Impact of feeding biochar sources in rohu (Labeo rohita): Evaluating the growth, nutrient absorption, carcass composition, haematology and mineral status.

Biochar, an organic carbonaceous matter, is a unique feed additive that is now being used in aquaculture industry to formulate a cost-effective and eco-friendly diet. This experiment (in door) was conducted over course of 90 days to determine the most effective form of biochar, produced from various sources, for supplementation in Moringa oleifera seed meal-based diet. These sources were: farmyard manure biochar, parthenium biochar (PB), vegetable waste biochar, poultry waste biochar (PWB) and corncob waste biochar, added at 2 g/kg concentration to determine the effect of supplementation on the growth indices, nutrient absorption, carcass composition, haematology and mineral status of Labeo rohita (rohu) fingerlings. The research design consisted of six test diets with three replications (6 × 3) of each. Total of 270 fingerlings (6.30 ± 0.020 g) were fed at 5% body weight and 15 of them were kept in separate steel tanks. The results indicated that PWB was most effective in improving weight gain (285.58 ± 4.54%) and feed conversion ratio (1.060 ± 0.040) compared to control diet and other test diets. The same type of biochar (PWB) produced the best results for nutrient digestibility, that is, crude protein, crude fat and gross energy and carcass composition. In terms of haematology and mineral status, PWB showed the best results. In conclusion, it was found that PWB significantly enhanced (p < 0.05) L. rohita fingerling's growth, carcass composition, nutrient digestibility, haematological parameters (red blood cells, white blood cells, platelets and haemoglobin) and mineral composition (Ca, Na, P, Mg, Fe, Mn, Zn, K and Cu) whereas PB negatively affected all parameters. It is anticipated that the potential use of biochar will increase in aquaculture industry, as research on its incorporation in fish feeds is still limited.

Therapeutic Role of Hemp (Cannabis sativa) Against Copper-Induced Toxicity in Labeo rohita and Cirrhinus mrigala.

For decades hemp has been used as a therapeutic agent for enhancing immunity in animals. Current study was conceptualized to find out the protective role of dietary hemp seed products (hemp seed oil (HO) and hemp seed (HS)) against copper-induced toxicity in fish. Fingerlings of Labeo rohita (Rohu) and Cirrhinus mrigala (Mrigal) were exposed to copper at 20% of the 96 h LC50 (1.34 and 1.52 ppm, respectively) for 30 days. Following Cu exposure, fish were maintained on two types of hemp (Cannabis sativa)-supplemented feeds, on graded levels of hemp seed oil (HO: 1%, 2%, 3%) and hemp seed (HS: 5%, 10%, 15%) for 50 days, while one group was the control (without any copper exposure as well as any supplementation). Copper exposure significantly increased (P < 0.05) WBCs, hematocrit, MCHV, eosinophils, and lymphocytes in L. rohita and also in C. mrigala as compared to control. Copper exposure also significantly (P < 0.05) changed lysozymes, plasma protein, and IgM in both species, in comparison to control. Moreover, alkaline phosphatase, bilirubin, serum glutamic-pyruvic transaminase, and aspartate transaminase were significantly (P < 0.05) changed by copper exposure in comparison to control in both species. Additionally, Antioxidant enzymes like catalase, superoxide dismutase, glutathione reductase, and glutathione peroxidase were also significantly (P < 0.05) increased in the brain, gills, liver, and muscle of copper-exposed group in both species as compared to control. Interestingly, all the altered parameter of blood, serum, liver function tests, and antioxidant enzymes (in different organs) because of copper toxicity were successfully reverted to normal level in hemp seed oil (HO) and hemp seed (HS)-supplemented fed groups of both species. In conclusion, hemp seed supplementation showed significant (P < 0.05) improved results against copper toxicity. Thus, it could be recommended as an animal feed ingredient for its therapeutic role.

Potentials of Aloe barbadensis inclusion in fish feeds on resilience to Aeromonas hydrophila infection in freshwater fish Labeo rohita.

Aquatic bacterial pathogens can cause severe economic loss in aquaculture industry. An opportunistic pathogen, Aeromonas hydrophila is responsible for Motile Aeromonas Septicemia, leading to high mortality rates in fish. The present study was focused on the efficacy of Aloe barbadensis replacing fishmeal diets on hematological, serum biochemical, antioxidant, histopathological parameters, and disease resistance against A. hydrophila infection in Labeo rohita. Isonitrogenous fishmeal replaced diets (FMR) were prepared with varying levels of A. barbadensis at D1 (0%) (control), D2 (25%), D3 (50%), D4 (75%) and D5 (100%) then fed to L. rohita. After 60 days of post-feeding, the experimental fish were challenged with A. hydrophila. Blood and organs were collected and examined at 1- and 15-days post infection (dpi). The results demonstrated that on 1 dpi, white blood cells (WBC), total protein, cholesterol and low-density lipoprotein (LDL) levels were significantly increased in D3 diet fed groups. The D2 and D3 diet fed group showed decreasing trends of serum glutamic pyruvic transaminase (SGPT) and antioxidant enzymes activity on 15 dpi. The histopathological architecture results clearly illustrated that the D3 diet fed group had given a higher protective effect by reducing the pathological changes associated with A. hydrophila infection in liver, intestine and muscle. Higher percentage of survival rate was also observed in D3 diet fed group. Therefore, the present study suggested that the dietary administration of A. barbadensis up to 50% fishmeal replacement (D3 diet) can elicit earlier antioxidant activity, innate immune response and improve survival rate in L. rohita against A. hydrophila infection.

