Genome-wide identification and structural analysis of the BMP gene family in Triplophysa dalaica.

BACKGROUND: Bone morphogenetic proteins (BMPs) are part of the transforming growth factor beta (TGF-ß) superfamily and play crucial roles in bone development, as well as in the formation and maintenance of various organs. Triplophysa dalaica, a small loach fish that primarily inhabits relatively high elevations and cooler water bodies, was the focus of this study. Understanding the function of BMP genes during the morphogenesis of T. dalaica helps to clarify the mechanisms of its evolution and serves as a reference for the study of BMP genes in other bony fishes. The data for the T. dalaica transcriptome and genome used in this investigation were derived from the outcomes of our laboratory sequencing. RESULTS: This study identified a total of 26 BMP genes, all of which, except for BMP1, possess similar TGF-ß structural domains. We conducted an analysis of these 26 BMP genes, examining their physicochemical properties, subcellular localization, phylogenetic relationships, covariance within and among species, chromosomal localization, gene structure, conserved motifs, conserved structural domains, and expression patterns. Our findings indicated that three BMP genes were associated with unstable proteins, while 11 BMP genes were located within the extracellular matrix. Furthermore, some BMP genes were duplicated, with the majority being enriched in the GO:0008083 pathway, which is related to growth factor activity. It was hypothesized that genes within the BMP1/3/11/15 subgroup (Group I) play a significant role in the growth and development of T. dalaica. By analyzing the expression patterns of proteins in nine tissues (gonad, kidney, gill, spleen, brain, liver, fin, heart, and muscle), we found that BMP genes play diverse regulatory roles during different stages of growth and development and exhibit characteristics of division of labor. CONCLUSIONS: This study contributes to a deeper understanding of BMP gene family member expression patterns in high-altitude, high-salinity environments and provides valuable insights for future research on the BMP gene family in bony fishes.

Spexin acts as a novel glucose-lowering factor in grass carp (Ctenopharyngodon idella).

At present, the physiological roles of various hormones in fish glucose metabolism have been elucidated. Spexin, a 14-amino acids polypeptide, is highly conserved in many species and has functions such as reducing body weight and improving insulin resistance. In this paper, the open reading frame (ORF) of spx21 in grass carp (Ctenopharyngodon idella) was cloned, and the tissue distribution of spx1 and spx2, their direct and indirect regulatory effects on glucose metabolism of grass carp were investigated. The ORF of spx2 gene in grass carp was 279 bp in length. Moreover, spx1 was highly expressed in the adipose tissue, while spx2 was highly expressed in the brain. In vitro, SPX1 and SPX2 showed opposite effects on the glycolytic pathway in the primary hepatocytes. In vivo, intraperitoneal injection of SPX1 and SPX2 significantly reduced serum glucose levels and increased hepatopancreas glycogen contents. Meanwhile, SPX1 and SPX2 promoted the expression of key genes of glycolysis (pk) and glycogen synthesis (gys) in the hepatopancreas at 3 h post injection. As for indirect effects, 1000 nM SPX1 and SPX2 significantly increased insulin-mediated liver type phosphofructokinase (pfkla) mRNA expression and enhanced the inhibitory effects of insulin on glucose-6-phosphatase (g6pase), phosphoenolpyruvate carboxykinase (pepck), glycogen phosphorylase L (pygl) mRNA expression. Our results show that SPX1 and SPX2 have similar indirect effects on the regulation of glucose metabolism that enhance insulin activity, but they exhibit opposite roles in terms of direct effects.

Microbacterium salsuginis sp. nov., a halotolerant actinobacterium with antimicrobial activity.

