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Simple Summary: The application of recombinant technology provides the possibility for the establishment of ideal probiotics as an alternative novel therapeutic approach by integrating promising heterologous proteins into the chromosome of conventional probiotics. In our previous study, we found that Nile tilapia CC chemokine genes had multiple roles in homeostatic functions in both immune and non-immune tissues. Moreover, they are crucially involved in the early immune responses to pathogens. Therefore, in this study, we isolated and characterized the potential probiotic Bacillus subtilis from the intestine of Nile tilapia to use as a live vehicle for delivering CC chemokine proteins directly to the intestine. Our results suggest that dietary supplementation with either wild-type or recombinant probiotics could enhance the immune responses of Nile tilapia, with stronger stimulation observed in the recombinant probiotics-supplemented group. It could be concluded that applying genetically engineered probiotics for developing novel strategies would provide greater health benefits through a combination efficacy of potential probiotics and desirable heterologous proteins. This study aimed to investigate the potential probiotic Bacillus spp. from the intestine of Nile tilapia in order to construct a recombinant probiotic for the enhancement of the Nile tilapia immune response. One hundred bacterial isolates from the intestine of Nile tilapia were characterized for species identification using the 16s ribosomal RNA (rRNA). Only Bacillus isolates with exhibited antagonistic activity were investigated for their biological functions, which included protease-producing capacity, bile salts and pH tolerance, antibiotic susceptibility, and pathogenicity tests. According to the best results, Bacillus isolate B29, as closely related to B. subtilis, was selected to construct a recombinant probiotic for the delivery of CC chemokine protein (pBESOn-CC). The existence of recombinant probiotics was confirmed by Western blotting before the feeding trial. In addition, the CC chemokine mRNA level was quantified in the intestine of fish fed probiotics after 30 days of feeding. Total immunoglobulin, lysozyme activity, alternative complement 50 activity (ACH50), and phagocytic activity of fish fed either wild-type or recombinant probiotics were significantly increased, indicating that probiotics could stimulate the Nile tilapia immune system through different processes. Interestingly, the dietary supplementation of recombinant probiotics has a stronger immune response enhancement than the wild-type strain. [ABSTRACT FROM AUTHOR]

The effects of various sous-vide (SV) cooking conditions (50-60℃, 30-60 min) on physicochemical properties related to the texture characteristics, protein structure/degradation, and sensory acceptability of tilapia fillet (Oreochromis niloticus) were investigated. With an increasing temperature and processing time of SV cooking, protein degradation (of both myofibrils and connective tissue) was more pronounced, as evaluated by the decrease in water- and salt-soluble proteins, total collagen, as well as the changes in the ratio of secondary protein structures (α-helix, β-sheet, β-turn, etc.), which were determined by synchrotron-FTIR (SR-FTIR). These degradations were associated with the improvement of meat tenderness, as estimated by shear force and texture profile analyzer (TPA) results. Among all SV conditions, using 60 ℃ for 45 min seems to be the optimal condition for tilapia meat, since it delivered the best results for texture characteristics and acceptability (p < 0.05). Moreover, principal component analysis (PCA) results clearly demonstrated that the highest texture-liking score of this condition was well associated with the intensity of β-sheets, which seem to be the crucial component that affected the texture of SV-cooked tilapia more so than other parameters. The findings demonstrated the potential of SR-FTIR to decipher the biomolecular structure, particularly the secondary protein structure, of SV-cooked tilapia. This technique provided essential information for a better understanding of the changes in biomolecules related to the textural characteristics of this product. [ABSTRACT FROM AUTHOR]

Simple Summary: Gene markers that are specific to germ cells are needed for the development of reproductive biotechnologies for fish. To provide valuable molecular information on ddx4 and dnd, which can be applied for further monosex production in snakeskin gourami (Trichopodus pectoralis), this study cloned and characterized ddx4 (ddx4) and dnd (dnd1) homologs and described their temporal expressions in snakeskin gourami (Trichopodus pectoralis). The expressions of ddx4 and dnd1 mRNAs were detectable only in the gonads, particularly in germ cells. The expression of ddx4 was high during early larval development and decreased with increasing developmental age, whereas the expression of dnd1 increased with developmental age. In adult fish, the expression levels of ddx4 and dnd1 were higher in the ovary than in the testis. The purpose of this study was to clone and characterize ddx4 and dnd1 homologs in snakeskin gourami (Trichopodus pectoralis) and to determine their expression levels during larval development and in the gonads of males and females. Both cDNAs contained predicted regions that shared consensus motifs with the ddx4 family in teleosts and the dnd family in vertebrates. Phylogenetic tree construction analysis confirmed that these two genes were clustered in the families of teleosts. Both ddx4 and dnd1 mRNAs were detectable only in the gonads, particularly in germ cells. These two genes were expressed during early larval development. The expression of ddx4 was high during early larval development and decreased with increasing developmental age, whereas dnd1 expression increased with developmental age. In adult fish, the expression levels of both genes were higher in the ovary than in the testis. Overall, these findings provide valuable molecular information on ddx4 and dnd, and can be applied in future reproductive biological studies relating to sex dimorphism in snakeskin gourami. [ABSTRACT FROM AUTHOR]

