Speciation-specific Cr bioaccumulation, morphologic and transcriptomic response in liver of Plectropomus leopardus exposed to dietary Cr(III) and Cr(VI).

Trivalent chromium (Cr(III)) and hexavalent chromium (Cr(VI)) are the two mainly stable oxidation states of Cr in aquatic environments, while the difference of their bioavailability and toxicity by dietary exposure has been rarely known in aquatic organisms. Using juvenile coral trout (Plectropomus leopardus), Cr(III) and Cr(VI) as model system, this study tested the hypothesis that the dietary Cr bioaccumulation and toxicity in fish were highly dependent on Cr speciation. The fish were chronically exposed to 200 mg kg-1 of dietary Cr(III) and Cr(VI) for 8 weeks, and then the Cr bioaccumulation, morphologic change, and RNA-Seq in fish liver were determined. The results showed that dietary Cr(III) and Cr(VI) exposure significantly induced fish weight gain, while 1.17 folds and 1.26 folds increased in relation to Control group, respectively. Cr contents in liver was increased significantly in dietary Cr(VI) but not in Cr(III) groups. Both Cr treatment induced lipid deposition in liver tissue structure, moreover, pancreatic part was increased in dietary Cr(III) but its reduced in Cr(VI) exposure. RNA-Seq in fish liver were significantly different as well. Specifically, there were 138 differentially expressed genes (DEGs) in dietary Cr(III) group, including 76 up-regulated and 62 down-regulated, and these DEGs were mainly involved in lipid metabolism, while there were 175 DEGs in dietary Cr(VI) group, including 85 up-regulated and 90 down-regulated, and these DEGs were mainly involved in immune system. The qRT-PCR confirmed the RNA-seq data were reliable. Overall, these results supported our hypothesis that the chronic dietary Cr(III) and Cr(VI) exposure resulted in apparently different Cr bioaccumulation and toxicity in fish. Our findings here help us to fill in a big gap in our knowledge of speciation-specific Cr bioavailability and toxicity in aquatic organisms, which has been largely unclear previously. CAPSULE: Dietary Cr(III) increased lipid metabolism and dietary Cr(VI) activated immune system in liver of coral trout at transcription levels.

Waterborne Cr3+ and Cr 6+ exposure disturbed the intestinal microbiota homeostasis in juvenile leopard coral grouper Plectropomus leopardus.

BACKGROUND: Chromium (Cr) mainly has two stable forms: Cr3+ and Cr6+. Cr and its compound are widely used in the printing, dyeing, leather making, and metallurgy industry. They are evitably released into the environment and pose a significant threat to creatures, for instance, the excessive chromium (Cr) burden in the marine ecosystem is often harmful to fish. Intestinal microbiota greatly affects fish performance, but how waterborne Cr affects fish intestinal microbiota is unclear. To test the hypothesis that the waterborne Cr exposure could significantly affect fish' intestinal microbiota homeostasis, and the effect was highly dependent on Cr concentration and speciation, the juvenile leopard coral grouper Plectropomus leopardus were exposed to waterborne Cr3+ and Cr6+ (0.1, 0.5 ppm) for 7 days, and the intestinal microbiota was determined by Amplicon sequencing of the 16S rRNA. RESULTS: In all Cr treatment groups, the alpha diversity of intestinal microbiota communities of P. leopardus was decreased. The phyla Proteobacteria, Firmicutes, Actinobacteria, Chloroflexi and Bacteroidetes were the dominant intestinal microbiota. The Chao index diversity significantly declined in Cr treatment group, indicating the intestinal microbiota community structure was changed. Among the dominant intestinal microbiota, Proteobacteria was most sensitive to Cr exposure, and it increased after xposure. The PICRUSt predicted that 0.5 ppm Cr3+ expousure caused metabolism disordered in the intestinal of P. leopardus. CONCLUSIONS: Waterborne Cr3+ and Cr6+ significantly disturbed intestinal microbiota homeostasis in P. leopardus, including their diversity, composition, and community structure. The metabolism level of intestinal microbiota in P. leopardus was decreased by Cr3+ exposure. High concentrations of Cr3+ may pose potential risks to the intestinal homeostasis of P. leopardus.

Characterization and functional analysis of myostatin and myogenin genes involved in temperature variation and starvation stress in Golden pompano, Trachinotus blochii.