A novel Gram-staining-positive actinobacterium with antimicrobial activity, designated CFH 90308T, was isolated from the sediment of a salt lake in Yuncheng, Shanxi, south-western China. The isolate exhibited the highest 16S rRNA gene sequence similarities to Microbacterium yannicii G72T, Microbacterium hominis NBRC 15708T and Microbacterium xylanilyticum S3-ET (98.5, 98.4 and 98.2 %, respectively), and formed a separate clade with M. xylanilyticum S3-ET in phylogenetic trees. The strain grew at 15-40 ºC, pH 6.0-8.0 and could tolerate NaCl up to a concentration of 15 % (w/v). The whole genome of strain CFH 90308T consisted of 4.33 Mbp and the DNA G+C content was 69.6 mol%. The acyl type of the peptidoglycan was glycolyl and the whole-cell sugars were galactose and mannose. The cell-wall peptidoglycan mainly contained alanine, glycine and lysine. The menaquinones of strain CFH 90308T were MK-12, MK-13 and MK-11. Strain CFH 90308T contained anteiso-C15:0, anteiso-C17:0, iso-C16:0 and iso-C15:0 as the predominant fatty acids. The average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) values between CFH 90308T and the other species of the genus Microbacterium were found to be low (ANIb <81.3 %, dDDH <25.6 %). The secondary metabolite produced by strain CFH 90308T showed antibacterial activities against Bacillus subtilis, Pseudomonas syringae, Aeromonas hydrophila and methicillin-resistant Staphylococcus aureus. Based on genotypic, phenotypic and chemotaxonomic results, the isolate is considered to represent a novel species of the genus Microbacterium, for which the name Microbacterium salsuginis sp. nov. is proposed. The type strain is CFH 90308T (=DSM 105964T=KCTC 49052T).

Flagellimonas halotolerans sp. nov., a novel bacterium isolated from the South China Sea.

Two Gram-stain-negative strains, designed SYSU M86414T and SYSU M84420, were isolated from marine sediment samples of the South China Sea (Sansha City, Hainan Province, PR China). These strains were aerobic and could grow at pH 6.0-8.0 (optimum, pH 7.0), 4-37 °C (optimum, 28 °C), and in the presence of 0-10 % NaCl (w/v; optimum 3 %). The predominant respiratory menaquinone of strains SYSU M86414T and SYSU M84420 was MK-6. The primary cellular polar lipid was phosphatidylethanolamine. The major cellular fatty acids (>10 %) in both strains were iso-C15 : 0, iso-C15 : 1 G, and iso-C17 : 0 3-OH. The DNA G+C content of strains SYSU M86414T and SYSU M84420 were both 42.10 mol%. Phylogenetic analyses based on 16S rRNA gene sequences and core genes indicated that these novel strains belonged to the genus Flagellimonas and strain SYSU M86414T showed the highest 16S rRNA gene sequence similarity to Flagellimonas marinaquae JCM 11811T (98.83 %), followed by Flagellimonas aurea BC31-1-A7T (98.62 %), while strain SYSU M84420 had highest 16S rRNA gene sequence similarity to F. marinaquae JCM 11811T (98.76 %) and F. aurea BC31-1-A7T (98.55 %). Based on the results of polyphasic analyses, strains SYSU M86414T and SYSU M84420 should be considered to represent a novel species of the genus Flagellimonas, for which the name Flagellimonas halotolerans sp. nov. is proposed. The type strain of the proposed novel isolate is SYSU M86414T (=GDMCC 1.3806T=KCTC 102040T).

Improvement of sea buckthorn (Hippophae rhamnoides L.) flavonoids on the antioxidant and immune performance of Yellow River carp (Cyprinus carpio L.) fed high-carbohydrate diet.

High-carbohydrate (HC) diets may lead to the deterioration of the antioxidant and immune properties of Yellow River carp and the healthy development of the industry. Studies in mammals have found that sea buckthorn flavonoids (SF) improve antioxidant and immune performance. Therefore, this study comprehensively evaluated the effects of SF on Yellow River carp using in vitro and feeding trials with an HC diet. Control (C, 27.23 %), high-carbohydrate (HC, 42.99 %), and HC + SF (0.1 %, 0.2 %, and 0.4 %) groups were studied in a 10-week aquaculture experiment. The main findings were as follows: (1) SF scavenged O2·-, ·OH, and DPPH free radicals in vitro, which gradually increased with the SF concentration. (2) The antioxidant and immune performance of Yellow River carp was enhanced by dietary supplementation with SF, which involved the regulation of activities of antioxidant and immune enzymes, as well as their changes at the transcription and protein levels. In terms of antioxidant properties, compared to the HC group, HC + SF significantly decreased the activities of glutamic-oxaloacetic transaminase and glutamic-pyruvic transaminase and the contents of H2O2 and malondialdehyde in the serum and hepatopancreas. The activities of glutathione, glutathione-Px, superoxide dismutase, catalase, and total antioxidant activity in the HC-diet group. In contrast, the addition of SF increased antioxidant enzyme activity. In the hepatopancreas and muscles, SF regulated and activated Nrf2-Keap1, a key signaling pathway for oxidative stress. SF significantly increased the mRNA expression levels of downstream genes (gr, ho-1, cat, and sod) regulated by nrf2. In terms of immune performance, 0.4 % SF markedly increased the activity of immune-related enzymes. SF inhibited the gene expression of pro-inflammatory factors induced by the HC diet and promoted the gene expression of anti-inflammatory factors. In addition, the resistance of Yellow River carp to Aeromonas hydrophila was enhanced by SF. In summary, SF supplementation can reduce oxidative stress and inflammatory harm caused by the HC diet and improve the antioxidant and immune performance of Yellow River carp to varying degrees.

