Effect of leaves and seeds of Achyranthes aspera as feed supplements on the immunological and stress parameters and related gene expressions of Asian catfish (Clarias batrachus).

The immunostimularory properties of Achyranthes aspera leaves and seeds supplemented feeds were evaluated in Asian catfish, Clarias batrachus (0.352±0.008 g). The experimental feeds contained 0.25% leaves (EFL1), 0.5% leaves (EFL2), 0.5% seeds (EFS) and 0% leaves or seeds (control feed; CF). In CF, leaves and seeds were absent. Fish were immunized with chicken-RBC after 60 days of feeding. The blood and tissue samples were collected on 7th, 14th and 21st days after immunization for various assays. The average weight of magur was significantly higher in EFS compared to the other treatments throughout the study period and EFL2 followed this group. Serum lysozyme level of fish was significantly higher in EFS on 7th and 14th days and in EFS and EFL2 on 21st day after immunization compared to the other treatments. Myeloperoxidase and nitric oxide synthase levels were always significantly higher in EFS diet fed fish compared to other treatments. The highest hemagglutination titer level was found in EFS throughout the study period. Thiobarbituric acid reactive substances (TBARS) and carbonyl protein levels were significantly lower in liver and kidney of enriched diets fed magur compared to CF treatment. TBARS and carbonyl protein levels were minimum in EFS diets fed fish. In EFS and EFL2 treatments, the expressions of TNF-α, iNOS and NF-kB were significantly higher compared to the CF group. A. aspera seeds and leaves showed significant immunostimulatory properties in Asian catfish fry.

Freshwater Macrophytes: A Potential Source of Minerals and Fatty Acids for Fish, Poultry, and Livestock.

The freshwater macrophytes are abundant in tropical and subtropical climates. These macrophytes may be used as feed ingredients for fish and other animals. The nutritional value of twelve freshwater-cultured macrophytes was evaluated in the present study. Significantly higher crude protein (36.94-36.65%) and lipid (8.13-7.62%) were found in Lemna minor and Spirodela polyrhiza; ash content was significantly higher in Hydrilla verticillata, Wolffia globosa, and Pistia stratiotes (20.69-21.00%) compared with others. The highest levels of sodium, magnesium, chromium, and iron levels were recorded in P. stratiotes. H. verticillata was a rich source of copper, manganese, cobalt, and zinc; the contents of calcium, magnesium, strontium, and nickel were highest in S. polyrhiza. Selenium and potassium contents were higher in Salvinia natans and W. globosa, respectively. The n-6 and n-3 polyunsaturated fatty acids (PUFAs) contents were significantly higher in W. globosa and Ipomoea aquatica, respectively compared with others. Linoleic and α-linolenic acids were dominant n-6 and n-3 PUFAs. The highest value (4.04) of n-3/n-6 was found in I. aquatica. The ratio ranged from 0.61 to 2.46 in other macrophytes. This study reveals that macrophytes are rich sources of minerals, n-6 and n-3 PUFAs.

Evaluation of UV-B protective properties of leaves and seeds of Achyranthes aspera in Asian catfish Clarias batrachus (Linn.).

The ultraviolet-B (UV-B) radiation is harmful to the aquatic organisms. The UV-B protective properties of leaves and seeds of herb Achyranthes aspera were evaluated in Clarias batrachus. Fish were fed with four diets-EFL1, EFL2 containing 0.25 and 0.5% leaves, EFS containing 0.5% seeds and control, CF. After 83 days of feeding, fish were exposed to UV-B (157 µW/cm2) for 7 days at the rate of 15 min/day. One batch of fish in each treatment was kept unexposed. Significantly higher final weight was found in EFS followed by EFL2 and EFL1 treatments. It was higher in unexposed fish compared to the exposed ones. Among exposed fish, significantly higher lysozyme was found in EFS and myeloperoxidase in EFS and EFL2 compared to others. Nitric oxide synthase and superoxide dismutase levels were significantly higher in liver and head kidney of EFS diet fed fish compared to others. Thiobarbituric acid reactive substances (TBARS) and carbonyl protein levels were minimum in EFS followed by EFL2. The independent sample t-test showed that nitric oxide synthase was significantly higher and myeloperoxidase and TBARS levels were significantly lower in unexposed group compared to the exposed fish in respective treatment. There were up-regulations of TNF-α, iNOS, NF-kB, BAX, Cytochrome c, SOD-c, Caspase 3, Caspase 9, BCL2 in liver and head kidney of leaves and seeds incorporated diets fed fish compared to control. Supplementation of A. aspera seeds and leaves at 0.5% level in diets gave UV-B protection to the fish.