Role of Citrullus colocynthis and Psidium guajava Mediated Green Synthesized Silver Nanoparticles in Disease Resistance against Aeromonas hydrophila Challenge in Labeo rohita.

Green synthesis of metallic nanoparticles is an auspicious method of preparing nanoparticles using plant extracts that have lesser toxicity to animal cells and the host. In the present work, we analyzed the antibacterial activity of Citrullus colocynthis and Psidium guajava-mediated silver nanoparticles (Cc-AgNPs and Pg-AgNPs, respectively) against Aeromonas hydrophila (A. hydrophila) in an in vivo assay employing Labeo rohita (L. rohita). L. rohita were divided into six groups for both Cc-AgNPs and Pg-AgNPs treatments separately: Control, A. hydrophila infected, A. hydrophila + Ampicillin, A. hydrophila + Cc/Pg-AgNPs (25 µg/L), A. hydrophila + Cc/Pg-AgNPs (50 µg/L), and A. hydrophila + Cc/Pg-AgNPs (75 µg/L). Changes in different bio-indicators such as hematological, histological, oxidative stress, and cytokine analysis were observed. Interestingly, the infected fish treated with both types of AgNPs (Cc-AgNPs and Pg-AgNPs) exhibited a higher survival rate than the untreated infected fish and demonstrated signs of recovery from the infection, providing a compelling indication of the positive impact of phytosynthesized AgNPs. Disruptions in hematological and histological parameters were found in the infected fish. Both Cc-AgNPs and Pg-AgNPs showed recovery on the hematological and histological parameters. Analysis of oxidative stress and cytokine markers also revealed provoking evidence of the positive impact of Cc-AgNPs and Pg-AgNPs treatment against disease progression in the infected fish. The major finding of the study was that the higher concentrations of the nanoparticles (50 µg/L in the case of Cc-AgNPs and 75 µg/L in the case of Pg-AgNPs) were more effective in fighting against disease. In conclusion, our work presents novel insights for the use of green-synthesized AgNPs as economic and innocuous antibacterial candidates in aquaculture.

Complimentary effect of exogenous enzymes, essential amino acids and essential fatty acids supplemented de-oiled rice bran (DORB) based diets on hematology, liver and intestinal histoarchitecture in Labeo rohita.

A 60-day feeding trial was conducted to study the hematology, liver, and intestinal histoarchitecture of Labeo rohita fed with a combination of exogenous enzymes, essential amino acids, and essential fatty acids to DORB (De-oiled rice bran) based diets. Three treatments viz., T1 [DORB + phytase and xylanase (0.01% each)], T2 [DORB + phytase (0.01%) + xylanase (0.01%) + L-lysine(1.4%) + L-methionine (0.4%) + EPA and DHA (0.5%)] and T3 [DORB + phytase (0.01%), xylanase and cellulase (0.075%) + L-lysine (1.4%) +L-methionine (0.4%) + EPA and DHA (0.5%)] were used in the present study. Serum total protein, albumin content and A/G ratio varied significantly (p < 0.05) among groups. Globulin content did not vary significantly among groups (p ≥ 0.05). The Hb content, RBC and MCV count varied significantly (p < 0.05) whereas MCH, MCHC content, WBC and lymphocyte count did not vary significantly among groups (p > 0.05). The liver and intestine examination revealed no visible alteration and showed normal histo-architecture. Based on the finding it is concluded that DORB supplemented with exogenous enzymes, essential amino acids and essential fatty acids with phytase (0.01%), xylanase and cellulase (0.075%), L-lysine (1.4%), DL-methionine (0.4%) and EPA and DHA (0.5%) improves the health of L. rohita.

An insight into the interaction between Argulus siamensis and Labeo rohita offers future therapeutic strategy to combat argulosis.

Aquaculture and fisheries are salient flourishing sectors in the world but their sustainability is often afflicted by several pathogenic diseases. Among all the pathogenic diseases of fish, parasitic diseases are found to be a major cause of concern. Argulosis is one of the dominant parasitic problems encountered in Indian aquaculture practices. Argulus siamensis is the most prevalent argulid species harming the Indian major carp species including Labeo rohita. The major carps respond to parasitic infestation by elevating various immune relevant genes. The therapeutic chemicals, synthetic drugs and other plant extracts have made a progress in the fight against argulosis. However, there is no effective vaccine and drugs are available for this disease. Thus, designing efficient, cost-effective and eco-friendly control and treatment strategies for argulosis is presently needed. Keeping the aforementioned facts in mind, the current review elaborated the immunological interaction between A. siamensis and L. rohita, available combat tactics, highlighted the already identified vaccine candidates to design effective control measures and illustrated the use of omics technology in future to combat argulosis.

