A comparative metabolomics study on anadromous clupeid Tenualosa ilisha for better understanding the influence of habitat on nutritional composition.

INTRODUCTION: Fish inhabiting different aquatic habitats adapts to the environment by metabolomic readjustments. Understanding the combined activities of all the metabolic pathways (metabolome) helps in better understanding the complex interactions between gene and environment. OBJECTIVES: The anadromous migratory Tenualosa ilisha is a high value food fish comprising the dominant fishery of the rivers Padma and Hooghly. The present study aimed at understanding the influence of the two habitats on the nutritional composition of hilsa. METHODS: Metabolite profiling was carried out by GC/MS. De novo assembly of hilsa liver transcriptome was generated under Illumina HiSeq platform and multivariate analysis was employed for correlation and comparison. RESULTS: GC/MS fingerprinting showed C16:0, C18:1, C20:5 and C22:6 to be the predominant fatty acids present in hilsa liver, which were also found to be significantly higher in Hooghly hilsa. Comparative transcriptome analysis revealed that the differentially expressed genes were mainly associated with 'lipid metabolism' and 'amino acid metabolism' pathways. Multivariate analysis between the metabolites amino acid, fatty acid and corresponding gene expression showed that few genes of amino acid metabolism (EZH1, ALAS2 and ALDH4A1) positively correlated with individual amino acids (lysine, glycine and glutamate) in Hooghly hilsa. Similarly, the key genes for LC-PUFA biosynthesis (ELOVL5, FADS2, CPT1) showed positive correlation with individual LC-PUFAs (C18:3, C20:4, C20:5, C22:6), indicating higher LC-PUFA biosynthesis potential in Hooghly hilsa. CONCLUSION: Comparative metabolomic study in hilsa from the two different habitats showed that the habitats influence the nutritional composition as evidenced by high abundance of amino acids lysine, leucine and arginine and LC-PUFAs C18:3, C20:4, C20:5, C22:6 in Hooghly hilsa.

Dietary supplementation of curcumin augments heat stress tolerance through upregulation of nrf-2-mediated antioxidative enzymes and hsps in Puntius sophore.

Heat stress is one of the major environmental concerns in global warming regime and rising temperature has resulted in mass mortalities of animals including fishes. Therefore, strategies for high temperature stress tolerance and ameliorating the effects of heat stress are being looked for. In an earlier study, we reported that Nrf-2 (nuclear factor E2-related factor 2) mediated upregulation of antioxidative enzymes and heat shock proteins (Hsps) provide survivability to fish under heat stress. In this study, we have evaluated the ameliorative potential of dietary curcumin, a potential Nrf-2 inducer in heat stressed cyprinid Puntius sophore. Fishes were fed with diet supplemented with 0.5, 1.0, and 1.5% curcumin at the rate 2% of body weight daily in three separate groups (n = 40 in each group) for 60 days. Fishes fed with basal diet (without curcumin) served as the control (n = 40). Critical thermal maxima (CTmax) was determined for all the groups (n = 10, in duplicates) after the feeding trial. Significant increase in the CTmax was observed in the group fed with 1.5% curcumin- supplemented fishes whereas it remained similar in groups fed with 0.5%, and 1% curcumin-supplemented diet, as compared to control. To understand the molecular mechanism of elevated thermotolerance in the 1.5% curcumin supplemented group, fishes were given a sub-lethal heat shock treatment (36 °C) for 6 h and expression analysis of nrf-2, keap-1, sod, catalase, gpx, and hsp27, hsp60, hsp70, hsp90, and hsp110 was carried out using RT-PCR. In the gill, expression of nrf-2, sod, catalase, gpx, and hsp60, hsp70, hsp90, and hsp110 was found to be elevated in the 1.5% curcumin-fed heat-shocked group compared to control and the basal diet-fed, heat-shocked fishes. Similarly, in the liver, upregulation in expression of nrf-2, sod, catalase, and hsp70 and hsp110 was observed in 1.5% curcumin supplemented and heat shocked group. Thus, this study showed that supplementation of curcumin augments tolerance to high temperature stress in P. sophore that could be attributed to nrf-2-induced upregulation of antioxidative enzymes sod, catalase, gpx, and the hsps.

Micronutrient Composition of 35 Food Fishes from India and Their Significance in Human Nutrition.