Effect of Lemna minor supplemented diets on growth, digestive physiology and expression of fatty acids biosynthesis genes of Cyprinus carpio.

The potential nutritional value of duckweed Lemna minor (Lemnaceae) was evaluated for common carp Cyprinus carpio fry. Fish were fed diets containing five graded levels of duckweed: 0% (LM0, control), 5% (LM5), 10% (LM10), 15% (LM15) and 20% (LM20). The final weight and specific growth rate were significantly higher in LM15 and LM20 diets fed fish compared to others. Feed conversion ratio was minimum in fish fed diet LM20. Amylase activity was significantly higher in LM0 treatment. Total protease, trypsin and chymotrypsin activities showed linear relationships with the increased level of duckweed in the diet. Protein and essential amino acids contents were significantly higher in carp fed diets LM15 and LM20 compared to others. Lipid content was significantly higher in fish fed duckweed-based diets compared to control. A direct relationship was found between the inclusion level of duckweed in the diet and n-3 long-chain polyunsaturated fatty acid (LC-PUFA) content of carp. Contents of desaturated and elongated products of dietary linolenic acid (18:3n-3) including 20:4n-3, 20:5n-3, 22:5n-3 and 22:6n-3 increased in a graded manner with increasing dietary duckweed. The monounsaturated fatty acids and n-6 PUFA contents reduced significantly in fish fed duckweed. Expression of fads2d6, elovl2, elovl5 and fas were higher in carp fed diets LM10, LM15 and LM20 compared to control fish. The inclusion of L. minor in diet enhanced the nutritional value of carp by increasing protein, lipid, amino acids and n-3 PUFA contents.

The impact of Achyranthes aspera seeds and leaves supplemented feeds on the survival, growth, immune system and specific genes involved in immunostimulation in Clarias batrachus fry challenged with Aeromonas hydrophila in pond conditions.

The present study was conducted to evaluate the effect of dietary inclusion of Achyranthes aspera seeds and leaves on the immune system of magur Clarias batrachus challenged with Aeromonas hydrophila in pond conditions. Magur fry (0.51 ± 0.032 g) were cultured in hapas set inside a pond and were fed with three feeds. Two experimental feeds FS1 and FS2 were supplemented with 0.5% seeds and leaves of A. aspera, respectively and FC3 was the control one. After 90 days of feeding, fish were challenged with A. hydrophila. In FC3, 70% fish died within 48 h of challenge, while 25 and 30% mortality were recorded in FS1 and FL2, respectively. The cumulative mortality rates were 70, 45 and 35% in FC3, FL2 and FS1, respectively. The average weight and specific growth rate of magur were significantly higher in FS1 compared to others. Serum lysozyme, myeloperoxidase, nitric oxide synthase and superoxide dismutase levels were significantly higher in FS1 compared to others. Thiobarbituric acid reactive substances and carbonyl protein levels were significantly lower in FS1 compared to others. In liver and head kidney of FS1 and FS2 fed magur, the iNOS, SOD-C, TNF-α, Cytochrome c, Caspase 9 were up-regulated. Caspase 3 was also significantly up-regulated in FS1 and it was followed by FL2 treatment. A. aspera incorporated feeds improved the immune system of fish and gave protection against bacteria even in the pond conditions.

Effect of temperature on food consumption, immune system, antioxidant enzymes, and heat shock protein 70 of Channa punctata (Bloch, 1793).