Environmental cadmium exposure perturbs systemic iron homeostasis via hemolysis and inflammation, leading to hepatic ferroptosis in common carp (Cyprinus carpio L.).

Cadmium (Cd) pollution is considered a pressing challenge to eco-environment and public health worldwide. Although it has been well-documented that Cd exhibits various adverse effects on aquatic animals, it is still largely unknown whether and how Cd at environmentally relevant concentrations affects iron metabolism. Here, we studied the effects of environmental Cd exposure (5 and 50 µg/L) on iron homeostasis and possible mechanisms in common carp. The data revealed that Cd elevated serum iron, transferrin saturation and iron deposition in livers and spleens, leading to the disruption of systemic iron homeostasis. Mechanistic investigations substantiated that Cd drove hemolysis by compromising the osmotic fragility and inducing defective morphology of erythrocytes. Cd concurrently exacerbated hepatic inflammatory responses, resulting in the activation of IL6-Stat3 signaling and subsequent hepcidin transcription. Notably, Cd elicited ferroptosis through increased iron burden and oxidative stress in livers. Taken together, our findings provide evidence and mechanistic insight that environmental Cd exposure could undermine iron homeostasis via erythrotoxicity and hepatotoxicity. Further investigation and ecological risk assessment of Cd and other pollutants on metabolism-related effects is warranted, especially under the realistic exposure scenarios.

Identification of Isthmin-1 in common carp (Cyprinus carpio L.) and the effects on glucose metabolism in vivo and in vitro.

Isthmin-1 (Ism1) plays roles in glucose uptake in mammals as an adipokine. To investigate its role in the glucose metabolism of common carp (Cyprinus carpio. L), the Ism1 sequence was cloned, and its expression and distribution in tissues were detected. In addition, we prepared and purified the recombinant Ism1 protein using the E. coli expression system and assessed changes in the expression of key genes related to glucose metabolism through both in vivo injection experiments and primary hepatocyte experiments in vitro. The results revealed that the open reading frame of Ism1 was 1377 bp long, encoding 458 amino acids. Similarity analysis indicated that Ism1 exhibited a close evolutionary relationship with goldfish (Carassius auratus), sharing 98.35% amino acid similarity. Ism1 was expressed in all tissues of common carp, with the highest level observed in the heart, followed by the gill, head kidney, and hepatopancreas. Distinct patterns of Ism1 expression were identified during the oral glucose tolerance test and long-term high-carbohydrate and high-fat diet feeding experiments. In vivo studies demonstrated that the serum glucose concentration was reduced on treatment with Ism1, accompanied by a significant upregulation of mRNA levels for gk, hk, and pfk genes in hepatopancreas; conversely pepck and g6pase mRNA levels were significantly downregulated in the hepatopancreas under these conditions as well. Furthermore, our primary hepatocyte experiment confirmed that Ism1 could inhibit pepck and g6pase mRNA expression, while promoting gk, hk, and pfk mRNA expression levels. In conclusion, Ism1, in common carp, could participate in the glucose metabolism, which provides essential information for future studies on the function of Ism1.

In vivo evidence of sea buckthorn relieving oxidative stress and improving immune performance of common carp (Cyprinus carpio L.).