Withania coagulans fruit extract: a possible useful additive in ameliorating growth and immunity of Labeo rohita (Hamilton, 1822).

This study aimed to assess the Withania coagulans fruit extract's effects on growth and haematological parameters of Labeo rohita. Healthy fish (n = 120) were divided in four groups. Experimental diet was prepared with different extraction of W. coagulans (0%, 1%, 1.5% and 2%). Diet was provided for eight weeks to all groups except the control group which fed on basal diet. The 1.5% and 2% W. coagulans extracts lead improvement in feed conversion ratio and specific fish growth rate; haematological parameters in fish group fed with 2% of W. coagulans showed increased in WBC, RBC, Hb, MCHC, PCV. The values of plasma proteins and lysozyme activity were higher in the experimental group compared control group. Fish fed with W. coagulans extracts showed increased of immunity and less mortality respect to others. It suggested that W. coagulans could be considered valuable support for the improve growth and immunity in Labeo rohita.

Limiting amino acids supplementation in low crude protein diets and their impacts on growth performance and carcass composition in Labeo rohita (rohu) adult fish / Limitação da suplementação de aminoácidos em dietas com baixo teor de proteína bruta e seus impactos no desempenho de crescimento e composição da carcaça em peixes adultos Labeo rohita (rohu)

Abstract Ninety days study was conducted in hapas installed in earthen ponds. Fish of an average initial weight (220g) were evenly distributed in triplicate groups within fifteen hapas. Five experimental diets labeled as T1 (25% CP and NRC recommended amino acid level) as control diet, T2 (with 2% low protein and 5% amino acid supplementation), T3 (with 2% low protein and 10% amino acid supplementation), T4 (with 4% low protein and 10% amino acid supplementation) and T5 (with 4% low protein and 20% amino acid supplementation) were prepared. Fish were fed with @3% of their body weight twice a day at 10.00 & 16:00 hour. Significantly higher percent weight gain (420.18 ± 66.84a) and specific growth rate (13499.33±1273.54a) along with improved feed conversion ratio (1.29 ± 0.09b) and hundred percent survivals were recorded during the trial. Furthermore proximate analysis of meat showed significant improvement in the crude protein level (81.77 ± 0.19a) served with diet containing 20% limiting amino acids mixture. Therefore, limiting amino acids can be a source of cost effective feed and use safely in L. rohita diet.

Resumo O estudo de 90 dias foi realizado em hapas instalados em tanques de terra. Peixes com peso inicial médio (220 g) foram distribuídos uniformemente em grupos triplicados em 15 hapas. Cinco dietas experimentais rotuladas como T1 (25% de CP e NRC recomendado nível de aminoácidos) como dieta controle, T2 (com 2% de proteína baixa e 5% de suplementação de aminoácidos), T3 (com 2% de proteína baixa e 10% de suplementação de aminoácidos), T4 (com 4% de baixa proteína e 10% de suplementação de aminoácidos) e T5 (com 4% de baixa proteína e 20% de suplementação de aminoácidos) foram preparadas. Os peixes foram alimentados com 3% do seu peso corporal duas vezes por dia às 10h00 e 16h00. Ganho de peso significativamente maior (420,18 ± 66,84a) e taxa de crescimento específico (13499,33 ± 1273,54a) juntamente com taxa de conversão alimentar melhorada (1, 29 ± 0,09b) e sobrevivência de cem por cento foram registrados durante o ensaio. Além disso, a análise aproximada da carne mostrou melhora significativa no nível de proteína bruta (81,77 ± 0,19a) servida com dieta contendo 20% de mistura de aminoácidos limitantes. Portanto, a limitação de aminoácidos pode ser uma fonte de alimentação econômica e usada com segurança na dieta de L. rohita.

Impact of dietary leaf extracts of Black pepper Piper nigrum L on the growth, hematological and immunological parameters of Labeo rohita (Hamilton, 1822) cultured in glass aquaria / Impacto de extratos de folhas da dieta de pimenta-do-reino Piper nigrum L sobre o crescimento, parâmetros hematológicos e imunológicos de Labeo rohita (Hamilton, 1822) cultivado em aquários de vidro

A study was conducted to evaluate the effect of Piper nigrum (black pepper) leaf extract on on the growth performance, proximate composition, hematological parameters, and immune response of Labeo rohita fingerlings with an average weight of 22.14 ± 0.98g. After acclimation for two weeks, fish (n=25) were randomly selected and placed in four glass aquaria (T0, T1, T2 and T3) at constant water temperature (30.0 ± 1.0 °C), pH (7.50 ± 0.5) and total hardness (200 ± 2.0 mgL,-¹) for a period of 12 weeks, with three replicates each. Fish were fed with P. nigrum leaf extract supplemented feed @ 0.0%, 1.0%, 2.0% and 3.0% in T0, T1, T2 and T3, respectively. At the end of experiment, five fish were randomly selected from each aquaria for proximate composition, gut and skin microbial load, hematological parameters. Total proteins, albumins, and globulins were also recorded to evaluate immunological memory. The result revealed that fish in T2 showed better growth performance with an average weight gain of 56.11 ± 0.51 g. Thus, it had been concluded that Piper nigrum, a medicinal plant, can also be used to enhance the growth performance and immune response of Labeo rohita as attractive alternatives against antibiotics and vaccines and has shown no negative side effects on fish health as well as on its environment.

