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Eurycomanone (EN), a quassinoid present in E. longifolia plants acts as a testosterone booster in male animals. The present study was aimed to explore the role of EN in females using fish as model organism. Chitosan nano-conjugated EN nanoparticles were synthesised and tested for their role in reproductive response in female catfish. Three injections of the formulation were given for a period of 21 days and the mRNA expression profile of important genes as well as serum hormones involved in steroid hormone synthesis were evaluated. In gonads, the mRNA expression of genes was upregulated except CYP19a1 (aromatase), that significantly decreased in the treatment groups. The hormone genes, FSH-β and LH-β mRNA levels in brain were upregulated whereas CYP19a2 was sharply downregulated following treatments. The testosterone varied non-significantly while the estradiol-17β serum hormone was significantly diminished. The effect was sustained in chitosan conjugated treatments compared to EN alone. The Gonadosomatic index (GSI) varied non-significantly in different treatments and control. In treatment groups, histological analysis showed the ovarian follicles in the regressed phase. Analysis of the results of the undertaken research leads to conclusion that that EN may not be beneficial for female reproduction due to its obstructive role in estrogenic production.

To understand the carbohydrate utilization ability during developmental stages of Clarias magur, the mRNA levels and enzyme activity of amylase were studied. Furthermore, the transcript levels of some important growth genes like growth hormone (GH), insulin-like growth factor I (IGF-I) and insulin-like growth factor binding protein (IGFBP) were evaluated. The enzymatic activity and mRNA expression of alpha-amylase along with expression of growth-related genes were examined from hatching until 34 days post-hatching (dph) by quantitative reverse transcription-polymerase chain reaction (qRT-PCR). Subsequently, mRNA expression pattern revealed the significantly (P .05) in the mRNA expression of growth-related genes among the different time points of the present study. Moreover, the amylase activity revealed almost same pattern as that of its mRNA expression. In general, the present results suggested that C. magur has the ability to utilize the digestible carbohydrates during the early stages and can be helpful in developing low cost feeds for larvae.

The study was undertaken to explore the molecular mechanism of eurycomanone, a major compound of Eurycoma longifolia plant in increasing the reproductive processes in the male fish model. Chitosan-nanoconjugated eurycomanone nanoparticles with a significant particle size [130 nm (CED1); 144.1 nm (CED2)] and stable zeta potentials (+49.1 mV and +30 mV) were synthesized and evaluated against naked eurycomanone (ED1 and ED2). In present study, short-term and long-term experiments were conducted to evaluate the effect of nano-formulation on expression of endocrine-related genes, circulating hormone concentrations (Follicle stimulating hormone, FSH; luteinizing hormone, LH; progesterone, testosterone and 17-β estradiol) and reproductive capacity of male Clarias magur. In short-term experiment, the sampling of tissues was done on hourly basis after injection of eurycomanone either alone or with chitosan and long-term experiment was carried for 21 days and in this the injection was repeated after 7 days and 14 days. Treatments CED1 and CED2 showed controlled and sustained surge of the transcript level of selected genes (except aromatase) and serum hormones (except 17β-estradiol) compared to ED1 and ED2 groups. The transcript levels of aromatase and serum 17β-estradiol hormone showed the declining trend in the chitosan conjugated groups. The gonadosomatic index (GSI), reproductive capacity, intracellular calcium and selenium and cellular structure of testes were improved in CED1 and CED2 groups compared to other treatments. Furthermore, the effect of chitosan conjugated eurycomanone was evaluated in primary testicular cells and an increase in the mRNA expression level of endocrine-related genes was detected. This is the first report of the use of chitosan conjugated eurycomanone and present study elucidates the molecular mechanism of eurycomanone in increasing the reproductive output in animals.

Reproduction is arguably the most vital process in animals including fish that is being controlled by the brain through hypothalamic-pituitary-gonadal (HPG) axis. Several bioactive compounds from plants were recognized to be having a great role in promoting a variety of biological activities such as growth, immunomodulation, feed consumption and antioxidant activity or can act as antiviral agents. Moreover, phytochemicals have been used for several hundred years as additions to energize, vitalize and eventually curb the male and female reproductive functions. There can be different approaches to administer the plant-based bioactive compounds such as oral, immersion or injection methods. However, the nanodelivery approach forms an effective way to achieve the delivery to target tissues with less or no impediment. The nanodelivery approach improves the efficiency of targeted delivery of plant-based bioactive compounds. The phytochemicals can also be involved in phenotypic sex manipulations that sometimes become necessary in fisheries and aquaculture. The aim of this chapter is to provide an overview of available studies on the reproductive control via HPG axis along with different phytochemicals and their delivery approaches for reproductive modulation.

