Genomic organization and transcription of superoxide dismutase genes (sod1, sod2, and sod3b) and response to diazinon toxicity in platyfish (Xiphophorus maculatus) by using SOD enzyme activity.

The aim of this study is to determine the effects of 50% of 96 h LC50 (5.25 ppm) diazinon on the expression of superoxide dismutase (SOD) enzyme genes (sod1, sod2, and sod3b) and SOD enzyme activity at the end of 24, 48, 72, and 96 h in platyfish liver and gill tissues. To this end, we determined the tissue-specific distribution of sod1, sod2, and sod3b genes and performed in silico analyses in platyfish (Xiphophorus maculatus). It was determined that malondialdehyde (MDA) level and SOD enzyme activity were increased in the liver [(43.90 EU mg protein-1 (control), 62.45 EU mg protein-1 (24 h), 73.17 EU mg protein-1 (48 h), 82.18 EU mg protein-1 (72 h), 92.93 EU mg protein-1 (96 h)] and gill [(16.44 EU mg protein-1 (control), 33.47 EU mg protein-1 (24 h), 50.38 EU mg protein-1 (48 h), 64.62 EU mg protein-1 (72 h), 74.04 EU mg protein-1 (96 h)] tissues of platyfish exposed to diazinon, while the expression of the sod genes was down-regulated. The tissue-specific distribution of the sod genes varied, with the tissues and the sod genes expression were being predominant in the liver (628.32 in sod1, 637.59 in sod2, 888.5 in sod3b). Thus, the liver was considered a suitable tissue for further gene expression studies. Based on the phylogenetic analyses, platyfish sod genes can be reported to be orthologs of sod/SOD genes from other vertebrates. Identity/similarity analyses supported this determination. Conserved gene synteny proved that there are conserved sod genes in platyfish, zebrafish, and humans.

Fatty acid-binding protein genes in gilthead seabream: molecular cloning and nutritional regulation under low water temperatures.

The molecular characteristics and tissue disruption of 10 fatty acid-binding protein (fabp) genes in gilthead seabream (Sparus aurata) were investigated, and their expression levels were found in the fish fed diets with different vegetable oil (VO) sources, which may explore the potential function of fabp genes in S. aurata. For this purpose, the open reading frames of fabp genes involved in the transport and ß-oxidation of fatty acids (FA) were molecularly cloned and characterized. S. aurata was then exposed to a two-staged feeding trial (the grow-out period following a wash-out period) at low water temperatures. In the grow-out period, the fish were fed diets containing 50% and 100% ratios of various VOs for 60 days, and in the wash-out period, the fish were fed a diet containing 100% fish oil (FO) for 30 days. It has been determined that (a) S. aurata and vertebrate fabp/FABP genes are orthologues; (b) spatio-temporal differences in tissue-specific patterns of fabp genes differ importantly; for instance, the difference between the highest and lowest values reaches 13 × 105 -fold in the fabp10a; and (c) VO-based diets upregulated fabp transcript levels in the liver and muscle with some exceptions, such as liver fabp11a and muscle fabp7a. Gene expressions of only the hepatic fabp7b and fabp10a genes were diminished at the end of the wash-out period. In this study, the authors provide further evidence that dietary FAs affect fabp mRNA expressions in S. aurata. This might be useful in the nutritional control of fabp genes to maintain lipid homeostasis in marine fish fed VO-based diets at low water temperatures.

Molecular cloning and characterization of Cu-Zn superoxide dismutase (sod1) gene in brown trout and its expression in response to acute aquaculture stressors.