Um estudo foi conduzido para avaliar o efeito do extrato da folha de Piper nigrum (pimenta-do-reino) sobre o desempenho de crescimento, composição centesimal, parâmetros hematológicos e resposta imune de alevinos de Labeo rohita com peso médio de 22,14 ± 0,98g. Após aclimatação por duas semanas, os peixes (n = 25) foram selecionados aleatoriamente e colocados em quatro aquários de vidro (T0, T1, T2 e T3) em temperatura constante da água (30,0 ± 1,0 °C), pH (7,50 ± 0,5) e dureza total (200 ± 2,0 mgL-¹) por um período de 12 semanas, com três repetições cada. Os peixes foram alimentados com ração suplementada com extrato de folha de P. nigrum @ 0,0%, 1,0%, 2,0% e 3,0% em T0, T1, T2 e T3, respectivamente. Ao final do experimento, cinco peixes foram selecionados aleatoriamente de cada aquário para composição centesimal, carga microbiana intestinal e cutânea e parâmetros hematológicos. Proteínas totais, albuminas e globulinas também foram registradas para avaliar a memória imunológica. O resultado revelou que os peixes em T2 apresentaram melhor desempenho de crescimento com ganho de peso médio de 56,11 ± 0,51 g. Assim, concluiu-se que Piper nigrum, uma planta medicinal, também pode ser usado para melhorar o desempenho de crescimento e resposta imunológica de Labeo rohita como alternativas atraentes contra antibióticos e vacinas e não mostrou efeitos colaterais negativos na saúde dos peixes, bem como sobre seu ambiente.

Limiting amino acids supplementation in low crude protein diets and their impacts on growth performance and carcass composition in Labeo rohita (rohu) adult fish / Limitação da suplementação de aminoácidos em dietas com baixo teor de proteína bruta e seus impactos no desempenho de crescimento e composição da carcaça em peixes adultos Labeo rohita (rohu)

Ninety days study was conducted in hapas installed in earthen ponds. Fish of an average initial weight (220g) were evenly distributed in triplicate groups within fifteen hapas. Five experimental diets labeled as T1 (25% CP and NRC recommended amino acid level) as control diet, T2 (with 2% low protein and 5% amino acid supplementation), T3 (with 2% low protein and 10% amino acid supplementation), T4 (with 4% low protein and 10% amino acid supplementation) and T5 (with 4% low protein and 20% amino acid supplementation) were prepared. Fish were fed with @3% of their body weight twice a day at 10.00 & 16:00 hour. Significantly higher percent weight gain (420.18 ± 66.84a) and specific growth rate (13499.33±1273.54a) along with improved feed conversion ratio (1.29 ± 0.09b) and hundred percent survivals were recorded during the trial. Furthermore proximate analysis of meat showed significant improvement in the crude protein level (81.77 ± 0.19a) served with diet containing 20% limiting amino acids mixture. Therefore, limiting amino acids can be a source of cost effective feed and use safely in L. rohita diet.

O estudo de 90 dias foi realizado em hapas instalados em tanques de terra. Peixes com peso inicial médio (220 g) foram distribuídos uniformemente em grupos triplicados em 15 hapas. Cinco dietas experimentais rotuladas como T1 (25% de CP e NRC recomendado nível de aminoácidos) como dieta controle, T2 (com 2% de proteína baixa e 5% de suplementação de aminoácidos), T3 (com 2% de proteína baixa e 10% de suplementação de aminoácidos), T4 (com 4% de baixa proteína e 10% de suplementação de aminoácidos) e T5 (com 4% de baixa proteína e 20% de suplementação de aminoácidos) foram preparadas. Os peixes foram alimentados com 3% do seu peso corporal duas vezes por dia às 10h00 e 16h00. Ganho de peso significativamente maior (420,18 ± 66,84a) e taxa de crescimento específico (13499,33 ± 1273,54a) juntamente com taxa de conversão alimentar melhorada (1, 29 ± 0,09b) e sobrevivência de cem por cento foram registrados durante o ensaio. Além disso, a análise aproximada da carne mostrou melhora significativa no nível de proteína bruta (81,77 ± 0,19a) servida com dieta contendo 20% de mistura de aminoácidos limitantes. Portanto, a limitação de aminoácidos pode ser uma fonte de alimentação econômica e usada com segurança na dieta de L. rohita.