Animal growth and development is a complicated process and is regulated by multi-genes. Myostatin (Mstn) and myogenin (Myog) are a pair of negative and positive regulators respectively, which play an important role in the generation of muscle cells. In order to study the function of these two genes in muscle growth of Trachinotus blochii, full lengths of two mstn genes (mstn-1 and mstn-2) and myog gene were cloned using RACE. We first identified and characterized the complete cDNA sequences of mstn-1, mstn-2, and myog genes derived from T. blochii, an economically important mariculture species in China. Multiple sequence alignment of amino acids and phylogenetic analysis revealed that the Mstn and Myog were highly conserved to the other Perciformes. In addition, gene duplication of mstn in T. blochii was observed. mstn-1 mRNA was mainly expressed in the muscle and gonad, while mstn-2 and myog transcripts were detectable mainly in the brain and muscle, respectively. Moreover, the nutritional status and temperature influenced abundance levels in brain and muscle. Results suggested that mstn and myog genes play an important role in muscle growth of T. blochii, mstn may not be limited to control of muscle growth in fish and could also be involved in other biological functions.

Waterborne zinc bioaccumulation influences glucose metabolism in orange-spotted grouper embryos.

Fish embryos, as an endogenous system, strictly regulate an energy metabolism that is particularly sensitive to environmental pressure. This study used orange-spotted grouper embryos and stable isotope 67Zn to test the hypothesis that waterborne Zn exposure had a significant effect on energy metabolism in embryos. The fish embryos were exposed to a gradient level of waterborne 67Zn, and then sampled to quantify 67Zn bioaccumulation and mRNA expressions of key genes involved glucose metabolism. The results indicated that the bioaccumulated 67Zn generally increased with increasing waterborne 67Zn concentrations, while it tended to be saturated at waterborne 67Zn > 0.7 mg L-1. As we hypothesized, the expression of PK and PFK gene involved glycolysis pathway was significantly up-regulated under waterborne 67Zn exposure >4 mg L-1. Waterborne 67Zn exposure >2 mg L-1 significantly suppressed PCK and G6PC gene expression involved gluconeogenesis pathway, and also inhibited the AKT2, GSK-3beta and GLUT4 genes involved Akt signaling pathway. Our findings first characterized developmental stage-dependent Zn uptake and genotoxicity in fish embryos. We suggest fish embryos, as a small-scale modeling biosystem, have a large potential and wide applicability for determining cytotoxicity/genotoxicity of waterborne metal in aquatic ecosystem.

Dietary Trivalent Chromium Exposure Up-Regulates Lipid Metabolism in Coral Trout: The Evidence From Transcriptome Analysis.

Diet quality greatly affects an animal's performance and metabolism. Despite the fact that trivalent chromium [Cr(III)] is considered an essential element and is widely used in nutritional supplements for animals and humans, the potential toxicity of Cr(III) is unclear. Here, liver transcriptome sequencing was performed on coral trout (Plectropomus leopardus) exposed to 200 mg kg-1 of dietary organic Cr(III) [as chromium picolinate (CrPic)] for 8 weeks. One-hundred-and thirteen differentially expressed genes (DEGs) were identified in response to Cr(III) stress, in comparison to the control, including 31 up-regulated and 82 down-regulated DEGs. Clusters of Orthologous Groups of proteins (COG) classifies DEGs into 15 functional categories, with the predominant category being related to lipid transport and metabolism (9.73%). The Kyoto Encyclopedia of Genes and Genomes (KEGG) assigned DEGs to six major categories with robust DEGs as part of the lipid metabolism pathway (18.58%). Moreover, KEGG functional enrichment analysis showed that these DEGs are primarily related to steroid biosynthesis, terpenoid backbone biosynthesis, and steroid hormone biosynthesis pathways, of which steroid biosynthesis was the most significant pathway, and 12 key up-regulated DEGs (dhcr7, dhcr24, ebp, lss, msmo1, sqle, cyp51, tm7sf2, sc5dl, fdft1, nsdhl, and hsd17b7) were found for steroid biosynthesis pathways. To validate the RNA sequencing data using quantitative real-time PCR (qRT-PCR), qRT-PCR results indicate that the expression of genes encoding HMGCR, TM7SF2, TRYP2, CTRL, EBP, LSS, and CYP51 were induced, while those encoding THRSP, LCE, and MCM5 were reduced, consistent with RNA-seq results. This findings provides the first evidence that a long-term high dose of Cr(III) intake causes lipid metabolism disorder and potential toxicity in fish. Cautious health risk assessment of dietary Cr(III) intake is therefore highly recommended for the commercial and/or natural diets of aquatic animals, which has previously largely been ignored.

