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.

Effect of Different Early Weaning Diets on Survival, Growth, and Digestive Ontogeny of Channa striatus (Bloch, 1793) Larvae.

The present study was carried out to evaluate the survival, growth, and digestive ontogeny of C. striatus larvae fed with different experimental diets from 4 days post-hatch (dph) to 32 dph at three-day intervals. A total of 24,000 larvae, with 1600 larvae per tank in triplicate and an initial mean weight of 0.64 ± 0.01 mg at 4 days post hatch (dph) were subjected to five different early weaning diets, namely Artemia nauplii (T1), co-feed diet comprising Artemia nauplii and formulated micro diet (T2), formulated micro diet (T3), formulated micro diet with protease supplementation (T4), and a commercial diet (T5). All the early weaning diets significantly affected the survival, growth, and ontogeny of the digestive system. Initially at 8 dph, C. striatus fed with T1 showed better survival and growth performance compared to other treatments. By 12 dph, the larvae fed with T1 and T2 showed similar results in terms of survival and growth performance, outperforming other treatments. However, the larvae fed with T2 and T4 outperformed T1 in survival and growth performance at 16 dph. By 24-32 dph, the larvae fed with all treatments met the basic nutritional needs for survival, with T4 fed larvae showing better growth compared to other treatments. At the end of the trial, cumulative mortality was lowest in larvae fed with T1 and highest in the larvae fed with T3 and T5. Similarly, the larvae fed with T4 showed significantly higher weight gain, specific growth rate (SGR), and average daily growth (ADG), while T1 fed larvae exhibited better feed conversion ratio (FCR) and protein efficiency ratio (PER). The enzyme activity fluctuated throughout the experimental duration. Lavae fed with T1 and T2 showed higher enzyme activities initially. However, T4 fed larvae showed higher trypsin and chymotrypsin specific activity at 16 dph along with well-developed intestinal folds with dense microvilli, higher pepsin-specific activity at 20 dph onwards with fully developed gastric glands and thicker gastric mucosal epithelium, and higher amylase and lipase activity at 16 dph with large and prominent zymogen granules in the exocrine pancreas. Peaking at 4 dph, the activity of protein metabolic enzymes (AST and ALT) sharply declined at 8 dph and increased until 32 dph. Larvae fed with T1 showed higher AST and ALT activity along with increased lipid deposits, followed by those fed with T2 and the larvae fed with T4 showing higher activity without fat accumulation but significantly lower than those fed T1 and T2. From the present research findings, it is recommended to initiate weaning for Channa striatus larvae with Artemia nauplii (from 4 dph to 8 dph) followed by a co-feeding regime (Artemia nauplii and formulated diet) between 9 and 16 dph and transition to protease-supplemented micro diet (T4) from 17 dph onwards.

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.

Intestinal ascariasis in children.

BACKGROUND: Ascariasis is a staggering health problem commonly seen in children of endemic areas. In the abdomen, ascaris lumbricoides can cause a myriad of surgical complications. Intestinal obstruction by ascaris lumbricoides is commonly seen in children. Most cases are managed conservatively. The purpose was to study the clinical presentation and management of symptomatic intestinal ascariasis in children. METHODS: A 3-year study was performed from April 2006 to April 2009 of pediatric-age patients who had symptomatic intestinal ascariasis. All patients had detailed clinical history, examination, plain X-ray of abdomen, and ultrasonography of abdomen. Peroperative findings were recorded in all patients who had surgical intervention. RESULTS: This prospective study had 360 patients. Male to female ratio was 1.37:1. 187 patients (52%) presented within 2-4 days of duration of illness. Mean +/- standard deviation (SD) age of patients was 6.35 +/- 2.25 years. Age group of 4-7 years (80%) was commonest group affected. Abdominal pain was a leading symptom in 357 patients (99%) with the pain in periumbilical area present in 215 patients (60%). In 227 patients (63%) abdominal distension was seen and was the commonest physical finding. Palpable worm masses were seen in 129 patients (36%); 81 patients (63%) had palpable worm masses in the umbilical quadrant. On X-ray of abdomen, visible worm masses were seen in 83 patients (23%). Abdominal sonography showed interloop fluid in 177 patients (49%) and free fluid in the pelvis of 97 patients (27%). The number of patients who were managed conservatively was 281 (78%), and 79 patients (22%) had surgical intervention. In patients who had surgical intervention, 39 patients (49%) had enterotomy and 7 patients (9%) had kneading of worms. Postoperative complications occurred in 33 patients, and an overall mortality of 1% (1 patient) was seen. CONCLUSIONS: Ascaridial intestinal obstruction is common in children in the Kashmir. Abdominal pain is the leading symptom in intestinal ascariasis. Plain X-ray and ultrasonography of the abdomen are used to diagnosis intestinal ascariasis. The majority of the patients can be managed conservatively.