BACKGROUND: Sea buckthorn has the functions of antioxidation, antitumor, anti-inflammation and regulating energy metabolism. In order to investigate the effects of sea buckthorn powder and sea buckthorn flavonoids on the antioxidant properties, immune function and muscle fatty acid composition of common carp, an oral feeding experiment was carried out. RESULTS: The administration of glucose significantly reduced the levels of glutathione and the activity of total antioxidant capacity enzyme in serum and hepatopancreas, while concurrently upregulating the level of malondialdehyde (MDA)(P < 0.05). Conversely, oral intake of sea buckthorn powder and flavonoids increased antioxidant enzyme activity and decreased MDA levels. In terms of antioxidant molecular indicators, sea buckthorn powder and sea buckthorn flavonoids significantly increased the mRNA levels of nuclear factor NF-E2-related factor (nrf2) in the hepatopancreas and muscle. Meanwhile, mRNA expression levels of downstream antioxidant-related genes (gr, cat, gpx, and sod) regulated by Nrf2 were also upregulated. In the immune aspects, the mRNA expression levels of proinflammatory cytokines, such as interleukin-6 (il-6), interleukin-1ß (il-1ß) and nuclear factor-κB (nf-κb), were reduced but the expressions of anti-inflammatory cytokines, such as growth factor-ß (tgf-ß) and interleukin-10 (il-10), were enhanced in the head kidney and spleen tissues after oral administration with sea buckthorn. In terms of muscle fatty acid composition, the ratio of n-3 polyunsaturated fatty acid (PUFA)/n-6 PUFA was notably higher after administering sea buckthorn flavonoids than that of the glucose group (P < 0.05). CONCLUSION: This study demonstrated that oral administration of sea buckthorn powder and sea buckthorn flavonoids significantly enhanced the antioxidant capacity and immune response and improved the muscle fatty acid compositions in common carp, and also mitigated the adverse effects of glucose treatment to a certain extent. © 2024 Society of Chemical Industry.

Cereibacter flavus sp. nov., a novel member of the family Rhodobacteraceae isolated from seawater of the South China Sea and reclassification of Rhodobacter alkalitolerans as Cereibacter alkalitolerans comb. nov.

A Gram-stain-negative, aerobic, motile, ovoid-shaped and yellow-coloured strain, designated SYSU M79828T, was isolated from seawater collected from the South China Sea. Growth of this strain was observed at 4-37 °C (optimum, 28 °C), pH 6.0-8.0 (optimum, pH 7.0) and with 0-6% NaCl (optimum, 3.0 %, w/v). The respiratory quinone was found to be Q-10. Major fatty acid constituents were C18 : 1 ω7c/C18 : 1 ω6c, C18 : 1 ω7c11-methyl and C18 : 0 (>5 % of total). The major polar lipids were phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol, phosphatidylcholine, phosphoglycolipid, two unidentified phospholipid, one unidentified lipid and an unidentified glycolipid. The genomic DNA G+C content was 64.5 mol%. Phylogenetic analyses based on 16S rRNA gene sequences and core genes indicated that strain SYSU M79828T belonged to the genus Cereibacter and had the highest sequences similarity to 'Rhodobacter xinxiangensis' TJ48T (98.41 %). Based on 16S rRNA gene phylogeny, physiological and chemotaxonomic characterizations, we consider that strain SYSU M79828T represents a novel species of the genus Cereibacter, for which the name Cereibacter flavus sp. nov. is proposed. The type strain is SYSU M79828T (=GDMCC 1.3803T=KCTC 92893T). In addition, according to the results of phylogenetic analysis and similar taxonomic characteristics, we propose that Rhodobacter alkalitolerans should be reclassified as Cereibacter alkalitolerans comb. nov.

Establishment and characterization of Yellow River carp (Cyprinus carpio haematopterus) muscle cell line and its application to fish virology and immunology.

The Yellow River carp (Cyprinus carpio haematopterus) is a vital economically farmed fish of the Cyprinidae family. With the development of intensive aquaculture, carp production has increased dramatically, leading to the frequent occurrence of various diseases. Cell lines are considered the most cost-effective resource for in vitro studies and are widely used for physiological and pathological studies because of accessibility and convenience. This research established a novel immortal cell line CCM (Yellow River carp muscle cells) derived from the carp muscle. CCM has been passed over 71 generations for 1 year. The morphology of CCM and the adhesion and extension processes were captured by light and electron microscopy. CCM were passaged every 3 days with 20% FBS DMEM/F12 at 1:3. The optimum conditions for CCM growth were 28 °C and 20% FBS concentration. DNA sequencing of 16S rRNA and COI showed that CCM was derived from carp. CCM positively reacts to anti-PAX7 and anti-MyoD antibodies of carp. Analysis of chromosomes revealed that the chromosomal pattern number of CCM was 100. Transfection experiment demonstrated that CCM might be utilized to express foreign genes. Furthermore, cytotoxicity testing showed that CCM was susceptible to Aeromonas hydrophila, Aeromonas salmonicida, Aeromonas veronii, and Staphylococcus Aureus. The organophosphate pesticides (chlorpyrifos and glyphosate) or heavy metals (Hg, Cd, and Cu) exhibited dose-dependent cytotoxicity against CCM. After LPS treatment, the MyD88-IRAKs-NFκB pathway stimulates inflammatory-related factor il1ß, il8, il10, and nfκb expression. LPS did not seem to cause oxidative stress in CCM, and the expression of cat and sod was not affected. Poly (I:C) through TLR3-TRIF-MyD88-TRAF6-NFκB and TRIF-TRAF3-TBK1-IRF3 activated the transcription of related factors, increased expression of anti-viral protein, but no changes in apoptosis-related genes. To our knowledge, this is the first muscle cell line in Yellow River carp and the first study on the immune response signal pathways of Yellow River carp based on the muscle cell line. CCM cell line provides a more rapid and efficient experimental material for fish immunology research, and this study preliminarily elucidated its immune response strategy to LPS and poly (I:C).