Effects of temperature on the transcriptomes of pituitary and liver in Golden Pompano Trachinotus blochii.

Fish growth can be modulated dynamically through the brain-pituitary-liver regulation axis. In the present study, whole transcriptomes of the pituitary and liver from Golden Pompano Trachinotus blochii were sequenced in seawater at 20 °C (T_low) and 25 °C (T_high). A total of 187,277,583 paired-end reads were assembled to obtain 100,495 transcripts, corresponding to 83,974 genes. These reads were mapped to T. blochii transcripts, and the mapping accuracy ranged from 80.4 to 94.9%. Two lists of differentially expressed genes were obtained by comparisons of pituitary and liver T_low versus T_high groups, comprising 458 and 205 genes, respectively. Of these, 33 differentially expressed genes were common between the two lists. Twelve GO terms were overrepresented for the 458 differentially expressed genes in the pituitary, and it is noteworthy that the GO term galanin receptor activity (GO: 0004966) related to the modulation of appetite and metabolism, whose genes made up half of all assembled genes in the term. For the 205 differentially expressed genes in the liver, 19 overrepresented GO terms were mainly related to immune regulation, digestion, and protein metabolism. Among the common differentially expressed genes, there were 32 genes that had identical changing trends in both pituitary and liver comparisons. Furthermore, two GO terms inorganic diphosphatase activity and MHC protein complex were overrepresented. These results indicate that the brain could regulate pituitary function through galanin signal mechanism and that the metabolism of liver was further optimized to modulate immunity and growth under different temperatures.

Simultaneous uptake of Cd from sediment, water and diet in a demersal marine goby Mugilogobius chulae.

The embryonic state of our knowledge regarding the simultaneous uptake of trace metals via multiple routes in aquatic organisms makes it difficult to accurately assess the bioaccumulation and risk of metals. This study used cadmium (Cd) and a demersal marine fish (the yellowstripe goby) as a model system to determine tissue-specific uptake of Cd under conditions of simultaneous exposure to Cd from water, sediment and diet. A triple stable isotope tracing method was used in which each exposure route was spiked by a different stable isotope (110Cd, 111Cd and 113Cd). The results revealed that the fish took up waterborne and sedimentary Cd via gills and gastrointestinal tract (GT), and that of dietary Cd was via the GT. The gills absorbed Cd predominantly from water (77.2-89.4%) whilst the GT absorbed Cd mainly from diet (81.3-98.7%). In the muscle and carcass, the Cd uptake was mainly from the diet (47.1-80.4%) and water (22.8-51.6%). Our study demonstrated that when aquatic animals were subject to simultaneous exposure through multiple uptake routes, the uptake and relative importance of each route for metal accumulation was highly tissue-specific and more complex than a single route of metal exposure.

Biokinetic Modeling of Cd Bioaccumulation from Water, Diet and Sediment in a Marine Benthic Goby: A Triple Stable Isotope Tracing Technique.

Aquatic animals are often simultaneously exposed to metals through multiple routes in the natural environment. This study explored a triple stable isotope tracing method to quantify simultaneous cadmium (Cd) uptake biokinetics by yellow stripe goby from water (traced by 110Cd), sediment (traced by 111Cd), and diet (traced by 113Cd) when the fish were exposed to Cd for 24 h. The simultaneous uptake of Cd from multiple routes during 4 weeks was then predicted by the modified biokinetic model. The results demonstrated that the uptake rate constant of waterborne 110Cd, sediment-associated 111Cd, and dietary 113Cd was 3.1 L kg-1 d-1, 2.2 × 10-4 g g-1 d-1, and 3.3 × 10-3 g g-1 d-1 in the fish. Sedimentary Cd was less bioavailable than the waterborne and dietary Cd; however, sediment could become the predominant Cd source of the total Cd bioaccumulation when the partition coefficient of Cd between sediment and seawater ( Kd) is larger than 6 × 104 L kg-1. The simultaneous uptake of Cd from the three routes could be successfully predicted by the modified model. The model revealed that the Cd bioaccumulation generally increased with the increase of ambient Cd concentration in all the three routes. Overall, our findings demonstrated that the multiple stable isotopes tracing method and the modified biokinetic model have a wide generality and applicability for predicting Cd bioaccumulation under multiple routes of metal exposure scenario and may have application to other metals.