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.

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.

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.

Appendiceal ascariasis in children.

BACKGROUND: The propensity of Ascaris lumbricoides to wander leads to varied surgical complications in the abdomen. Wandering A lumbricoides may sometimes reach the vermiform appendix and its presence there may remain silent or incite pathology. Our aim was to study ascariadial appendicitis. METHODS: Over a period of 3 years, we identified children who were found to have appendiceal ascariasis during surgery for different intestinal complications due to ascariasis. We studied the relationship between ascariasis and its lodgement inside the vermiform appendix in these patients. No preoperative diagnosis was made in this series. RESULTS: We found 11 patients with appendiceal ascariasis. It was incidentally found that 8/11 (72.7%) patients had worms inside their vermiform appendix but not appendicitis, whereas the remaining three patients (27.2%) were found to have Ascaris-associated appendicitis. The characteristic finding in Ascaris-infested vermiform appendix was that the worm is positioned with its head at the base and its tail at the tip of the appendix. CONCLUSION: Migration of A lumbrocoides inside the vermiform appendix is an incidental finding and tends to pursue a silent course in most patients. Only rarely does the presence of Ascaris inside the vermiform appendix cause appendicitis.

Spectrum of abdominal organ injury in a primary blast type.

INTRODUCTION: Abdominal organ injury in a primary blast type is always challenging for diagnosis. Air containing abdominal viscera is most vulnerable to effects of primary blast injury. In any patient exposed to a primary blast wave who presents with an acute abdomen, an abdominal organ injury is to be kept in a clinical suspicion. AIM: Study various abdominal organ injuries occurring in a primary type of blast injury. MATERIAL AND METHODS: All those who had exploratory laparotomy for abdominal organ injuries after a primary blast injury for a period of 10 years from January 1998 - January 2008 were included in this retrospective study. RESULTS: Total 154 patients had laparotomy for abdominal organ injuries with a primary blast type of injury. Small intestine was damaged in 48 patients (31.1%) followed by spleen in 22.7% cases. 54 patients (35.06%) had more than one organ injured. Liver laceration was present in 30 patients (19.48%). Multiple small gut perforations were present in 37 patients (77.08%). Negative laparotomy was found in 5 patients (3.24%) whereas 3 (1.94%) had re-exploration. Mortality was present in 6 patients (3.89%). CONCLUSIONS: Primary blast injury causes varied abdominal organ injuries. Single or multiple organ damage can be there. Small intestine is commonest viscera injured. Laparotomy gives final diagnosis.

Torsion of vermiform appendix with fecalith: a case report.

INTRODUCTION: Torsion of the vermiform appendix is a rare disorder, which causes abdominal symptoms indistinguishable from acute appendicitis and is found by chance during the laparotomy. CASE PRESENTATION: We report a case (a 76-year-old male) suffering of torsion of the vermiform appendix with fecalith. It was twisted 540 degrees in an anti-clockwise direction. Appendectomy was done. CONCLUSION: Appendiceal torsion may be assocated with the presence of fecalith. This case is the oldest one among the patients with appendiceal torsion reported to literature.