Synthesis of Iron Nanoparticles Using Sargassum wightii Extract and Its Impact on Serum Biochemical Profile and Growth Response of Etroplus suratensis Juveniles.

The present study focuses on the green synthesis of iron nanoparticles using plant extracts as reducing, capping, and stabilizing agents. Aqueous seaweed extracts with the addition of iron solution were mixed using a magnetic stirrer which resulted in a color change indicating the formation of iron nanoparticles. The iron nanoparticles were successfully synthesized using Sargassum wightii extract. The synthesized iron nanoparticles were characterized by UV-Vis spectrophotometer, Fourier transform infrared spectroscopy (FTIR), and zeta potential techniques. The UV-Vis spectra showed a peak at 412 to 415 nm. Zeta potential revealed that the synthesized iron nanoparticles were negative and positive charges. FTIR spectroscopy analysis showed the presence of chemical bond and amide group likely to be responsible for the green synthesis of iron nanoparticles. The effect of nano-iron as a dietary iron source on the growth and serum biochemical profile of Etroplus suratensis fingerlings was evaluated. Iron nanoparticles were fed to E. suratensis fingerlings for 60 days with two levels 10 mg (T1) and 20 mg (T2) and a control group without iron nanoparticles. The highest WG% and SGR and lowest FCR were observed in the T2 group which is significantly different (p < 0.05) from other groups. The serum biochemical profile showed significantly increased activity on 20 mg/kg of nano-iron-supplemented diet. The findings of the present study concluded that supplementation of nano-iron at the 20 mg/kg level to the regular fish diet has a better impact not only on growth but also on the overall health of the fish.

Computational analysis and functional characterisation of Tor putitora toll-like receptor 4 with the elucidation of its binding sites for microbial mimicking ligands.

In the current study, full-length Toll-like receptor 4 (TLR4) cDNA was cloned and characterised in Tor putitora, an important fish inhibiting Himalayan rivers. The complete coding sequence of TpTLR4 is 2457 bp with nine key structural domains, including six leucine-rich repeats (LRRs). The phylogenetic tree revealed that TpTLR4 showed the closest relationship with TLR4 of Cyprinus carpio (96%), Labeo rohita (91%) and Megalobrama amblycephala (88%), all belonging to the Cyprinidae family. CELLO2GO tool revealed that TpTLR4 protein is highly localised in the plasma (67.7%), and the protein has a strong association with myeloid differentiation primary response 88 (MYD88) followed by Tumor necrosis factor receptor-associated factor (TRAF) family. In the toll-interleukin-1 receptor (TIR) domain of TpTLR4, the proline is replaced by the alanine amino acid, thus may give plasticity to the receptor to recognise both bacterial and viral ligands. Molecular docking has revealed that TpTLR4 showed the strongest affinity towards poly (I:C) with the binding energy of -6.1 kcal/mol and five hydrogen bonds among all ligands. Based on our molecular docking results, it can be presumed that TpTLR4 can sense bacterial, fungal and viral molecular patterns with binding sites mainly present in the TpTLR4 LRR9 motif, which spans between 515 and 602 amino acids. Tor putiora TLR4 transcript was ubiquitously expressed in all the tested fish tissues. Although, transcript level was found to be highest in blood and spleen followed by the kidney. The TpTLR4 transcripts showed peak expression in spleen and kidney at 12 h post-injection (hpi) (p < 0.05) of poly (I:C). The constitutive expression of TpTLR4 in various tissues, up-regulation in different tissues and strong binding affinities with poly (I:C) indicate that TpTLR4 may play an essential role in sensing pathogen-associated molecular patterns (PAMPs), particularly of viral origin.

Temporal changes in superoxide dismutase, catalase, and heat shock protein 70 gene expression, cortisol and antioxidant enzymes activity of Labeo rohita fingerlings subjected to starvation and refeeding.