This study aims to shed light on the association between non-volatile and volatile compounds related to flavor/taste characteristics as well as sensory acceptability of Nile tilapia fillet (Oreochromis niloticus) cooked by various sous-vide (SV) conditions (50–60 ℃, 30–60 min), with fish cooked with boiling water used as control. Higher temperatures and longer processing times of SV cooking led to greater protein and lipid oxidation as indicated by the increase in total sulfhydryl (-SH), carbonyl, free fatty acid (FFA) contents as well as peroxide values (PV) and thiobarbituric acid reactive substance (TBARS) values. The differences in flavor/taste components including adenosine triphosphate (ATP)-related compounds, free amino acids (FAAs) and volatiles were also obtained, which directly affect sensory acceptability as evaluated by using the hedonic scale. Based on principal component analysis (PCA) results, the acceptability score was strongly correlated with inosine monophosphate (IMP) and acetoin, which seem to be the most crucial flavor enhancers for cooked tilapia. Among all samples, tilapia processed at 60 °C for 45 and 60 min, which contained significantly higher IMP and acetoin (p < 0.05) than others, had significantly higher flavor-liking and overall-liking scores, with a more than 7.5 meaning for high acceptability (p < 0.05), indicating the optimal SV conditions for tilapia fillet. Overall, the present finding indicated that the SV-cooking technique, at the optimal conditions, can improve the meat quality of cooked fish, in terms of flavor/taste characteristics, compared with traditional cooking (control). [ABSTRACT FROM AUTHOR]

Simple Summary: Factors such as changes in water quality, the transport and handling of the fish, the presence of pollutants, and the high densities used in intensive farming of red tilapia, among others, can cause oxidative stress in fish, which can lead to immune system suppression and increase the risk of opportunistic infections. The use of Jerusalem artichoke (JA) as an oligofructose-rich prebiotic dietary supplement in fish feed could enhance growth performance, hematological and immunological parameters, as well as disease resistance. However, the effect of JA in diets for red tilapia is poorly reported. In the present study, we evaluated the effects of a Jerusalem artichoke-supplemented diet on the blood chemistry, growth performance, intestinal morphology, expression of antioxidant-related genes, and disease resistance against Aeromonas veronii challenge in juvenile red tilapia. The results of JA-supplemented (JA5 and JA10) diets showed beneficial effects in terms of growth performance, blood chemistry, and intestinal morphology. Importantly, the prebiotic diets (JA5 and JA10) were associated with a significant increase in the expression of gpx1 and gst antioxidant-related genes and disease resistance against A.veronii. In conclusion, prebiotic diets have the potential to be employed as prospective supplements in sustainable red tilapia farming. The aim of the present study was to evaluate the effects of a Jerusalem artichoke-supplemented diet on the blood chemistry, growth performance, intestinal morphology, expression of antioxidant-related genes, and disease resistance against Aeromonas veronii challenge in juvenile red tilapia. A completely randomized design (CRD) was followed to feed red tilapias with three experimental diets: control, 5.0 g/kg JA-supplemented (JA5), or 10.0 g/kg JA-supplemented (JA10) diets in triplicates for 4 weeks. The results revealed that the growth performance, weight gain (WG), specific growth rate (SGR), and average daily gain (ADG) of fish fed diets JA5 and JA10 were significantly higher (p < 0.05) than those of fish fed the control diet. Fish fed the control diet had significantly higher T-bilirubin, D-bilirubin, and ALT in blood serum than fish fed JA5 and JA10, as well as higher BUN than fish fed JA5. The number of goblet cells in the proximal and distal parts of the intestine revealed that the number of acid, neutral, and double-staining mucous cells of fish fed diets JA5 and JA10 was significantly higher (p < 0.05) than in fish fed the control diet. The diets including the prebiotic (JA5 and JA10) were associated with a significant increase in the expression of gpx1 and gst antioxidant-related genes and disease resistance against A. veronii in juvenile red tilapia. Therefore, JA5 and JA10 can be employed as promising prebiotics for sustainable red tilapia farming. [ABSTRACT FROM AUTHOR]