Impact of dietary leaf extracts of Black pepper Piper nigrum L. on the growth, hematological and immunological parameters of Labeo rohita (Hamilton, 1822) cultured in glass aquaria

Abstract A study was conducted to evaluate the effect of Piper nigrum (black pepper) leaf extract on on the growth performance, proximate composition, hematological parameters, and immune response of Labeo rohita fingerlings with an average weight of 22.14 ± 0.98g. Aftrer acclimation for two weeks, fish (n=25) were randomly selected and placed in four glass aquaria (T0, T1, T2 and T3) at constant water temperature (30.0 ± 1.0 °C), pH (7.50 ± 0.5) and total hardness (200 ± 2.0 mgL,-1) for a period of 12 weeks, with three replicates each. Fish were fed with P. nigrum leaf extract supplemented feed @ 0.0%, 1.0%, 2.0% and 3.0% in T0, T1,T2 and T3, respectively. At the end of experiment, five fish were randomly selected from each aquaria for proximate composition, gut and skin microbial load, hematological parameters. Total proteins, albumins, and globulins were also recorded to evaluate immunological memory. The result revealed that fish in T2 showed better growth performance with an average weight gain of 56.11 ± 0.51 g. Thus, it had been concluded that Piper nigrum, a medicinal plant, can also be used to enhance the growth performance and immune response of Labeo rohita as attractive alternatives against antibiotics and vaccines and has shown no negative side effects on fish health as well as on its environment.

Resumo Um estudo foi conduzido para avaliar o efeito do extrato da folha de Piper nigrum (pimenta-do-reino) sobre o desempenho de crescimento, composição centesimal, parâmetros hematológicos e resposta imune de alevinos de Labeo rohita com peso médio de 22,14 ± 0,98g. Após aclimatação por duas semanas, os peixes (n = 25) foram selecionados aleatoriamente e colocados em quatro aquários de vidro (T0, T1, T2 e T3) em temperatura constante da água (30,0 ± 1,0 °C), pH (7,50 ± 0,5) e dureza total (200 ± 2,0 mgL-1) por um período de 12 semanas, com três repetições cada. Os peixes foram alimentados com ração suplementada com extrato de folha de P. nigrum @ 0,0%, 1,0%, 2,0% e 3,0% em T0, T1, T2 e T3, respectivamente. Ao final do experimento, cinco peixes foram selecionados aleatoriamente de cada aquário para composição centesimal, carga microbiana intestinal e cutânea e parâmetros hematológicos. Proteínas totais, albuminas e globulinas também foram registradas para avaliar a memória imunológica. O resultado revelou que os peixes em T2 apresentaram melhor desempenho de crescimento com ganho de peso médio de 56,11 ± 0,51 g. Assim, concluiu-se que Piper nigrum, uma planta medicinal, também pode ser usado para melhorar o desempenho de crescimento e resposta imunológica de Labeo rohita como alternativas atraentes contra antibióticos e vacinas e não mostrou efeitos colaterais negativos na saúde dos peixes, bem como sobre seu ambiente.

Limiting amino acids supplementation in low crude protein diets and their impacts on growth performance and carcass composition in Labeo rohita (rohu) adult fish

Abstract Ninety days study was conducted in hapas installed in earthen ponds. Fish of an average initial weight (220g) were evenly distributed in triplicate groups within fifteen hapas. Five experimental diets labeled as T1 (25% CP and NRC recommended amino acid level) as control diet, T2 (with 2% low protein and 5% amino acid supplementation), T3 (with 2% low protein and 10% amino acid supplementation), T4 (with 4% low protein and 10% amino acid supplementation) and T5 (with 4% low protein and 20% amino acid supplementation) were prepared. Fish were fed with @3% of their body weight twice a day at 10.00 & 16:00 hour. Significantly higher percent weight gain (420.18 ± 66.84a) and specific growth rate (13499.33±1273.54a) along with improved feed conversion ratio (1.29 ± 0.09b) and hundred percent survivals were recorded during the trial. Furthermore proximate analysis of meat showed significant improvement in the crude protein level (81.77 ± 0.19a) served with diet containing 20% limiting amino acids mixture. Therefore, limiting amino acids can be a source of cost effective feed and use safely in L. rohita diet.

Resumo O estudo de 90 dias foi realizado em hapas instalados em tanques de terra. Peixes com peso inicial médio (220 g) foram distribuídos uniformemente em grupos triplicados em 15 hapas. Cinco dietas experimentais rotuladas como T1 (25% de CP e NRC recomendado nível de aminoácidos) como dieta controle, T2 (com 2% de proteína baixa e 5% de suplementação de aminoácidos), T3 (com 2% de proteína baixa e 10% de suplementação de aminoácidos), T4 (com 4% de baixa proteína e 10% de suplementação de aminoácidos) e T5 (com 4% de baixa proteína e 20% de suplementação de aminoácidos) foram preparadas. Os peixes foram alimentados com 3% do seu peso corporal duas vezes por dia às 10h00 e 16h00. Ganho de peso significativamente maior (420,18 ± 66,84a) e taxa de crescimento específico (13499,33 ± 1273,54a) juntamente com taxa de conversão alimentar melhorada (1, 29 ± 0,09b) e sobrevivência de cem por cento foram registrados durante o ensaio. Além disso, a análise aproximada da carne mostrou melhora significativa no nível de proteína bruta (81,77 ± 0,19a) servida com dieta contendo 20% de mistura de aminoácidos limitantes. Portanto, a limitação de aminoácidos pode ser uma fonte de alimentação econômica e usada com segurança na dieta de L. rohita.