Dietary Bile Acid Supplementation Could Regulate the Glucose, Lipid Metabolism, and Microbiota of Common Carp (Cyprinus carpio L.) Fed with a High-Lipid Diet.

This study sought to examine the role of bile acids in the regulation of glucose and lipid metabolism, intestinal flora, and growth in high-fat diet-fed common carp (Cyprinus carpio L.). Fish (6.34 ± 0.07 g) were fed for 56 days with three different diets, the control diet (CO, 5.4% lipid), high-fat diet (HF, 11% lipid), and high-fat diet with 60 mg/kg bile acids (BAs, 11% lipid). The results showed that high-fat diets resulted in poor growth performance and increased triglyceride (TG) in serum and the liver. The addition of bile acids significantly alleviated the adverse effects of a high-fat diet. The mRNA expression results indicated that bile acids may improve lipid metabolism through the enhancement of the peroxisome proliferator-activated receptor (PPARa). The expression of gluconeogenesis-related phosphoenolpyruvate carboxykinase (PEPCK) mRNA was inhibited, while fibroblast growth factor 19 (FGF19) was significantly higher. Bile acids reshaped the intestinal microflora community, with the level of Bacteroidetes increasing. The correlation analysis indicated that Patescibacteria, Dependentiae, Myxococcota, and Planctomycetota in the gut are associated with genes involved in glucose and lipid metabolism. These results indicated that bile acids could ameliorate the negative effects of high-fat diets on common carp.

Effects of Genistein on Lipid Metabolism, Antioxidant Activity, and Immunity of Common Carp (Cyprinus carpio L.) Fed with High-Carbohydrate and High-Fat Diets.

A 56-day feeding trial was conducted to investigate the effects of genistein on growth, lipid metabolism, antioxidant capacity, and immunity of common carp fed with high-carbohydrate or high-fat diets. Five diets were used to feed fish: control diet (5% fat; CO), high-fat diet (11% fat; HF), high-carbohydrate diet (45% carbohydrate; HC), and HF or HC diet with 500 mg/kg genistein (FG or CG). Results showed that final body weight (FW) and specific growth rate (SGR) were significantly reduced, but the supplementation with genistein resulted in higher values of FW and SGR than the HF or HC group. Both high carbohydrate and high fat belong to high-energy diets, which may promote lipid deposition. Genistein obviously decreased liver triglyceride (TG) content and alleviated hepatic fat vacuolation in the HF and HC groups. The expression of lipid metabolism genes (cpt-1 and atgl) was markedly higher in the FG group than in the HF group. The lipid synthesis-related genes (fas, acc, and pparγ) were elevated in high-energy diets but recovered to the control level or reduced after genistein treatments. With respect to fatty acid transporter genes, fatp increased in the FG group, and cd36 increased in the CG group. Furthermore, the antioxidant and immune indexes, such as total antioxidant capacity (T-AOC), glutathione peroxidase (GSH-PX), superoxide dismutase (SOD), acid phosphatase (ACP), and lysozyme (LZM) activities, were decreased, while malonate aldehyde (MDA) content, activities of alanine aminotransferase (ALT), and aspartate aminotransferase (AST) were enhanced in the HF and HC groups. The antioxidant and immunity values could be ameliorated by treatment with genistein. Moreover, the transcript levels of antioxidant-related genes (cat, gr, and nrf2) in the liver and anti-inflammatory factors (tgf-ß and il-10) and lyz in the head kidney tissue were promoted, although the expression levels of proinflammatory factors (tnf-α and il-6) declined in the genistein supplementation group, which confirmed the antioxidant and immune-enhancing effects of genistein. Therefore, 500 mg/kg genistein could ameliorate the negative effects of high-energy diets on immunity.