The aim of this study was to explore for the first time in omnivorous fish the concept of nutritional programming. A nutritional stimulus was accomplished by microinjecting 2 M glucose into yolk reserves during the alevin stage in Nile tilapia (Oreochromis niloticus). At the molecular level in fry, at 1 week post-injection, glucose stimuli were associated with the up-regulation of genes involved in glycolysis (pklr , hk1 , hk2 , and pkma), glucose transport (glut4) pathways and down-regulation of genes related to gluconeogenesis (g6pca1 , g6pca2 , and pck1) and amino acid catabolism (asat , alat) (P < 0.05), demonstrating that the larvae well received the glucose stimulus at a molecular level. Moreover, 20 weeks after glucose injection, early glucose stimuli were always linked to permanent effects in juvenile fish, as reflected by a higher level of glycolytic enzymes [ gck , hk1 and hk2 at both mRNA and enzymatic levels and pyruvate kinase (PK) activity]. Finally, the effects of the glucose stimulus history were also examined in fish fed with two different dietary carbohydrate/protein levels (medium-carbohydrate diet, CHO-M; high-carbohydrate diet, CHO-H) in juvenile fish (during weeks 20–24). As expected, the CHO-H diet induced the expression of glycolytic and lipogenic genes (gck , pklr , hk1 , hk2 , fpkma, fasn , and g6pd) and suppressed the expression of gluconeogenic and amino acid catabolism genes (g6pca1 , pck1 , pck2 , asat , alat , and gdh). Nevertheless, the early glucose stimulus led to persistent up-regulation of glycolytic enzymes (gck , pklr , hk1 , and hk2) at both the mRNA and enzyme activity levels and glucose transporter glut4 as well as lower gluconeogenic pck1 gene expression (P < 0.05). More interestingly, the early glucose stimulus was associated with a better growth performance of juvenile fish irrespective of the diets. These permanent changes were associated with DNA hypomethylation in the liver and muscles, suggesting the existence of epigenetic mechanisms at the origin of programming. In conclusion, for the first time in tilapia, early glucose stimuli were found to be clearly associated with a positive metabolic programming effect later in life, improving the growth performance of the fish. [ABSTRACT FROM AUTHOR]

The rapid and intensive growth of aquaculture over the last decade, poses a tremendous challenge to this industry in order to comply with the latest guidelines, established to minimise its negative effects on the environment, animal welfare and public health. Farmed fish welfare has become one of the main priorities towards sustainable aquaculture production with several initiatives launched by the European Union within the framework of the 2030 agenda. It is clear that an unbiased and reliable way to access farmed fish welfare needs to be implemented due to the lack of reliable indicators and standardised methods that are used at present. In this review, we start by addressing the status quo of animal and fish welfare definition in particular, describing the methods and assays currently used to measure it. We then explain why we believe these methods are unreliable and why there is a need to establish new ones that will promote productivity and consumer's acceptance of farmed fish. The establishment of a new type of welfare biomarkers using cutting‐edge technologies like proteomics and other omics technologies is proposed as a solution to this issue. Therefore, we provide a brief description of these new methodologies, describing for each one how they can improve our scientific knowledge and the role they can play in farmed fish welfare biomarker discovery. [ABSTRACT FROM AUTHOR]

In general, promoters have significant influence on recombinant protein production. Herein, we compared the performance of actin (pACT), phosphoglycerate kinase (pPGK), and translational elongation factor (pTEF) promoters for driving the expression of fatty acid delta-6 (Δ6) desaturase from Nile tilapia (Oreochromis niloticus; Oni-fads2) in Saccharomyces cerevisiae. Our results showed that by applying real-time RT-PCR, the highest level of Oni-fads2 mRNA was observed in S. cerevisiae carrying the expression vector driven by pTEF promoters. Exogenous substrate C18:2n-6 was used to determine Δ6 activity by quantitatively determining the C18:3n-6 product. The results showed that highest Δ6 desaturation was observed when using pTEF as a promoter. Recombinant S. cerevisiae cells expressing Oni-fads2 driven by pTEF were tested with the substrate C18:3n-3, and Δ6 desaturation efficiently converted C18:3n-3 to C18:4n-3. Furthermore, crude extract of recombinant yeast also exhibited Δ6 activity. Thus, recombinant S. cerevisiae cells expressing Oni-fads2 driven by the pTEF promoter have potential as a yeast factory for the sustainable production of long-chain polyunsaturated fatty acids. [ABSTRACT FROM AUTHOR]