The impact of water temperature on the physiology of Channa punctata (Bloch, 1793) was evaluated in the present study. Fish were acclimated at 25 ± 1 °C and then exposed at six different temperatures: 10, 15, 20, 25, 30, and 35 °C. C. punctata exposed at 10, 15, and 20 °C showed 30, 21, and 11% reduced food consumption, respectively compared to 25 °C. Significantly higher respiratory burst and myeloperoxidase activities were found in fish exposed at 20 and 25 °C after 12 h of exposure compared to other treatments. Nitric oxide synthase was significantly higher at 25 °C after 12 h and at 25 and 30 °C exposed fish after 7 days compared to others. The reduced glutathione level was significantly higher at 25 °C compared to other treatments after 7 days of exposure. The thiobarbituric acid reactive substances level was minimum at 25 °C. Significantly lower antioxidant enzymes, catalase, glutathione peroxidase, and glutathione S-transferase were found in gills of fish exposed at 25 °C compared to others in both samples. The highest antioxidant enzyme levels were found at 10 °C. Heat shock protein (Hsp) 70 levels were significantly lower in liver and muscle of fish exposed at 25 °C compared to other treatments. The Hsp level was significantly higher at 35 and 30 °C exposed fish compared to others after 12 h, and the level reduced after 7 days in these treatments. Thermal stress affects food consumption rate, immune system, antioxidant enzymes, and enzyme systems in fish. The elevated Hsp70 level serves as a biomarker of stress in C. punctata. Graphical Abstract.

Effect of light intensity on survival, growth and physiology of rohu, Labeo rohita (Cyprinidae) fry.

Purpose: Light intensity is one of the important environmental factors that have strong influence on larviculture. Culture of fish at high intensity of light generates harmful radicals in the body that can compromise their health and production. The present study aims to evaluate the effect of various light intensities on the physiology of rohu Labeo rohita fry.Materials and Methods: Rohu fry (13.56 ± 0.4 mg) were exposed at five different light intensities: 0.17 ± 0.005 (Lc, control), 1.45 ± 0.23 (L1), 2.69 ± 0.47 (L2), 3.93 ± 0.72 (L3) and 5.06 ± 0.95 Wm-2 (L4). After 90 days of culture, rohu were harvested.Results and Conclusions: A 2-5% mortality of rohu was recorded in L3 and L4 treatments. The average weight and specific growth rate were significantly (p < .05) higher in Lc treatment compared to others. The light intensity and swimming activity of rohu showed direct relationship, whereas, light intensity showed inverse relation with nitric oxide synthase and reduced glutathione levels. Significantly (p < .05) higher glutathione S-transferase and glutathione peroxidase activies were found in rohu exposed at L4 treatment. Higher light intensities resulted in oxidative stress in the muscles of rohu. Thiobarbituric acid reactive substances, carbonyl protein and heat shock protein 70 were significantly (p < .05) higher in rohu exposed at L4 compared to other treatments. Exposure of rohu fry to intense light resulted into physiological stress and immunosupression.

The study of ameliorative effect of dietary supplementation of vitamin C, vitamin E, and tryptophan on Labeo rohita (Cyprinidae) fry exposed to intense light.

The stress ameliorating effect of dietary supplementation of vitamin C, vitamin E, and tryptophan on rohu Labeo rohita fry was evaluated. Rohu fry (1.1 ± 0.03 g) were cultured under five different feeding regimes: enriched with 0.08% vitamin C (D1), 0.02% vitamin E (D2), 1.42% tryptophan (D3), a combination of these three ingredients at similar doses (D4), and control diet (D5). Rohu fry of D5 were divided into two groups-exposed to experimental light (D5FL) and ambient light (114 ± 4 lx, D5AL). All fry (except D5AL) were exposed at light intensity of 3442 ± 648 lx. Feeding of rohu with enriched diets significantly (P < 0.05) enhanced the survival rate and average weight. A 15-25% higher survival and 1.3-1.8-fold higher average weight were recorded in rohu fed with enriched diet compared to D5FL treatment. Supplementation of vitamin C in diet (D1) of rohu resulted in 4.1-fold and 6.9-fold higher nitric oxide synthase and reduced glutathione (GSH) levels, respectively compared to the D5FL treatment. The tryptophan-enriched diet (D3) showed 5.8-fold higher melatonin and 4.4-fold lower cortisol levels in rohu compared to the D5FL treatment. Significantly (P < 0.05) higher nitric oxide synthase, GSH and melatonin, and lower cortisol, glucose, thiobarbituric acid reactive substances, carbonyl protein, glutathione S-transferase, and glutathione peroxidase levels were found in D4 diet fed rohu compared to the other treatments. Reduced level of stress in D4 treatment resulted in best performance of rohu in terms of less swimming activity and higher survival and growth compared to the other treatments.