A short term starvation and refeeding experiment was conducted to study the temporal changes in SOD, CAT and HSP70 gene expression of Labeo rohita fingerlings. The study was carried out for 15 days with initial 7 days of starvation and then refeeding up to 15th day of the experimental trial. The expressions of SOD and CAT genes of liver and gills were significantly up-regulated after 7 days of starvation, down-regulated after 3 days of refeeding, and returned to the basal values after 8 days of refeeding. The HSP70 gene expression was significantly (p < 0.05) increased after starvation, with highest mRNA expression found on 7th day and reduced to the levels of control on refeeding. The activities of antioxidant enzymes, SOD and CAT were also studied to correlate with the results of gene expression. The changes in activities of SOD and CAT were found significantly (p < 0.05) higher in the starved group compared to the fed group. The dynamics of AST and ALT in serum revealed a progressive increase till the 7th day and decreased upon refeeding, cortisol level also has shown significant increase up to 7th day of starvation and sharp decline on refeeding. The concentration of blood glucose level start declining on 3rd day onwards with lowest level found on 7th day of starvation and was quickly restored to the levels of control on refeeding. The present study reveals that starvation elicits oxidative stress response as revealed by enhanced expression and activities of antioxidant enzymes, HSP 70 and serum biochemical alterations. However, these alterations were restored upon refeeding of L. rohita within 7 days.

Fabrication and characterization of chitosan conjugated eurycomanone nanoparticles: In vivo evaluation of the biodistribution and toxicity in fish.

Chitosan nanoparticles (CNPs) have been proven considerable delivery agents due to their remarkable physicochemical properties. Present study reports the fabrication of CNPs by ionic gelation process and their characterization by different approaches. The constructed nanoparticles were successfully conjugated with eurycomanone with significant entrapment efficiency. Particle size of chitosan and chitosan conjugated eurycomanone nanoparticles were 126.2nm and 130nm respectively. Scanning electron microscopy showed that the particles were spherical in shape and well dispersed. Cross-linking between CNPs and eurycomanone (CENPs) were confirmed by Fourier-transform infrared (FTIR) spectroscopy. Fluorescent nanoparticles were prepared by using Rhodamine-6G dye, characterised by SEM and confirmed for conjugation by FTIR. Biodistribution of CENPs showed the presence of fluorescent nanoparticles in liver, kidney, testes and brain of C. magur. The toxicity of CENPs was evaluated by comparing the histological sections of catfish testes collected from treated and control group. No signs of toxicity were seen in testes after the delivery of CENPs. Molecular docking study revealed high spontaneous binding ability of chitosan with eurycomanone and aromatase enzyme. The study reports that CNPs can act as a stabilizing agent for eurycomanone formulation and could be a promising approach to increase the reproductive performance of the fishes.

Cloning, characterisation, docking and expression analysis of 3-beta-hydroxysteroid dehydrogenase during ontogenetic development and annual reproductive cycles in catfish, Clarias batrachus.

Fish like higher animals, have a well-defined mechanism to produce sex steroids that play a critical role in gonadal development and maturation. In this study, we aimed to analyse the expression pattern of 3ß-HSD in different tissues, during ontogenetic development and gonadal recrudescence of Clarias batrachus. A full-length cDNA of 1617 bp including an open reading frame (ORF) of 1125 bp encoding 374 amino acids was isolated from testes of C. batrachus. The docking analysis between C. batrachus 3ß-HSD protein and eurycomanone exhibited high binding affinity toward each other with total energy of -108.292 kcal/mol and van der Waals (VDW) interaction of -84.2838 kcal/mol. The 3ß-HSD transcript level during ontogeny was detected in all the stages starting from the fertilized egg. The mature C. batrachus showed more expression of 3ß-HSD mRNA in gonads and brain while weak expression was detected in the remaining tissues analysed. The 3ß-HSD mRNA expression during annual reproductive phases of gonads was more in preparatory and pre-spawning stages than that of spawning and post-spawning phases. The mRNA expression results together suggest that 3ß-HSD plays an important role in gonadal development. Furthermore, the active binding sites on 3ß-HSD protein could be targeted in pharmacological drug designing to cope with reproductive dysfunctions in fish.