This study investigated the effects of dietary inulin or Jerusalem artichoke (JA) on the growth performance, haematological, blood chemical and immune parameters of Nile tilapia fingerlings. Five treatment diets were designed to incorporate inulin at 0 (basal diet), 2.5 and 5.0 g/kg and JA at 5.0 and 10.0 g/kg. Two basal diets including fish meal and formulated experimental feed were used for fry and fingerling growing periods, respectively. During the fry growing period, larvae were fed treatment diets for 4 weeks. There were no significant differences in growth performance or survival rate. Fingerlings were then nursed with the formulated experimental diets from weeks 5 to 12. Fingerlings fed on inulin at 5.0 g/kg or JA at either level had better growth performance and survival rate than that fed on the basal diets. There were no significant differences in body composition. Dietary prebiotic inulin and JA increased red blood cell number ( p < .05). Among the five blood chemistry parameters examined, both inulin at 5.0 g kg and JA (5.0 and 10.0 g/kg) increased blood protein ( p < .05). Dietary inulin at 5.0 g/kg and JA at 5.0 and 10.0 g/kg increased total immunoglobulin and lysozyme activity ( p < .05). Both inulin and JA inclusion diets increased alternative complement activity ( p < .05). Taken together, dietary inulin at 5 g/kg and JA at 5.0 and 10.0 g/kg had beneficial effects on the growth performance, survival rate and immune of Nile tilapia fingerlings. [ABSTRACT FROM AUTHOR]

This present study investigated the potential of Morinda citrifolia leaf aqueous extract to induce osteogenic differentiation and matrix mineralization of human periodontal ligament (hPDL) cells. Human periodontal ligament cells were cultured in complete medium, ascorbic acid with β-glycerophosphate, or Morinda citrifolia leaf aqueous extract. Morinda citrifolia leaf aqueous extract significantly increased alkaline phosphatase activity compared to culturing in complete medium or ascorhic acid with β-glycerophosphate. Matrix-containing mineralized nodules were formed only when the cells were cultured in the presence of Morinda citrifolia leaf aqueous extract. These nodules showed positive alizarin red S staining and were rich in calcium and phosphorus according to energy dispersive X-ray analysis. In conclusion, Morinda citrifolia leaf extract promoted osteogenic differentiation and matrix mineralization in human periodontal ligament cells, a clear indication of the therapeutic potential of Morinda citrifolia leaves in bone and periodontal tissue regeneration. [ABSTRACT FROM AUTHOR]

Abstract: In this study, we characterized the neuropeptide Y (NPY) mRNA in snakeskin gourami (Trichogaster pectoralis) (TpNPY). TpNPY displayed characteristics typical of previously reported NPYs, and it exhibited a high degree of homology with the NPY proteins of other vertebrates. A phylogenetic analysis demonstrated that TpNPY was closely related to the NPYs found in the acanthomorpha and salmoniformes fish species. TpNPY was found to be ubiquitously expressed in all brain regions when assessed by real-time RT-PCR and in situ hybridization. In addition, a graded expression level of TpNPY was observed in peripheral tissues; for example, a moderate level of TpNPY was found in the gills, liver, kidney, stomach, intestine, spleen and gonads, while a low level of TpNPY was found in the muscle. The change in expression of TpNPY with respect to daily feeding habits was investigated in distinct brain regions, including the telencephalon, mesencephalon, metencephalon, and diencephalon. Fluctuations in the expression level of TpNPY were observed for a 24h post-prandial period. Except for the telencephalon, a reduction in TpNPY expression was found after a meal, while a peak level of TpNPY was observed 1h before the scheduled breakfast. Furthermore, there was a positive correlation between TpNPY and TpMC4R in the telencephalon and diencephalon throughout the circadian feeding cycle, which suggests that there is a connection between the function of NPY and the melanocortin system for the regulation of daily feeding. Fish brains were incubated with an MC4R antagonist (i.e., HS024), and the expression of TpNPY and TpMC4R was measured. Interestingly, there was a significant relationship between the expression of TpNPY and TpMC4R under the effects of HS024, which demonstrates that there are interactions between MC4R and NPY, particularly in a hyperphagic state. [Copyright &y& Elsevier]