Impact of dietary leaf extracts of Black pepper Piper nigrum L. on the growth, hematological and immunological parameters of Labeo rohita (Hamilton, 1822) cultured in glass aquaria / Impacto de extratos de folhas da dieta de pimenta-do-reino Piper nigrum L. sobre o crescimento, parâmetros hematológicos e imunológicos de Labeo rohita (Hamilton, 1822) cultivado em aquários de vidro

Abstract A study was conducted to evaluate the effect of Piper nigrum (black pepper) leaf extract on on the growth performance, proximate composition, hematological parameters, and immune response of Labeo rohita fingerlings with an average weight of 22.14 ± 0.98g. Aftrer acclimation for two weeks, fish (n=25) were randomly selected and placed in four glass aquaria (T0, T1, T2 and T3) at constant water temperature (30.0 ± 1.0 °C), pH (7.50 ± 0.5) and total hardness (200 ± 2.0 mgL,-1) for a period of 12 weeks, with three replicates each. Fish were fed with P. nigrum leaf extract supplemented feed @ 0.0%, 1.0%, 2.0% and 3.0% in T0, T1,T2 and T3, respectively. At the end of experiment, five fish were randomly selected from each aquaria for proximate composition, gut and skin microbial load, hematological parameters. Total proteins, albumins, and globulins were also recorded to evaluate immunological memory. The result revealed that fish in T2 showed better growth performance with an average weight gain of 56.11 ± 0.51 g. Thus, it had been concluded that Piper nigrum, a medicinal plant, can also be used to enhance the growth performance and immune response of Labeo rohita as attractive alternatives against antibiotics and vaccines and has shown no negative side effects on fish health as well as on its environment.

Resumo Um estudo foi conduzido para avaliar o efeito do extrato da folha de Piper nigrum (pimenta-do-reino) sobre o desempenho de crescimento, composição centesimal, parâmetros hematológicos e resposta imune de alevinos de Labeo rohita com peso médio de 22,14 ± 0,98g. Após aclimatação por duas semanas, os peixes (n = 25) foram selecionados aleatoriamente e colocados em quatro aquários de vidro (T0, T1, T2 e T3) em temperatura constante da água (30,0 ± 1,0 °C), pH (7,50 ± 0,5) e dureza total (200 ± 2,0 mgL-1) por um período de 12 semanas, com três repetições cada. Os peixes foram alimentados com ração suplementada com extrato de folha de P. nigrum @ 0,0%, 1,0%, 2,0% e 3,0% em T0, T1, T2 e T3, respectivamente. Ao final do experimento, cinco peixes foram selecionados aleatoriamente de cada aquário para composição centesimal, carga microbiana intestinal e cutânea e parâmetros hematológicos. Proteínas totais, albuminas e globulinas também foram registradas para avaliar a memória imunológica. O resultado revelou que os peixes em T2 apresentaram melhor desempenho de crescimento com ganho de peso médio de 56,11 ± 0,51 g. Assim, concluiu-se que Piper nigrum, uma planta medicinal, também pode ser usado para melhorar o desempenho de crescimento e resposta imunológica de Labeo rohita como alternativas atraentes contra antibióticos e vacinas e não mostrou efeitos colaterais negativos na saúde dos peixes, bem como sobre seu ambiente.

Immune-biochemical response and immune gene expression profiling of Labeo rohita fingerlings fed with ethanolic tea leaf extracts and its survivability against Aeromonas hydrophila infection.

The present study was conducted to evaluate the immunostimulatory effect of tea leaf extract (Camellia sinensis) on Labeo rohita and its resistance against Aeromonas hydrophila infection. The ethanolic extract of green tea (GTEE) was found to be the most potent as compared to other solvent extract which was used for further study. It was used to evaluate immune-biochemical response of L. rohita fingerlings, fed with tea leaf extract (control- 0.0%, 0.2% (T1), 0.4% (T2), 0.8% (T3) and 1% (T4) of GTEE kg-1 feed). Different biochemical parameters like glucose, ALP, GPT, GOT, and immunological parameters like lysozyme activity, NBT, anti-protease activity, myeloperoxidase activity, plasma protein, and immune relevant genes (IL-10, C3, Lysozyme G type and iNOS) expressions were carried out. The immunological parameters such as lysozyme activity, NBT and myeloperoxidase activity showed significantly high value once fed with GTEE incorporated diets. Significant up-regulation of immune genes indicated the enhancement of immune response at molecular level. The biochemical parameters were found to be significantly decreasing, indicating that the extract had hepato-protective effect and can help to overcome stress. The fish, fed with GTEE incorporated diets, showed resistance against A. hydrophila when compared with the control group. 0.2% GTEE showed the highest post-challenged survival (76.67%). From the present study, it is concluded that GTEE @ 0.2% can be used as potent immunostimulant as a sustainable alternative prophylactic and therapeutic agent in aquaculture.

Effect of Fortified Feed with Phyto-Extract on the First Physical Barrier (Mucus) of Labeo rohita.