The micronutrients (vitamins and minerals) are required in small amounts but are essential for health, development, and growth. Micronutrient deficiencies, which affect over two billion people around the globe, are the leading cause of many ailments including mental retardation, preventable blindness, and death during childbirth. Fish is an important dietary source of micronutrients and plays important role in human nutrition. In the present investigation, micronutrient composition of 35 food fishes (includes both finfishes and shellfishes) was investigated from varying aquatic habitats. Macrominerals (Na, K, Ca, Mg) and trace elements (Fe, Cu, Zn, Mn, Se) were determined by either atomic absorption spectroscopy (AAS) or inductively coupled plasma mass spectrometry (ICP-MS)/atomic emission spectrometry (ICP-AES). Phosphorus content was determined either spectrophotometrically or by ICP-AES. Fat-soluble vitamins (A, D, E, K) were analyzed by high-performance liquid chromatography (HPLC). The analysis showed that, in general, the marine fishes were rich in sodium and potassium; small indigenous fishes (SIFs) in calcium, iron, and manganese; coldwater fishes in selenium; and the brackishwater fishes in phosphorous. The marine fishes Sardinella longiceps and Epinephelus spp. and the SIFs were rich in all fat-soluble vitamins. All these recommendations were made according to the potential contribution (daily value %) of the species to the recommended daily allowance (RDA). Information on the micronutrients generated would enhance the utility of fish in both community and clinical nutrition.

Effect of storage temperature as a preanalytical variable on the lens crystallins protein quality for proteomic studies.

PURPOSE: To study the effect of storage temperature on lens crystallins quality for proteomic analysis, using αA-crystallins as internal marker. EXPERIMENTAL DESIGN: Lenses were stored at -40°C, -10°C and ice for up to 10 days. Protein extracts were prepared from samples stored at -40°C and -10°C on completion of 10 days; for samples kept under ice-storage, lenses were taken out at every 24 h, extracts prepared and stored. Fresh lens extracts served as the control. RESULTS: SDS-PAGE analysis of proteins from lens stored at -40°C and -10°C for 10 days did not show any appreciable change in profiles; however, two protein bands of 27 and 29 kDa disappeared from lens stored in ice. A time-course analysis showed that such changes in ice-stored lens occurred beyond six days of storage. Appearance of additional αA-crystallin fragments on 1-D and 2-D immunoblots confirmed degradation of αA-crystallins. Protein spots altered in abundance and identified by peptide mass fingerprinting include αA-, αB-, ßB1-, ßB2- and γ-crystallins. CONCLUSIONS AND CLINICAL RELEVANCE: For proteomic studies, quality of the starting material must be ensured to avoid erroneous and misleading interpretation of results. Under field conditions where deep freezing or immediate preparations of samples are not the options, eye lens can be transported under ice-storage for about six days without deterioration in protein quality.

Proteome analysis of the Atlantic salmon (Salmo salar) cell line SHK-1 following recombinant IFN-gamma stimulation.

Type II IFN exists as a single molecule (IFN-gamma) in contrast to type I IFN, of which there are a number of different forms. IFN-gamma is involved both directly and indirectly in antiviral activity, stimulation of bactericidal activity, antigen presentation and activation of macrophages. Recently IFN-gamma was cloned from a salmonid fish, the rainbow trout and a functional recombinant protein produced exhibited IFN-gamma activity. This recombinant IFN-gamma was used to stimulate an Atlantic salmon cell line, SHK-1, to monitor the changes in protein expression by proteomic analysis 24 h after stimulation compared to unstimulated control cells. An SHK-1 cell proteome map was developed and proteins altered in abundance by the IFN-gamma stimulation were identified. Under the analytical conditions used, 22 proteins were found to be altered in abundance, 15 increased and 7 decreased. Several proteins were excised from the gel and identified, following trypsin digestion and MALDI-MS/MS/LC-MS and database interrogation. Transcriptional analysis of five mRNAs encoding proteins increased in abundance by IFN-gamma in the proteome analysis was determined by real-time PCR. We assessed the correlation between gene expression and change in abundance of proteins for these genes.

Brugia malayi adult low molecular weight IgG4-reactive antigens induce differential cytokine response in lymphocytes of endemic normal and asymptomatic microfilariae carriers in vitro.

To characterize putatively protective immune response in bancroftian filariasis, Th1/Th2 cytokine profile induced in peripheral blood mononuclear cells (PBMCs) of endemic normal (EN) and asymptomatic microfilaremic (ASM) individuals were studied using different molecular weight fractions of Brugia malayi adult soluble antigens (BmA), which are differentially recognized by IgG4 antibodies present in their sera. Infection free and putatively immune individuals living in a filaria endemic area were identified and included in the present study as EN only after careful longitudinal follow up for three years. It was observed that the low molecular weight antigens present in Fr4 and Fr5 induced differential cytokine response; EN individuals showed a strong Th1 bias whereas ASM individuals showed a strong Th2 bias even though both the groups produced Th1 cytokines, albeit of different quantity, when a nonhelminthic antigen like H37Rv whole cell lysate was used. Since antigens present in Fr5 induced a highly polarized response, they should be examined for their diagnostic potential in lymphatic filariasis.