This present study investigated the potential of Morinda citrifolia leaf aqueous extract to induce osteogenic differentiation and matrix mineralization of human periodontal ligament (hPDL) cells. Human periodontal ligament cells were cultured in complete medium, ascorbic acid with p-glycerophosphate, or Morinda citrifolia leaf aqueous extract. Morinda citrifolia leaf aqueous extract significantly increased alkaline phosphatase activity compared to culturing in complete medium or ascorbic acid with β-glycerophosphate. Matrix-containing mineralized nodules were formed only when the cells were cultured in the presence of Morinda citrifolia leaf aqueous extract. These nodules showed positive alizarin red S staining and were rich in calcium and phosphorus according to energy dispersive X-ray analysis. In conclusion, Morinda citrifolia leaf extract promoted osteogenic differentiation and matrix mineralization in human periodontal ligament cells, a clear indication of the therapeutic potential of Morinda citrifolia leaves in bone and periodontal tissue regeneration. [ABSTRACT FROM AUTHOR]

This study was conducted to evaluate the effect of dietary levels of Brewtech dried brewers yeast (BY) and GroBiotic-A (GB) on growth performance, proximate body composition, immune response, and resistance of juvenile Nile tilapia to Streptococcus iniae challenge. A practical basal (control) diet formulated to contain approximately 32% crude protein and 6% lipid was supplemented with 1% and 2% of BY or GB. Each diet was fed to Nile tilapia in quadruplicate aquaria for 12 weeks. Weight gain, feed intake, survival, and whole body proximate composition of fish were not significantly affected by dietary treatments. Serum total protein, total immunoglobulin, lysozyme, and agglutinating antibody titer to S. iniae were not significantly affected by dietary supplementation of BY or GB. However, serum haemolytic complement activity (SH50) of fish fed 1% BY was significantly higher than those of fish fed the control diet and diets supplemented with 2% BY or GB. Cumulative mortality of fish 20 days post-challenge with S. iniae was unaffected by dietary treatments. However, fish fed diets supplemented with 1% BY and 2% BY or GB had substantially reduced and earlier cessation of mortality. [ABSTRACT FROM PUBLISHER]

Abstract: The influence of 17α-methyltestosterone (MT) on growth responses, biological parameters and the expression of genes involved in the GH–IGF pathway of the hypothalamic–pituitary–liver–gonadal axis were investigated in female, male, and sex-reversed Nile tilapia to evaluate the relationship between sex and MT-induced changes in these parameters. Female fish had a lower growth rate than male and sex-reversed fish, and MT increased growth performance and duodenal villi in females. Most but not all biological parameters of sex-reversed fish were similar to those of male fish. Male fish had higher red blood cell counts and hemoglobin levels than female and sex-reversed fish, suggesting that these hematological indices reflect a higher metabolic rate in male fish. Greater blood triglyceride levels indicated the vitellogenin process in female fish. MT increased the alternative complement activity in female fish (P <0.05). Sex and MT had no significant effects on the hypothalamic mRNAs of GHRH and PACAP. Although not statistically significant, females tended to have higher GH mRNA levels than male and sex-reversed fish. Additionally, MT tended to decrease and increase GH mRNA levels in female and male fish, respectively. There were significant differences among sexes in the expression of GHR, and IGF mRNAs at the peripheral level in the liver and gonads. Females had lower hepatic GHRs and higher ovarian GHRs than male and sex-reversed fish. While the mRNA levels of IGF-1 were lower in the ovary, the levels of IGF-2 were higher compared with those in testes. A significant correlation between GHRs and IGFs was demonstrated in the liver and gonad (except for IGF-1). Multiple regression analysis showed a significant relationship between GH mRNA and both GHRs and IGFs in the liver and gonad. MT exerted androgenic and, to some extent, estrogenic effects on several physiological parameters and GH–IGF action. [Copyright &y& Elsevier]

Abstract: In this study, melanocortin 4 receptor (MC4R) was characterized in Snakeskin Gourami (Trichogaster pectoralis) and designated as TpMC4R. The TpMC4R contained predicted regions that were structural features of MC4R subtypes of vertebrates. In addition, phylogenetic analyses showed that TpMC4R were in MC4R orthologues and closely related to piscine MC4Rs. The TpMC4R transcripts were detected in embryos at all developmental stages. Further, its mRNA was detectable in unfertilized eggs, suggesting that TpMC4R could be classified as maternal mRNA. Using real-time RT-PCR, graded expression at each central and peripheral tissue was demonstrated. Localization of TpMC4R expression in the brain was performed by in situ hybridization. An experiment was conducted to determine the expression profile of TpMC4R during daily feeding in each region of the brain. Fish were fed twice a day at 10:00 and 16:00, and the level of TpMC4R expression was analyzed at 9:00, 11:00, 13:00 and 15:00. The expression level of TpMC4R was significantly reduced at 15:00 compared to that at all previous times (9:00, 11:00 and 13:00). Moreover, significantly higher expression levels of TpMC4R were observed at 18- and 24-h fasting periods. The finding of variation in MC4R expression level during daily feeding and fasting period revealed some direct and/or indirect relations of the MC4R expression to circadian feeding rhythm and short-term food deprivation in fish. [Copyright &y& Elsevier]