Validation of growth enhancing, immunostimulatory and disease resistance properties of Achyranthes aspera in Labeo rohita fry in pond conditions.

The immunostimulatory and disease resistance properties of herb Achyranthes aspera L. (Amaranthaceae) were evaluated in rohu Labeo rohita in pond. Rohu fry (1.9 ± 0.08 g) were cultured in hapas (25 hapa-1) set inside a pond and were fed with two experimental diets containing 0.5% seeds (D1) and leaves (D2) of A. aspera and control diet (D3). Fish were challenged with Aeromonas hydrophila after 80 days. The cumulative mortality rate of fish was significantly (P < 0.05) higher in D3 (28-48%) compared to others. Average weight was significantly (P < 0.05) higher in D1 (6.5-12.5%) compared to other treatments. Myeloperoxidase and nitric oxide synthase levels were significantly (P < 0.05) higher in D1 and D2 compared to D3. Thiobarbituric acid reactive substances and carbonyl protein levels were significantly (P < 0.05) lower in hepatopancreas and kidney of D1 compared to others. In hepatopancreas, the expressions of lysozyme C, loysozyme G, TNF-α, IL-10 and IL-1ß were significantly (P < 0.05) higher in D1 compared to others. This treatment was followed by D2. In kidney, lysozyme G and TNF-α levels were significantly (P < 0.05) higher in D1 and D2 compared to D3. Whereas, IL-10 and IL-1ß were significantly (P < 0.05) down-regulated and up-regulated, respectively in kidney of D2. There was up-regulation (P < 0.05) of TLR-4 in hepatopancreas and kidney of D1 and D2 diets fed rohu, respectively compared to others.

Oxygen stress: impact on innate immune system, antioxidant defence system and expression of HIF-1α and ATPase 6 genes in Catla catla.

Catla catla catla (2.28 ± 0.1 g) were exposed to six different levels of dissolved oxygen: 1 (DO-1), 3 (DO-3), 5 (DO-5), 7 (DO-7), 9 (DO-9) and 11 (DO-11) mg/L. DO-5 served as control. In DO-1 and DO-3, the number of red blood cells (RBC), lysozyme, respiratory burst activity and nitric oxide synthase were significantly (p < 0.05) lower compared to the control one. In DO-7 and DO-9, RBC and lysozyme were significantly (p < 0.05) higher compared to the control one. Thiobarbituric acid reactive substances was significantly (p < 0.05) higher in catla exposed at low (1 and 3 mg/L) and high (9 and 11 mg/L) dissolved oxygen compared to others. In muscles and hepatopancreas, reduced glutathione was significantly (p < 0.05) higher in DO-5 and DO-7 and in gills of DO-5 compared to others after 1 h. In muscles, glutathione S-transferase (GST) was significantly (p < 0.05) lower in DO-5 and DO-7 compared to others. In hepatopancreas, GST and glutathione peroxidise (GPx) were significantly (p < 0.05) higher in DO-1 and DO-3 compared to others. In gills, GPx was higher in DO-9 and DO-11 after 48 h. In brain, hypoxia-inducible factor (HIF)-1α mRNA level was induced in DO-1 and DO-3 compared to others after 1 h of exposure. In gills and hepatopancreas, HIF-1α mRNA level was significantly (p < 0.05) higher in DO-1 compared to others after 1 h. The ATPase 6 mRNA level was significantly (p < 0.05) higher in brain and hepatopancreas of DO-1 after 1 h and in gills and hepatopancreas of DO-3 and DO-9, respectively, after 48 h compared to others.

Biological mosquito control is affected by alternative prey.

