Neelam Saharan

Multi-response optimization of Artemia hatching process using split-split-plot design based response surface methodology

A novel method, BBD-SSPD is proposed by the combination of Box-Behnken Design (BBD) and Split-Split Plot Design (SSPD) which would ensure minimum number of experimental runs, leading to economical utilization in multi- factorial experiments. The brine shrimp Artemia was tested to study the combined effects of photoperiod, temperature and salinity, each with three levels, on the hatching percentage and hatching time of their cysts. The BBD was employed to select 13 treatment combinations out of the 27 possible combinations that were grouped in an SSPD arrangement. Multiple responses were optimized simultaneously using Derringer’s desirability function. Photoperiod and temperature as well as temperature-salinity interaction were found to significantly affect the hatching percentage of Artemia, while the hatching time was significantly influenced by photoperiod and temperature, and their interaction. The optimum conditions were 23 h photoperiod, 29 °C temperature and 28 ppt salinity resulting in 96.8% hatching in 18.94 h. In order to verify the results obtained from BBD-SSPD experiment, the experiment was repeated preserving the same set up. Results of verification experiment were found to be similar to experiment originally conducted. It is expected that this method would be suitable to optimize the hatching process of animal eggs.

Molecular characterization of inhibin-A: Structure and expression analysis in Clarias batrachus

The inhibins are disulphide-linked heterodimeric glycoproteins that belong to the TGFβ superfamily. Inhibins have been well studied in mammals but the information about their structure and function is very limited in lower vertebrates. The aim of the present study was to characterize inhibin-A and to understand its receptor binding interaction, and to evaluate its biological function in Clarias batrachus. Structure prediction of inhibin-A revealed two glycosylation sites on inhibin-α (Asp262 and Asn334). Docking of inhibin-A with its receptor; betaglycan and Act RIIA showed that residues Ser321, Gly324 and Leu325 of inhibin-α are involved in high affinity binding with betaglycan while inhibin-βA bound to Act RIIA by forming hydrogen bonds. The mRNA transcript analysis of various tissues indicated the presence of higher to moderate expression of inhibin-α and inhibin-βA in the gonads and the extra-gonadal tissues. Further, stage specific expression showed decreased levels of inhibin-α in the gonads during the annual reproductive cycles. Inhibin-βA, activin-βB and Act RIIA increased in the brain during spawning while FSHr increased in the gonads during the preparatory phase. Our study provides molecular, structural and functional insights of inhibin-A for the first time in C. batrachus.

Inhibin anti-peptide antibody macromolecule: An approach to improve fecundity in Clarias batrachus

Inhibins are members of the transforming growth factor beta (TGFβ) superfamily known to regulate ovarian functions through stimulation of the hypothalamo-pituitary-gonadal axis. In the present study, we aimed to design a species-specific inhibin-α chimeric peptide (ICP) and evaluate the effect of immunoneutralization using anti-ICP antisera to enhance the reproductive performance in female Clarias batrachus. Injection of anti-ICP antisera caused a significant increase in the number of oocytes at a medium dose (200 μl) in comparison to high dose (400 μl) and control (normal rabbit serum). Histological observations confirmed the dose-dependent advancement in oocyte maturation. Furthermore, anti-ICP antisera treated groups exhibited a significant increase in the serum concentrations of follicle stimulating hormone (FSH) and 17β-estradiol (E2) hormones. The anti-ICP antisera decreased the mRNA expression levels of inhibin-α while stimulated the transcript levels of inhibin-βA, FSHβ, CYP 19a1, 3β-HSD and StAR respectively in a dose-dependent manner. Collectively, these findings indicate that anti-ICP antibody macromolecules modulate the endogenous reproductive hormonal secretion and enhance oocyte quality and quantity in female C. batrachus. This is the first report wherein antibodies against inhibins were used to promote reproductive performances and investigate the underlying molecular mechanisms in fishes.

Growth and survival of marine sponges, Stylissa massa (Carter, 1887) and Liosina paradoxa (Thiele, 1899) in sea and land based culture systems

Two species of marine sponges Stylissa massa and Liosina paradoxa were cultured in cages (in situ) and in land based aquaria (ex situ) over a period of 120 days. Growth and survival were evaluated using four substrates viz., tile, block, coral rock and rope) under both in situ and ex situ conditions. L. paradoxa in cages recorded significant increase (p<0.05) on day 90 i.e., by 70% compared to initial volume whereas at 120 days there was a significant (p<0.05) decrease (54.22%). S. massa showed significant increase (p<0.05) in growth by 95.6% at 120 days compared to initial volume. Negative growth was recorded in S. massa under aquarium conditions while L. paradoxa recorded good growth as well as survival and performed exceptionally well in aquarium during the entire experimental period of 120 days. In cages, 90.62% survival was recorded for S. massa in 120 days. Among the different substrates used, coral rocks gave 100% survival whereas other substrates such as tiles, blocks and ropes showed 87.5% survival. Under aquarium conditions, S. massa showed overall survival of 81.25% and among the different substrates, 100% survival was obtained with coral rocks, 87.5% with blocks, 75% with ropes and the lowest survival of 62.5% was recorded with tiles.