 150 mg kg^?1α‐amylase). The fish were acclimatized to the experimental condition for 24 days during which control diet (NG, 35% CP) was fed. Weight gain percentage, specific growth rate, protein efficiency ratio and apparent net protein utilization of NG corn‐fed groups at 42.43% were significantly higher (P<0.05) than the same level of G corn‐fed groups. We conclude that 42.43% NG corn supplemented with 50 mg α‐amylase kg^?1 feed at sub‐optimum level of CP (27%) significantly enhanced the growth and protein in L. rohita fingerling. Addition of dietary α‐amylase to G corn had no added advantage on growth of L. rohita fingerling.     

Comparative performance of Gamma Amino Butyric Acid (GABA) and 5-Hydroxytryptamine (5-HT) in the diet of larvae and post larvae of giant freshwater prawn, Macrobrachium rosenbergii: Effect of dose and route of administration on growth and survival

Three experiments were conducted to evaluate the effect of different dose and route of administration of 5-Hydroxytryptamine (5-HT) and Gamma Amino Butyric Acid (GABA) on growth, survival and pigmentation of giant freshwater prawn, Macrobrachium rosenbergii, larvae (experiment 1) and post larvae (experiment 2 and 3). In experiment 1 larva were stocked at 100 /L in seven different treatments with each of three replicates. The treatments were T1 (control), T2 (100 μg/ml of 5-HT bath exposure for 2 days), T3 (1 μM of GABA bath exposure for 2 days), T4 (0.5% 5-HT in feed), T5 (0.25% 5-HT in feed), T6 (0.5% GABA in feed) and T7 (0.25% GABA in feed). Highest growth, transformation rate, pigmentation and survival of larvae were recorded in T2 group in experiment 1. In experiment 2, post larvae were stocked at 70 per tank with 200 L of water for 45 days. This experiment consisted of 5 treatment groups with each of 3 replication viz. T1 (control), T2 (0.5% 5-HT in feed), T3 (0.25% 5-HT in feed), T4 (0.5% GABA in feed) and T5 (0.25% GABA in feed). At the end of this experiment growth was found to be higher in the control than the other treatment group showing inhibitory effect of 5-HT and GABA on growth of post larvae. To confirm the result, a third experiment was conducted for 30 days. Thirty numbers of post larvae having similar size were segregated from the previous experimental tanks (experiment 2) and stocked in the tanks of 100 L of water for experiment 3. All treatments were fed with control diet. There was similar growth pattern in all the treatments, which were higher (P < 0.05) than control, confirms the inhibitory effect of neurotransmitter in the diet of PL. Therefore, the overall results of the present study suggest that the bath treatment of neurotransmitter is superior to the dietary addition with regard to the growth, survival and pigmentation of M. rosenbergii larvae. 5-HT is more effective than the GABA for larvae of M. rosenbergii.     

Effect of inland water salinity on growth,feed conversion efficiency and intestinal enzyme activity in growing grey mullet, <Emphasis Type="Italic">Mugil cephalus</Emphasis> (Linn.): Field and laboratory studies

Two experiments were conducted to investigate the effect of inland water salinity on growth performance, feed conversion efficiency and intestinal enzyme activity in grey mullet. In experiment I, a 90 day monoculture of grey mullet at different salinity levels (0, 10, 15, 20 and 25%) was carried out. The fingerlings were stocked at 5000 per hectare and fed on a supplementary diet at 5% BW d^–1. This Study revealed that fish growth mean body weight (90.5 ± 4.5 g) and mean length (21.6 ± 0.4 cm), SGR (4.70%) and growth per day (0.99 g d^–1) were significantly (p < 0.05) enhanced in fish maintained at 10% salinity in comparison with other treatments. Nutrient levels, phytoplankton population, NPP and chlorophyll a all decreased with an increase in salinity (>10%). In addition, zooplankton populations increased with an increase in the salinity level. Most of the other hydrochemical characteristics remained at optimal levels in all other treatments. Fish weight gain showed a significant positive correlation with productivity indicating parameters viz. alkalinity (r = 0.53), turbidity (r = 0.62), NPP (r = 0.75) and chlorophyll a (r = 0.46), clearly revealing that fish growth is also related to the trophic status of the ponds. In the second experiment (Experiment II), mullet fry were exposed to five different salinity levels (10, 15, 20, 25 and 30%) and maintained for 70 days in the laboratory. Significantly (p < 0.05) high growth, (SGR and per cent increase in body weight), feed conversion efficiency and intestinal enzyme activity were observed in the group maintained at 10 salinity in comparison with other groups maintained at similar salinity levels. Carcass composition, musc1e and liver glycogen levels were also significantly (p < 0.05) affected by salinity changes.