BACKGROUND: Mosquitofish were introduced to several countries of the tropics and subtropics as biological agents for the control of mosquito larvae. Meanwhile, they became a threat to native communities and fish worldwide, similar to other invasive species through resource competition, overexploitation, or habitat alteration. We investigated prey selectivity patterns of Gambusia affinis (mosquitofish) preying on larvae of the two Indian major carps (Catla catla and Labeo rohita) in the presence of varied proportions of alternative prey (rotifers, cladocerans, chironomid and mosquito larvae) under laboratory conditions. RESULTS: The patterns of prey selectivity in mosquitofish were influenced by the presence of alternative prey and theirrelative abundance in the environment. Carp larvae, when present in equal proportions, were randomly selected by mosquitofish, however, positively selected when present in higher proportions.Inthe presence of Hexarthramira, Daphnia similoides or the mosquito larval instar-IV as an alternative prey, the mosquitofish preferred fish larvae regardless of prey proportions. In the medium where either mosquito larval instar-I or chironomid larvae were givenas alternative prey, the mosquitofish either rejected or randomly selected the carp larvae. Given a multispecies prey combination, mosquitofish primarily selected the larvae of L.rohita and mosquito larval instar-I. We also found a prey switching ability of mosquitofish in relation to varying abundances of prey species in the environment. CONCLUSIONS: Thepresent results suggest that mosquito immatures are not the preferred food of mosquitofish when fish larvae are present in their natural habitats. Since mosquitofish and carp larvae have overlapping natural habitats and prey preferences are the invasive mosquitofish may have a substantial impact on native communities of invertebrates and fish. This way, they are equally important for extensive fisheries and conservation management.

Effect of seeds of Achyranthes aspera on the immune responses and expression of some immune-related genes in carp Catla catla.

Evaluation of functional mechanism of immunostimulant in fish is essential for realizing their therapeutic potential. The impact of dietary supplementation of seeds of Achyranthes aspera on the immune system of Catla catla was studied. Catla (37.73 ± 6.7 g) were fed with test diets containing 0.5 (D1) and 1.0% (D2) seeds of A. aspera and control diet (D3) for 50 days. Then fish were immunized with heat-killed Aeromonas hydrophila. Blood and tissue samples were collected after 7 days of immunization. Three fish were sacrificed for each parameter. Respiratory burst activity, bactericidal activity and antibody response were significantly (P < 0.05) higher in test diets fed catla compared to the control one. Nitric oxide synthase and serum lysozyme were significantly (P < 0.05) higher in fish fed with diet D2 compared to the other treatments. There were up-regulations of TNFα, lysozyme c and g gene expressions in kidney and IL-10, lysozyme c and g expressions in hepatopancreas of test diets fed fish compared to the control one. The expression of IL-10 was down-regulated in the kidney of D2 diet fed fish compared to others. In the gill, significantly (P < 0.05) higher expressions of TNFα was found in D1 diet feed fish and lysozyme c and in D2 diet fed fish compared to the other groups. This study showed tissue-specific gene expression pattern in catla. An inverse relationship was found between the expression of TNFα and IL-10 in kidney of catla.

Impact of UV-B radiation on the digestive enzymes and immune system of larvae of Indian major carp Catla catla.

PURPOSE: Ultraviolet radiation is a potent threat to the aquatic animals. Exposure to such stressor affects metabolic and immunological processes. The present investigation aims to study the effect of UV-B radiation on digestive enzymes and immunity of larvae of Catla catla. MATERIALS AND METHODS: Larvae were exposed to ultraviolet-B (UV-B, 280-320 nm) radiation (145 microW/cm(2)) for three different exposure times of 5, 10 and 15 min on every other day. After 55 days, important digestive enzymes were assayed. For immunological study, lysozyme, glutamate oxaloacetate transaminase (GOT) and glutamate pyruvate transaminase (GPT) levels were measured. Then the fish were kept for one month without radiation and lysozyme level was measured. RESULTS: Protein concentration varied directly with the duration of exposure and was highest among fish that had received the 15 min UV-B irradiation. Significantly higher amylase, protease, trypsin and chymotrypsin activities were found in 5 min exposed fish compared to others. Lysozyme level was significantly higher in control group compared to the UV-B treated fish. The lysozyme level decreased with the increasing duration of UV-B radiation. When fish were kept without UV-B radiation for one month, lysozyme level was brought to the normal level in all treatments, except 15 min exposed fish. The GOT and GPT levels were significantly higher in the 15 min exposed group than others. CONCLUSIONS: The effects of UV-B radiation on the digestive physiology and immune system of catla have been clearly observed in the present study. The decreased enzyme activities in UV-B radiated fish results into improper digestion and poor growth.