So far, there are a number of fish genome projects, including experimental and economically important fish that provide available DNA sequence information. However, the function of a gene cannot be deduced only by its DNA sequence. Therefore, a technique with which to investigate the function of the fish gene is needed. Gene knockdown (GKD), or antisense technology, is now being used as a powerful technique to study gene functions in living organisms. GKD effects result from the introduction of an antisense molecule into living cells. The antisense agents bind to target messenger RNA, thus inactivating the target gene expression. The appropriately spatial inhibitory effects on protein production from corresponding gene resulted in the phenotypic change. Therefore, the function of the gene can be understood. To date, there are a number of antisense molecules that can affect efficient GKD in fish. These include antisense oligonucleotides, small interfering RNA, and ribozyme. These antisense molecules cause specific gene inhibitor effects with different mechanisms. The various antisense mechanism types facilitate a number of GKD applications with various approaches in animals. In this review, we demonstrate the characteristics of each antisense molecule, its mechanism, and its application, especially for gene functional analysis in fish. [ABSTRACT FROM AUTHOR]

Abstract: We report an effective gene knockdown technique in rainbow trout embryos using additional RNA components combined with ribozymes (Rzs). Chimeric Rzs (tRzCs) containing tRNAVal, Rz against GFP, and a constitutive transport element were microinjected into transgenic embryos. tRzCs induced greater gene interference than Rzs alone. Control tRzCs did not affect unpaired bases of target RNA, and the tRzC did not interfere with non-relevant gene expression, suggesting that the tRzC-mediated gene-interference effects were sequence-specific. Furthermore, the tRzC-containing expression vector specifically suppressed target GFP expression in transgenic trout. tRzCs enhance Rz cleavage and could therefore be powerful tools for studying unknown gene function in vertebrates. [Copyright &y& Elsevier]

Recently, small interfering RNAs (siRNAs) have been used for gene knockdown in mammalian cultured cells, but their utility in fish has remained unexplored. Here we demonstrate a siRNA-mediated gene silencing technique in rainbow trout embryos. We found that siRNAs effectively suppressed the transient expression of episomally located foreign GFP genes at an early developmental stage and inhibited the expression of GFP genes in stable transgenic trout embryos. Similar gene silencing was observed with an siRNA against the endogenous tyrosinase A gene. siRNAs interfered with the expression of maternally inherited mRNA. siRNAs did not affect non-relevant gene expression and siRNAs with a 4 base mismatch did not affect target gene expression. siRNA gene silencing is therefore highly sequence-specific. Our findings are the first evidence that siRNA-mediated gene silencing is effective in fish. This technique could be a powerful tool for studying gene function during embryonic development in aquacultural fish species, zebrafish, and medaka. [Copyright &y& Elsevier]

Simple Summary: This study presents the effects of different dietary vegetable lipid sources (DLs) on several blood parameters, which may indicate the health status of Nile tilapia (Oreochromis niloticus). The tested DLs had no effects on haematological indices. However, the immune response analysis showed that DLs promoted immune systems via different processes. Dietary polyunsaturated fatty acids (PUFAs) increased expression of several proteins involved in coagulation activity, while saturated fatty acids (SFAs) might influence heme lipid-oxidation. Overall, different DLs had effects on several parameters corresponding to health status in Nile tilapia, and dietary PUFAs appear to promote health in this fish. This study aimed to investigate the effects of DLs, including palm oil (PO; an SFAs), linseed oil (LO; n-3 PUFAs) and soybean oil (SBO; n-6 PUFAs) on the health status of Nile tilapia (Oreochromis niloticus) during adulthood. Three experimental diets incorporating PO, LO or SBO were fed to adult Nile tilapia for a period of 90 days, and haematological and innate immune parameters were evaluated. Proteome analysis was also conducted to evaluate the effects of DLs on plasma proteins. The tested DLs had no significant effects on red blood cell (RBC) count, haematocrit, haemoglobin, and total immunoglobulin and lysozyme activity. Dietary LO led to increased alternative complement 50 activity (ACH50), and proteome analysis revealed that PO and SBO enhanced A2ML, suggesting that different DLs promote immune system via different processes. Dietary LO or SBO increased the expression of several proteins involved in coagulation activity such as KNG1, HRG and FGG. Increased HPX in fish fed with PO suggests that SFAs are utilised in heme lipid-oxidation. Overall, DLs with distinct fatty acids (FAs) affect several parameters corresponding to health status in Nile tilapia, and dietary LO and SBO seemed to strengthen health in this species. [ABSTRACT FROM AUTHOR]