The aim of the current study was to assess the effect of two different fortified feeds with different concentrations of two important medicinal plants (Withania coagulans and Zingiber officinale) on the mucosal immunity of Labeo rohita. After a dietary intervention, mucus was tested against five pathogenic bacteria (in-vitro), while experimental fish were tested against the ectoparasite (Lernaea) (in-vivo). Our results revealed that all fish groups fed with different concentrations (1, 1.5, and 2%) of Z. officinale had low molecular weight proteins and did not develop any significant signs of parasitic infection, with low mortality rate; whereas the groups that were fed with W. coagulans (particularly with 1% and 2%), including a control group, developed rapid signs of infection with high mortality rate. The highest hemagglutination titer value was recorded for the fish fed with 1% and 1.5% of Z. officinale. The lowest value was found for the fish fed with 2% of W. coagulans. The mucus of all fish of fortified groups was active and inhibited the growth of tested bacterial pathogens as compared to the control group. Further, Z. officinale groups showed greater efficacy against bacteria as compared to the W. coagulans groups. In conclusion, Z. officinale can be considered as a potential and functional ingredient in aquaculture feed. Furthermore, future studies should be conducted to investigate more details on the subject.

Improvement in the growth status and carbohydrate utilization of Labeo rohita (Hamilton, 1822) fingerlings with dietary supplementation of chromium picolinate.

A 60-day feeding experiment was conducted to evaluate the effects of dietary chromium (Cr) on carbohydrate utilization and growth performance of Labeo rohita fingerlings. Fishes were fed with four high carbohydrate (53%), isonitrogenous (crude protein 35%), and isocaloric (415 Kcal, 100 gm-1) experimental diets containing different levels of dietary chromium picolinate (Cr-Pic) viz.0, 400, 800, and 1200 µg kg-1 diet. Weight gain (WG%), specific growth rate (SGR), feed efficiency ratio (FER), and protein efficiency ratio (PER) were significantly increased at 800 µg kg-1 diet chromium supplementation (P < 0.05). Cr-Pic supplementation (800 µg kg-1) also significantly (P < 0.05) enhanced the protein: DNA ratio in muscle, while DNA: RNA and DNA: tissue ratios were significantly (P < 0.05) decreased indicating higher growth. Significantly higher amylase, protease, and lipase activities were recorded in 800 µg Cr-Pic kg-1 diet fed fishes (P < 0.05), while any of the experimental groups showing no significant (P > 0.05) change in hexokinase activity, indicating normal glycolysis in all. Furthermore, significant (P < 0.05) decrease of glucose-6-phospatase activity in 800 µg Cr-Pic kg-1 diet fed group, showcasing an evidence for protein-sparing action with Cr-Pic supplementation. Significantly (P < 0.05) higher serum insulin and liver glycogen in 800 µg Cr-Pic kg-1 diet fed fishes denote an improvement in carbohydrate metabolism. However, significantly (P < 0.05) higher ATPase and SOD activities were also observed when chromium supplementation was more than 800 µg kg-1 diet, indicating stress at higher level. The present study indicates that growth and carbohydrate utilization can significantly (P < 0.05) be improved by feeding the L. rohita fingerlings with Cr-Pic (800 µg kg-1 diet) supplemented diet in laboratory condition.

Impact of grape pomace flour (GPF) on immunity and immune-antioxidant-anti-inflammatory genes expression in Labeo rohita against Flavobacterium columnaris.

This study evaluates the effects of dietary inclusion of grape pomace flour (GPF) on growth, antioxidant, anti-inflammatory, innate-adaptive immunity, and immune genes expression in Labeo rohita against Flavobacterium columnaris. In both normal and challenged fish the growth rate, hematology and biochemical parameters significantly increased when fed with 200 and 300 mg GPF enriched diets; similarly the activities of antioxidants and innate-adaptive immune parameters, such as malondialdehyde (MDA), superoxide dismutase (SOD), glutathione peroxidase (GPx), glutathione (GSH), phagocytic (PC), respiratory burst (RB), alternative pathway complement (ACP), lysozyme (Lyz), and total immunoglobulin M (IgM) significantly increased in both groups. Similarly, the immune, antioxidant, and anti-inflammatory-related gene mRNA expression was significantly up-regulated in head kidney (HK) tissues. The challenged fish fed without GPF always exhibited lower values of all the studied parameters. The results indicate that both normal and challenged fish treated with 200 mg GPF inclusion diet had significantly enhanced growth rate, antioxidant status, and immune defense mechanisms than with 300 mg GPF diet in L. rohita against F. columnaris.

Effects of dietary supplementation of vitamin-E and commercial probiotics on the innate immunity of Labeo rohita against Aeromonas hydrophila infection.

Immunomodulation is one of the useful tools to prevent diseases in aquaculture. In this study, the immunomodulatory effects of vitamin-E (100 mg/kg feed) and commercial probiotic consortia, Rhodomax™ (5 g/kg feed) on the innate immunity of Labeo rohita and their protective effect against Aeromonas hydrophila infection were evaluated and compared. Three groups of fish at 30 numbers/tank were fed with vitamin-E, probiotic and control diets at 3% body weight for 30 days, in triplicate. Following this, the fish of all groups were injected intramuscularly with A. hydrophila N10P at 2.40 × 107 cells/fish. The growth indices like specific growth rate (SGR) and feed conversion ratio (FCR) during the feeding and the non-specific immune responses during the feeding and post-challenge regimen were recorded. The dietary supplementation of vitamin-E and/or commercial probiotics caused significant improvements in the innate immunity of L. rohita compared to control. Nevertheless, the vitamin-E diet offered markedly better results in terms of SGR, FCR, ceruloplasmin, antiprotease, myeloperoxidase and phagocytic activities of L. rohita during the feeding and post-challenge regimen. While the respiratory oxidative burst activity was enhanced in probiotic diet-fed L. rohita only during the feeding regimen. All the immune parameters reached normalcy on day 15 post-injection with A. hydrophila. These findings revealed that supplementation vitamin-E at 100 mg/kg feed may improve the growth indices, prime the non-specific immune responses of L. rohita against A. hydrophila infection and enhance the overall health status than the tested commercial probiotics.