Aquaculture species are often exposed to acute stressors such as low water levels and handling during routine aquaculture procedures. This might result in oxidative stress by the increased reactive oxygen species (ROS)' production (e.g., superoxide anion). The harmful effects of ROS are eliminated by a defense system, referred antioxidant defense system (ADS). sod1 is the first gene involved in the ADS. Therefore, we cloned and characterized the open reading frame of the sod1 in brown trout. Then, we determined the effects of low water level and handling stress on sod1 mRNA expression in the liver and gills at 0 min, 1 and 2 h. The total RNA isolated was used to synthesize cDNA for RT-qPCR analysis. Phylogenetic tree, identity/similarity percentages, genomic organization, and conserved gene synteny analyses were applied to characterize Sod1/sod1. While low water level stress upregulated sod1 expression in the liver compared to the control group, no significant differences were observed in the gills between experimental groups. However, brown trout differently responded to handling stress at different time intervals in both tissues. Transcriptional differences were also noted between the sexes. This study contributes to the current understanding of the molecular mechanism between oxidative stress and ADS.

Identification and Characterization of Carnitine Palmitoyltransferase 1 (cpt 1) Genes in Nile Tilapia, Oreochromis niloticus.

Four cpt 1 genes (cpt 1α1a, cpt 1α2a, cpt 1α2b, and cpt 1ß) were identified in the Nile tilapia genome. Two transmembrane helix domains (TMH) were identified for Cpt 1α1a, Cpt 1α2a, and Cpt 1ß, while Cpt 1α2b had only one TMH domain. Evidence was found of conserved gene synteny between cpt 1 genes from Nile tilapia and the cpt 1/CPT 1 genes of zebrafish and human. Phylogenetic analysis showed that Nile tilapia Cpt 1 sequences clustered in distinct clades with their orthologous Cpt 1/CPT 1 from other vertebrates. Nile tilapia cpt 1α1a, cpt 1α2a, cpt 1α2b, and cpt 1ß contain 18 coding exons encoding polypeptides of 771, 784, 788, and 786 amino acids in length, respectively. The cpt 1 genes were determined in all the tested tissues with varying tissue distribution patterns. These findings suggest that (1) cpt 1α1a, cpt 1α2a, and cpt 1α2b arose in the Nile tilapia genome as a result of the teleost-specific whole-genome duplication; (2) nonfunctionalization is the most likely cause of the loss of cpt 1α1b in the Nile tilapia genome; (3) the different tissue-specific transcription of cpt 1α2a and cpt 1α2b may be either due to the sub- or the neo-functionalization of transcriptional control side.

Divergent spatial regulation of duplicated fatty acid-binding protein (fabp) genes in rainbow trout (Oncorhynchus mykiss).

The increased use of plant oil as a dietary supplement with the resultant high dietary lipid loads challenges the lipid transport, metabolism and storage mechanisms in economically important aquaculture species, such as rainbow trout. Fatty acid-binding proteins (Fabp), ubiquitous in tissues highly active in fatty acid metabolism, participate in lipid uptake and transport, and overall lipid homeostasis. In the present study, searches of nucleotide sequence databases identified mRNA transcripts coded by 14 different fatty acid-binding protein (fabp) genes in rainbow trout (Oncorhynchus mykiss), which include the complete minimal suite of seven distinct fabp genes (fabp1, 2, 3, 6, 7, 10 and 11) discovered thus far in teleost fishes. Phylogenetic analyses suggest that many of these extant fabp genes in rainbow trout exist as duplicates, which putatively arose owing to the teleost-specific whole genome duplication (WGD); three pairs of duplicated fabp genes (fabp2a.1/fabp2a.2, fabp7b.1/fabp7b.2 and fabp10a.1/fabp10a.2) most likely were generated by the salmonid-specific WGD subsequent to the teleost-specific WGD; and fabp3 and fabp6 exist as single copy genes in the rainbow trout genome. Assay of the steady-state levels of fabp gene transcripts by RT-qPCR revealed: (1) steady-state transcript levels differ substantially between fabp genes and, in some instances, by as much as 30×10(4)-fold; (2) some fabp transcripts are widely distributed in many tissues, whereas others are restricted to one or a few tissues; and (3) divergence of regulatory mechanisms that control spatial transcription of duplicated fabp genes in rainbow trout appears related to length of time since their duplication. The suite of fabp genes described here provides the foundation to investigate the role(s) of fatty acid-binding proteins in the uptake, mobilization and storage of fatty acids in cultured fish fed diets differing in lipid content, especially the use of plant oil as a dietary supplement. These nutritional dietary supplements may well lead to high lipid loads with the resultant challenges to lipid homeostasis and, thus, health of cultivated fish which may be mediated by appropriate transcriptional control of fabp genes.