The Vago interferon-like protein participates in the interplay between interferon regulatory factors and the expression of immune-responsive genes. Vago was initially perceived to participate only in the antiviral activation through JAK/STAT pathway. However, certain isoforms of Vago can stimulate antimicrobial responses. Here we identify Vago isoforms in Fenneropenaeus merguiensis (Fm Vagos) and how they function in antiviral and antibacterial responses against highly invasive pathogens, including white spot syndrome virus (WSSV) and Vibrio parahaemolyticus (VP AHPND). Three isoforms of Fm Vagos were identified: Fm Vago4, Fm Vago5a, and Fm Vago5b, and expressed throughout tissues of the shrimp. During infection, Fm Vago4, Fm Vago5a, and Fm Vago5b, were up-regulated after WSSV and VP AHPND challenges at certain time points. Pre-injection of purified recombinant Fm Vago4 (rVago4), Fm Vago5a (rVago5a), and Fm Vago5b (rVago5b) proteins could significantly reduce the mortality of shrimp upon WSSV infection, while the increase of survival rate of VP AHPND -infected shrimp was observed only in rVago4 treatment. The immunity routes that Fm Vagos might instigate in response to the pathogens were examined by qRT-PCR, revealing that the JAK/STAT pathway was activated after introducing rVago4, rVago5a, and rVago5b, while the Toll/IMD pathway and proPO system, combined with PO activity, were provoked only in the rVago4-treated shrimp. Our finding suggests cross-talk between Vago's antiviral and antimicrobial responses in shrimp immunity. These findings complement previous studies in which Vago and its specific isoform could promote viral and bacterial clearance in shrimp. • We identify three isoforms of Vago in Fenneropenaeus merguiensis. • By qRT-PCR, Fm Vagos expression is up-regulated upon challenges. • rVago4, rVago5a, and rVago5b could mitigate mortality upon WSSV infection. • rVago4 enhanced the survival rate against both WSSV and VP AHPND challenges. • The JAK/STAT and Toll/IMD pathways and phenoloxidase system were activated by rVago4. [ABSTRACT FROM AUTHOR]

Abstract Synbiotics, a synergistic combination of probiotics and prebiotics, are currently regarded as one of the most practical nutritional supplements in tilapia farms. In this study, the effect of supplementing the diet of red tilapia (Oreochromis spp.) with Jerusalem artichoke (Helianthus tuberosus) and Lactobacillus rhamnosus GG (LGG) was evaluated. Growth performance, serum biochemical parameters, intestinal morphology, goblet cell counts, immune parameters and protection against Aeromonas veronii challenge were determined. The results showed that fish fed with synbiotic-supplemented diets had a significantly higher (P < 0.05) feed conversion ratio (FCR), specific growth rate (SGR), and average daily gain (ADG) than fish fed with a control diet. The synbiotic-supplemented diet increased glucose, total protein and the total cholesterol levels. The absorptive area of the proximal and distal intestine of fish fed on the synbiotic diet was significantly higher (P < 0.05) than in those fed with probiotics (LGG), prebiotic-supplemented diets (JA), and the control diet. Goblet cell counts revealed that the numbers of acid mucous cells, neutral mucous cells and double-staining mucous cells of fish fed the synbiotic-supplemented diet (JA + LGG) were significantly higher (P < 0.05) in the proximal and distal intestine. Fish fed the synbiotic-supplemented diets also exhibited significantly higher (P < 0.05) lysozyme activity. The cumulative mortalities of fish fed with a synbiotic-supplemented diet were significantly lower than those of fish fed other diets. The results suggested the beneficial effect of JA and LGG synbiotic diet on growth performance and health status of red tilapia. Direct administration of JA and LGG in fish feed can be used as a practical nutritional supplement in red tilapia. Highlights • JA and L. rhamnosus GG can used as natural synbiotic ingredient for red tilapia. • JA and L. rhamnosus GG improved the absorptive area of the intestine which correlate to the better growth performance in red tilapia. • JA and L. rhamnosus GG enhanced blood glucose, total protein and total cholesterol levels in red tilapia. • JA and L. rhamnosus GG enhanced gut mucosal immunity and lysozyme activity against A. veronii challenge in red tilapia. [ABSTRACT FROM AUTHOR]