Genome-wide characterization of the Triplophysa dalaica slc4 gene family and expression profiles in response to salinity changes.

BACKGROUND: The solute carrier 4 (SLC4) gene family is involved in a variety of physiological processes in organisms and is essential for maintaining acid-base balance in the body. The slc4 genes have been extensively studied in mammals, and they play important roles in intracellular and extracellular pH regulation and in the secretion and uptake of HCO3- and other ions (Na+ and Cl-) between transepithelial cells in different tissues. This study identified and characterized the entire slc4 gene family of Triplophysa dalaica. RESULTS: Fifteen slc4 genes were identified in the whole genome of Triplophysa dalaica in this study, including five copies of Na+-independent Cl-/HCO3- transporters, eight members of Na+-dependent HCO3- transporters, and two genes coding Na+-coupled borate transporters. The chromosomal location information, isoelectric points, and molecular weights of the 15 slc4 genes were analyzed. The results for gene structure, domain analysis, and phylogenetic relationships of this gene family showed that the slc4 genes (except for slc4a9, which is missing in teleosts) are significantly expanded in teleosts compared to higher vertebrates. This phenomenon suggests that the slc4 gene family played an important role in the transition from aquatic to terrestrial animals. RT-PCR results showed that different slc4 genes showed diversified expression patterns in the tissues of T. dalaica. For osmotic pressure regulating organs, slc4a1b, slc4a4b, slc4a7, and slc4a11a were highly expressed in gills. In the kidney, slc4a1a, slc4a3, and slc4a10b were highly expressed, suggesting that the slc4 genes play a specific role in the salinity adaptation of T. dalaica. Our study has deciphered the biological roles of the slc4 genes in maintaining ionic and acid-base homeostasis in teleost fishes and provides a foundation for future exploration of the highly differentiated gene family in Triplophysa. CONCLUSIONS: The results are relevant for the breeding of alkali-tolerant varieties in saline-alkali areas for aquaculture. Our findings have important implications for the adaptation process of freshwater species to saline-alkali water.

Agromyces agglutinans sp. nov., isolated from saline lake sediment.

A novel actinobacterium, designated strain CFH 90414T, was isolated from sediment sampled at a saline lake in Yuncheng, Shanxi, PR China. The taxonomic position of the strain was investigated by using a polyphasic approach. Cells of strain CFH 90414T were Gram-reaction-positive, aerobic and non-motile. Growth occured at 4-40 °C, pH 5.0-9.0 and in the presence of up to 0-3.0 % (w/v) NaCl. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain CFH 90414T was a member of the genus Agromyces. The 16S rRNA gene sequence similarity analysis indicated that strain CFH 90414T was most closely related to Agromyces italicus JCM 14320T (98.07 %) and Agromyces lapidis JCM 14321T (97.18 %). The whole genome of CFH 90414T was 3.64 Mb, and showed a G+C content of 71.5 mol%. The average nucleotide identity (ANI) values and digital DNA-DNA hybridization (dDDH) values between CFH 90414T and the other species of the genus Agromyces were found to be low (ANI <78.99 % and dDDH <22.9 %). The whole-cell sugars were rhamnose, mannose, ribose, glucose and galactose. The isolate contained l-2,4-diaminobutyric acid, d-alanine, d-glutamic acid and glycine in the cell-wall peptidoglycan. The predominant menaquinone was MK-12. The major cellular fatty acids were anteiso-C15 : 0, anteiso-C17 : 0 and iso-C16 : 0. The polar lipid profile contained diphosphatidylglycerol, phosphatidylglycerol and an unidentified glycolipid. On the basis of phenotypic, genotypic and phylogenetic data, strain CFH 90414T is considered to represent a novel species of the genus Agromyces, for which the name Agromyces agglutinans sp. nov. is proposed. The type strain is CFH 90414T (=DSM 105966T=KCTC 49062T).

Devosia ureilytica sp. nov., isolated from Kuche River in China.