Responses of Corpuscles of Stannius to intra-peritoneal vitamin-D3 administration in teleost Labeo rohita (Hamilton, 1822) reared in water with two different levels of calcium concentration.

This study assessed the responses of vitamin-D3 intraperitoneally injected to Rohu, Labeo rohita @ of 0 IU/kg bw (only solvent), 100 IU/kg bw and 500 IU/kg bw reared in 20 and 40 ppm of calcium (Ca) enriched water. The cellular changes in Corpuscles of Stannius (CS) gland, serum Ca, and inorganic phosphate (Pi) level were analysed up to the 60th day. Rohu administered with 100 IU/kg bw D3 and exposed to 40 ppm Ca-rich water exhibited notable hyperplasia of CS compared with their control groups. Notable changes with high serum Ca level (13.87 ± 0.3 mg/dl) was detected on the 5th day in fish exposed to 40 ppm Ca-rich water, while related values attained (13.74 ± 0.1 mg/dl) only after 7 days in 20 ppm Ca-rich water of 500 IU/kg bw vitamin D3 injection. Similarly, high serum Pi level (7.66 ± 0.2 mg/dl) in 40 ppm Ca injected with D3 at 500 IU/kg bw. The results demonstrated that the Ca homeostasis of Labeo rohita is influenced by intra-peritoneal vitamin D3. Progressive studies should be conducted by increasing the dose of vitamin D3 to investigate optimum dose/supplement in feed for commercially important aquaculture teleost Labeo rohita for maximum and sustainable absorption of Ca from the variable water Calcium levels to maintain Ca2+ homeostasis.

Molecular characterization and expressional quantification of lgp2, a modulatory co-receptor of RLR-signalling pathway in the Indian major carp Labeo rohita following pathogenic challenges and PAMP stimulations.

Lgp2 (laboratory of genetics and physiology 2) is a cytosolic viral sensor of the RLR (retinoic acid-inducible gene 1 like receptor) family member without the caspase recruitment domain, having both inhibitory and stimulatory roles in RLR-signalling pathway. In India, Labeo rohita (rohu) is one of the leading and economically favoured freshwater fish species. Several immunological sentry proteins have been reported in this fish species, but no information is available on the RLR members. This study was aimed at cloning and characterization of full-length lgp2-cDNA (complementary DNA) in rohu and investigation of its expressional modulations following various pathogen-associated molecular pattern stimulations and bacterial infections. The full-length lgp2-cDNA sequence obtained through rapid amplification of cDNA ends-PCR consisted of 2299 nucleotides with an open reading frame of 2034 bp encoding 677 amino acids. In rohu-Lgp2, four conserved domains - a DEAD/DEAH box helicase domain, Pfam type-III restriction enzyme domain, helicase superfamily c-terminal domain and RIG-I C-terminal regulatory domain - have been detected. Within these domains, several important functional motifs, such as ATP-binding site, ATPase motif, RNA unwinding motif and RNA-binding sites, have also been identified. In healthy rohu, lgp2 gene was abundantly expressed in gill, liver, kidney, spleen and blood. In response to both in vitro and in vivo treatments using double-stranded RNA (poly I:C), lgp2 gene expression was significantly (P < 0.05) upregulated in all tested tissues and also in the LRG (Labeo rohita gill) cells. lgp2 gene expression significantly (P < 0.05) increased on stimulation of LRG cells using γ-d-glutamyl-meso-diaminopimelic acid and muramyl dipeptide. In vivo treatment using lipopolysaccharide and Aeromonas hydrophila-derived RNA resulted in both up- and down-regulation of lgp2 gene expression. Upon gram-positive and gram-negative bacterial infections, the expression of the lgp2 gene increased at different times in almost all the tested tissues. These integrated observations in rohu suggest that Lgp2 is an antiviral and antibacterial cytosolic receptor. SIGNIFICANCE STATEMENT: Lgp2, a cytosolic viral sensor of retinoic acid-inducible gene 1 like receptor family member, has been cloned in Labeo rohita. The complete sequence of rohu lgp2-complementary DNA consisted of 2299 nucleotides with an open reading frame of 2034 bp encoding 677 amino acids. It consisted of a DExDc, RES-III, HELICc, Pfam RIG-I_C-RD, ATP-binding site, ATPase motif, RNA unwinding motif and RNA-binding site. Upon bacterial infection, double-stranded RNA and various pathogen-associated molecular pattern stimulations, lgp2 gene expression significantly increased, indicating its role as an antiviral and antibacterial cytosolic receptor.