Goose fat, a promising nutrient for fish feeding, activates antioxidant enzymes in rainbow trout, Oncorhynchus mykiss.

The objective of this experiment was to test effects of different dietary lipids in rainbow trout feeding on the activity and expression of antioxidant enzymes, superoxide dismutase (SOD), glutathione peroxidase (GPx) and glutathione S-transferase (GST). Four iso-nitrogenous and iso-lipidic casein-gelatin based experimental diets were formulated. The sources of dietary lipids were cod liver oil (CO, rich in polyunsaturated fatty acids), goose fat (GF, rich in saturated fatty acids and monounsaturated fatty acids), soybean oil (SO, rich in linoleic acid), and a blend of CO, GF and SO. Dietary treatments had no significant effect on growth performance and survival was not affected. SOD, GPx and GST enzymes had the maximum activity in GF diet. However qPCR data showed that SOD and GPx mRNA levels were minimum in GF group. Overall data showed that rainbow trout liver enzymes were activated upon GF diet probably activating the enzyme structure itself without stimulating gene expression.

Alterations in fatty acids of polar lipids in Salmo trutta on long-term exposure to a glyphosate-based herbicide (Roundup).

Abstract: In present study, the effects of sublethal doses (10 and 20 mg L(-1)) of Roundup on fatty acid pattern in muscle and liver of brown trout were investigated. For this purpose, fish were held in experiment tanks for 1 month. While total MUFA wasn't influenced, the highest total SFA and total n-6 PUFA were determined in group 10 mg L(-1) and the lowest values were determined in control group and group 20 mg L(-1) in muscle, respectively. The highest and the lowest total n-3 PUFA was found in control group and group of 10 mg L(-1) in muscle, respectively. Total n-3/n-6 PUFA ratio and EPA+DHA level of group 10 mg L(-1) were lower than other groups in muscle. The amount of total n-3/n-6 PUFA, EPA + DHA and total n-3 PUFA of control group were found higher than treatment groups in liver. While the highest total SFA was determined in group 10 mg L(-1), there wasn't difference between control group and group 20 mg L(-1) in liver. Both of doses herbicide had higher value than control for total MUFA in liver. While Roundup didn't inhibit n-3 PUFA synthesis in the muscle, both concentrations, exhibited inhibitory effect on n-3 PUFA synthesis in the liver. This result probably consequence of its indirect effect on the some enzyme activities or gene expressions in fatty acid metabolism of brown trout.

Metabolic responses to prolonged starvation, food restriction, and refeeding in the brown trout, Salmo trutta: oxidative stress and antioxidant defenses.

The effects of long-term starvation and food restriction (49 days), followed by refeeding (21 days) have been studied with respect to antioxidant defense in the liver and gills (branchial tissues) of the brown trout, Salmo trutta. Malondialdehyde levels in both tissues increased in parallel with starvation and food restriction and these values did not return to normal after the refeeding period. The activities of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), and glutathione reductase (GR) in liver and gills increased during the 49 days of starvation, but glucose-6-phosphate dehydrogenase (G6PD) activities decreased. Glutathione S-transferase (GST) activity decreased in the liver at the 49th day of starvation, but increased in the branchial tissues. Some of the antioxidant enzyme activities (such as hepatic GST and branchial G6PD) returned to control values of fed fish after the refeeding period, but others (e.g. hepatic SOD and branchial GPx) did not return to normal values. In conclusion, our study indicates that total or partial food deprivation induces oxidative stress in brown trout.