Two novel strains, designated XJ19-45T and XJ19-1, were isolated from water of Kuche River in Xinjiang Uygur Autonomous Region, China. Their cells were Gram-stain-negative, aerobic and motile rods. The phylogenetic analyses based on 16S rRNA genes and genomes showed that the two isolates belonged to the genus Devosia and the closest relative was Devosia subaequoris HST3-14T. The 16S rRNA genes sequences pairwise similarities, average nucleotide identities, digital DNA-DNA hybridizations and average amino acid identities between type strain XJ19-45T and other relatives were all less than 98.3, 80.3, 23.6 and 85.7 %, respectively, all below the species delineation thresholds. Pan-genomic analysis indicated that the novel isolate XJ19-45T shared 1594 core gene clusters with the 11 closely related type strains in Devosia, and the number of strain-specific clusters was 390. The major cellular fatty acids (>10 %) of the two isolates were summed feature 8, C18 : 1 ω7c 11-methyl and C16 : 0. Diphosphatidylglycerol, phosphatidylglycerol and glycolipids were the major polar lipids, and Q10 was the detected respiratory quinone. Based on the results of phenotypic, physiological, chemotaxonomic and genotypic characterizations, we propose that the isolates represent a novel species, for which the name Devosia ureilytica sp. nov. is proposed. The type strain is XJ19-45T (=CGMCC 1.19388T=KCTC 92263T).

Fatty hepatocytes-derived exosomal miR-122 reduces immune function and antioxidant defence in Ctenopharyngodon idella kidney (CIK) cells.

Exosomes are important for intercellular "cross talk", but the role of exosomes in communication between hepatocytes and C. idella kidney (CIK) cells remains unknown. In this study, we detected the changes in factors related to immune and oxidative stress to investigate the molecular mechanism by which fatty hepatocyte-derived exosomes (OA-Exos) reduced immunity and induced oxidative stress in CIK cells. After incubation of CIK cells by OA-Exos for 24 h, tumor necrosis factor-α (TNF-α), nuclear factor-κB (NF-κB) and interleukin-1ß (IL-1ß) were significantly upregulated in the OA-Exos group (P < 0.05), and Mn superoxide dismutase (Mn-SOD) and heme oxygenase-1 (HO-1) were significantly downregulated (P < 0.05). Surprisingly, miR-122 expression was also significantly elevated after OA-Exos incubation. We further identified the expression of miR-122 and found that it was notably increased in OA-Exos compared to hepatocyte-derived exosomes (Exos). Then we transfected CIK cells with miR-122 mimic, consistently, the expression of inflammatory cytokines was also significantly elevated (P < 0.05), and the expression of glutathione peroxidase (GPx), HO-1, and Mn-SOD were dramatically decreased (P < 0.05). Furthermore, HO-1 was improved to be a direct target of miR-122, and transfection with HO-1 siRNA indicated that changes in inflammatory cytokines and genes related to oxidative stress were consistent with the above results of CIK cells incubated with OA-Exos and miR-122 mimic. We concluded that OA-Exos may, through the miR-122/HO-1 pathway, reduce immune function and antioxidant defence in CIK cells.

Brachybacterium subflavum sp. nov., a novel actinobacterium isolated from the foregut of grass carp.

A novel actinobacterium, designated CFH 10395T, was isolated from the foregut of grass carp (Ctenopharyngodon idella), which had been fed with ginseng extract supplement. The taxonomic position was investigated by a polyphasic approach. Cells of CFH 10395T were Gram-staining-positive, aerobic, ovoid-shaped, non-spore-forming and non-motile. On the basis of the results of 16S rRNA gene sequence analysis, CFH 10395T was most closely related to Brachybacterium endophyticum KCTC 49087T, Brachybacterium squillarum JCM 16464T and Brachybacterium paraconglomeratum JCM 17781T (97.85%, 97.51 and 97.29% similarity, respectively). CFH 10395T grew at 4-37 °C, pH 5.0-9.0 and in the presence of up to 10.0 % NaCl (w/v). The dominant menaquinone was MK-7. The whole-cell sugars were rhamnose, glucose, mannose and galactose. meso-diaminopimelic acid was the diagnostic diamino acid in the cell-wall peptidoglycan. The major fatty acids were anteiso-C15 : 0, anteiso-C17 : 0 and iso-C16 : 0. The genome size was 3.99 Mbp with a DNA G+C content of 71.9 mol%. On the basis of the results of phylogenetic analysis, physiological properties, chemotaxonomic characteristics, low average nucleotide identity (ANI) and digital DDH (dDDH) results [ANI calculated using MUMmer (ANIm) <87 %, ANI calculated using blast (ANIb) <83 % and dDDH <23 %], it is concluded that CFH 10395T represents a novel species of the genus Brachybacterium, for which the name Brachybacterium subflavum sp. nov., is proposed. The type strain is CFH 10395T (=CGMCC 1.13804T=KCTC 49235T).

Transcriptome analysis of common carp (Cyprinus carpio) provides insights into the ovarian maturation related genes and pathways in response to LHRH-A and dopamine inhibitors induction.