The gene expression of the trypsinogen and pepsinogen and activity of trypsin and pepsin in Indian walking catfish (Clarias magur) were investigated during the larval period to study the development of the digestive system. To this end, cDNA encoding the partial trypsinogen and pepsinogen were cloned and sequenced. Subsequently, the mRNA expression of genes was evaluated by quantitative polymerase chain reaction (qRT-PCR) along with the estimation of the activity of trypsin and pepsin. The mRNA expression pattern of trypsin appeared on 3rd-day post-hatching (dph) with a peak at 9 dph and 22 dph, while mRNAs of pepsin although detected at 4 dph but exhibited the significantly (P

The study was undertaken to investigate the toxicity level and safe dosage of eurycomanone, a major compound of Eurycoma longifolia plant extract in catfish. For that, combined approaches like, genotoxicity studies, biochemical, molecular and histopathological analysis were followed. The 96 h LD 50 of eurycomanone was found to be 391.716 μg/kg (0.3917 μg/g). The sub-acute toxicity levels of eurycomanone at concentrations, 0.274 μg/g (70% LD 50 ; Ed1) and 0.137 μg/g (35% LD 50 ; Ed2) were analysed to determine the safer dose for fish. Significant genotoxic effects (micronuclei and DNA damage) and elevation in biochemical parameters (AST, ALT and LDH) were detected in the Ed1 group than Ed2. Transcript levels of apoptotic genes were significantly more ( P 50 of eurycomanone was detrimental while 35% was safer for the catfish. This is the first report showing the toxicity levels of eurycomanone on fish. Furthermore, the safe dose could be effectively used to exploit the beneficial effects of this compound as used in higher animals.

An experiment of 60 days was conducted to study the immunomodulatory response and growth performance of Labeo rohita fingerlings fed with fucoidan rich seaweed extract (FRSE) and methionine. Four hundred and twenty fingerlings were randomly distributed into seven different experimental groups in triplicates following completely randomised design (CRD). Fishes were fed to satiation with purified diets containing either 0% FRSE with 0.3% methionine (control), 0% FRSE with low dose (0.9%) methionine (T 1 ), 0% FRSE with high dose (1.5%) methionine (T 2 ), 1% FRSE with 0.3% methionine (T 3 ), 2% FRSE with 0.3% methionine (T 4 ), 1% FRSE with high dose methionine (T 5 ) or 2% FRSE with low dose methionine (T 6 ) in the feed. There was a significant ( P 6 group. At the end of the experimental trial fishes were challenged with Aeromonas hydrophila and the highest relative survival percentage was recorded in the T 6 group, which was not significantly different from T 4 group ( P > 0.05). Immunological parameters: respiratory burst activity, myeloperoxidase activity, lysozyme activity, total immunoglobulin, phagocytic activity and total leukocyte count (TLC) were increased with the 2% level of dietary FRSE along with low dose methionine, whereas serum albumin/globulin (A/G) ratio and blood glucose level showed a decreasing trend ( P L. rohita fingerlings.

Early larval developmental stages of fish are highly susceptible to opportunistic pathogens until the complete maturation of the lymphoid organs. Knowledge of the expression pattern of important markers of adaptive immune system during the ontogenetic development is essential before vaccinating the fish. In the present study, Pterophyllum scalare (angelfish) was taken to explore the relative expression profile of developmental markers of adaptive immunity, recombination activating gene-2 (RAG-2) and immunoglobulin M (IgM). The fishes were bred and early developmental stages (0–45 days post-hatched) were used to assess the expression profile. The genes, RAG-2 and IgM were cloned and sequenced with the base pair lengths of 1958 bp and 225 bp respectively. The mRNA expression of RAG-2 appeared at insignificant level at the first day of hatching, but the expression was significantly increased from 24 dph (days post-hatching) onwards and reached its peak at 27 dph. The results proved that the maturation of lymphoid organs was completed at 27 dph as the respective protein is involved in the V(D)J recombination, important for the maturation of lymphoid organs. A similar trend was also observed in the mRNA transcript levels of IgM gene and a significantly high expression was detected from 27 dph onwards. The present study suggested that the suitable time for vaccination in P. scalare could be taken at 27 dph, as the maturation and development of lymphoid organs is completed thus helps in stimulating the adaptive response of immunity against any pathogen.

The study was conducted to investigate the expression and activity of key lipolytic enzymes during the ontogenetic development of Clarias magur. After partial characterization, the messenger RNA (mRNA) expression analysis of lipoprotein lipase (LPL), pancreatic triacylglycerol lipase (PL), and bile salt-activated lipase (BAL) genes along with the specific lipase activity were performed in larvae from Day 1 after hatching till 34-day posthatch (dph). Heterogeneous patterns of mRNA expression were shown by the important lipolytic enzymes and were detected before first exogenous feeding during the yolk-sac stage. LPL started increasing from 13 dph and peaked at 16 dph followed by a declining trend till 34 dph. However, the PL observed to be peaking at 9, 22, and 30 dph. Similarly, BAL showed an increasing trend from 11 to 22 dph with a significantly high level of mRNA expression at 16 dph. Later, the specific lipase activity was evaluated which appears at Day 1 after hatching with a progressive increase from 7 to 16 dph and a further declining trend afterwards with a peak at 22 dph. The results indicated the development of exocrine pancreas at 16 dph. Furthermore, the transcript levels and the activity of lipases were regulated with the age. Hence, the present study can be helpful in devising different strategies containing optimum lipid levels at a suitable stage of development for improving the survival during larval rearing. Furthermore, the study could be a baseline for elucidating the optimized dietary lipid levels of this catfish during its larval rearing.

Large scale farming of Clarias magur (Indian walking catfish) is limited due to the non-availability of seed because of high mortality rate occurring during its larval rearing. The aim of the present study was to develop an optimal strategy based on the dietary lipids and proteins for enhancing the larval survival. A 3-week study from 14-35 day post hatching (dph) was conducted to elucidate the effects of dietary lipid and protein level on growth, survival and mRNA expression of growth related genes of C. magur larvae. Significantly (P