Luteinizing hormone-releasing hormone analog (LHRH-A) and dopamine inhibitors have been widely used to induce oocyte maturation and ovulation in domesticated fishes. Although this approach represents a reliable method for regulating fish reproduction, the underlying molecular mechanisms mediating LH action are largely unexplored. The objective of this study was to determine the transcriptional profile of gene programming in hormone-treated common carp. In the present study, female common carp were intraperitoneally injected with LHRH-A together with dopamine inhibitors, and control fish were injected with saline. Ovarian morphological changes were analysed by both light microscopy and scanning electron microscopy. Furthermore, gene expression profiling of the brain and ovarian tissues was performed by Illumina sequencing. Compared to the control carp, hormone treatment resulted in morphological changes including disappearance of nuclear membrane, breakdown of germinal vesicle (GVBD), and fusion of yolk globules, reflecting that hormones significantly promoted oocyte maturation. In comparison to control, we have identified 867 and 9,053 differentially expressed genes in the hormone-treated female brain and ovary, respectively. In the brain, most of the identified genes were significantly enriched in 18 KEGG pathways. In the ovarian tissue, the identified genes were significantly involved in 9 pathways. In the hormone-treated carp, genes were involved in calcium signalling pathway, cAMP signalling pathway, insulin secretion, and oxidative phosphorylation pathway, which showed obvious associations with ovarian maturation. The present study provides transcriptomic information for hormone-treated carp, which might be useful for studying the endocrine regulation and mechanisms of ovarian maturation in domesticated fishes.

Molecular identification of FNDC5 and effect of irisin on the glucose metabolism in common carp (Cyprinus carpio L.).

Irisin, encoded by fibronectin type III domain-containing protein 5 (FNDC5) gene, plays a role in energy expenditure and insulin sensitivity in mice. In fish, the function of irisin related to glucose metabolism is less reported. It may increase glucose utilization in fish. The aim of the present study was to characterize the regulatory role of irisin in glucose metabolism in common carp (Cyprinus carpio L.). In this study, FNDC5a and FNDC5b were isolated from common carp. The cDNA of FNDC5a and FNDC5b were 722 bp and 714 bp, encoding 221 and 207 amino acids, respectively. FNDC5a was abundantly expressed in the brain and gonad. FNDC5b was mainly expressed in brain. Different expression pattern of FNDC5a and FNDC5b under fasting/refeeding and OGTT experiment were identified. The recombinant common carp irisinA and irisinB were prepared by prokaryotic expression system. Glucose concentration was decreased in treatment with irisinA or irisinB in the in vitro and in vivo experiments. The mRNA expression levels of gluconeogenesis-related genes were significantly down-regulated, while the mRNA expression of glycolysis-related genes were significantly up-regulated after treatment with recombinant irisinA or irisinB in liver in vivo and in primary hepatocytes in vitro. Our research shows that irisin inhibits hepatic gluconeogenesis and promotes hepatic glycolysis. Taken together, this study for the first time revealed the two subtypes of FNDC5 and explored the function and mechanisms of irisinA and irisinB in fish glucose homeostasis.

Molecular identification of glucose transporter 4: The responsiveness to starvation, glucose, insulin and glucagon on glucose transporter 4 in common carp (Cyprinus carpio L.).

Glucose transporter 4 (GLUT4) is comprehensively investigated in mammals, while the comparative research of GLUT4 in common carp is deficient. To investigate the function of GLUT4, carp glut4 was first isolated. The open reading frame of carp glut4 was 1518 bp in length, encoding 505 amino acids. A high-sequence homology was identified in carp and teleost, and the phylogenetic tree displayed that the carp GLUT4 was clustered with the teleost. A high level of glut4 mRNA was analysed in fat, red muscle and white muscle. After fasting treatment, glut4 mRNA expression was increased significantly in muscle. In the oral glucose tolerance test experiment, glut4 mRNA was also significantly elevated in muscle, gut and fat. Furthermore, intraperitoneal injection of insulin resulted in the upregulation of glut4 gene expression significantly in white muscle, gut and fat. On the contrary, the glut4 mRNA level in the white muscle, gut and fat was markedly downregulated after glucagon injection. These results suggest that GLUT4 might play important roles in food intake and could be regulated by nutrient condition, insulin and glucagon in common carp. Our study is the first to report on GLUT4 in common carp. These data provide a basis for further study on fish GLUT4.