Circular RNAs (circRNAs) are non-coding RNAs (ncRNAs) originating from a post-transcriptional modification process called back-splicing. Despite circRNAs being traditionally considered by-products rather than independently functional, circRNAs play many vital roles, such as in host immunity during viral infection. However, in shrimp, these remain largely unexplored. Therefore, this study aims to identify circRNAs in Litopenaeus vannamei in the context of WSSV infection, one of the most eradicative pathogens threatening shrimp populations worldwide. We identified 290 differentially expressed circRNAs (DECs) in L. vannamei upon WSSV infection. Eight DECs were expressed from their parental genes, including alpha-1-inhibitor-3, calpain-B, integrin-V, hemicentin-2, hemocytin, mucin-17, proPO2, and rab11-FIP4. These were examined quantitatively by qRT-PCR, which revealed the relevant expression profiles to those obtained from circRNA-Seq. Furthermore, the structural and chemical validation of the DECs conformed to the characteristics of circRNAs. One of the functional properties of circRNAs as a miRNA sponge was examined via the interaction network between DECs and WSSV-responsive miRNAs, which highlighted the targets of miRNA sponges. Our discovery could provide insight into the participation of these ncRNAs in shrimp antiviral responses. • The circRNAs in Litopenaeus vannamei upon WSSV infection are herein described for the first time. • A total of 290 differentially expressed circRNAs (DECs) were identified using circRNA-Seq. • The transcriptional level determined by qRT-PCR was consistent with circRNA-Seq. • The DECs were confirmed by PCR amplification, RNase R treatment, and Sanger sequencing. • The miRNA-circRNA interaction network revealed various putative miRNA sponges for circRNAs. [ABSTRACT FROM AUTHOR]

The goal of this study was to investigate the effects of dietary supplementation with β-glucan and synbiotics (i.e., microencapsulated Bacillus subtilis and Pediococcus acidilactici ) on immune gene expression and immune parameters of Pacific white shrimp ( Litopenaeus vannamei ) as well as modulation of immune parameters when shrimp were exposed to ammonia stress. Shrimp were fed one of four treatment diets [basal diet (C), β-glucan-containing diet (β-glu), β-glucan plus B. subtilis -containing diet (β-glu + Bs ), and β-glucan plus P. acidilactici -containing diet (β-glu + Pa )] for 90 days. The microencapsulated bacteria colonized in the proximal and central parts of the shrimp intestine. The β-glu diet increased expression of lipopolysaccharide and β-1,3-glucan-binding protein ( LGBP ), and the β-glu + Bs diet induced expression of the serine protease and prophenoloxidase genes. In addition, β-glu + Pa increased the LGBP transcription. However, the mRNA level of peroxinectin did not differ significantly among the experimental groups. Shrimp fed the β-glu and β-glu + Pa diets had higher superoxide dismutase activity (SOD) compared to shrimp fed the basal diet. In addition, the β-glu + Bs and β-glu + Pa diets increased phenoloxidase activity (PO). However, there were no significant differences in total hemocyte count (THC) and lysozyme activity among the treatment diets. Ammonia stress led to increased THC and decreased SOD activity, but it seemed to have no adverse effects on PO and lysozyme activity. Thus, both β-glucan and synbiotic supplementation had beneficial effects on several immune parameters, and the effect of dietary synbiotics was superior to that of β-glucan only for PO activity. [ABSTRACT FROM AUTHOR]

Abstract: Fatty acid delta-6 desaturase (fads2)-like gene from Nile tilapia (Oreochromis niloticus) was characterized and designated as oni-fads2. The Oni-FADS2 showed the typical structure of microsomal FADS2. The presence of oni-fads2 transcripts in unfertilized eggs demonstrated the maternal role of Nile tilapia in providing the oni-fads2 transcript in their eggs. In addition, the expression of oni-fads2 was detectable in embryos throughout the hatching stage. Real-time reverse transcription-PCR revealed that oni-fads2 was expressed at a high level in all the brain regions, liver, and testis. Recombinant yeast (RY) was generated by transformation of Saccharomyces cerevisiae with the plasmid containing oni-fads2 driven by the Gal1 promoter (pYoni-fads2). The conspicuous expression of RY was detectable by RT-PCR after induction with galactose for 24h. When RY was induced with galactose, it exhibited 39% and 7% of delta-6 desaturase (∆6) activity toward C18:2n6 and C18:3n3, respectively. Additionally, it displayed 4% of delta-5 desaturase (∆5) activity toward C20:3n6, indicating that Oni-FADS2 had ∆5 and ∆6 bifunction. [Copyright &y& Elsevier]

Combining germ cell cryopreservation with transplantation is a powerful tool for preserving the genetic resources of various fish strains. In this germ cell transplantation system, since only type A spermatogonia (ASGs) among the various testicular cells have the potency to colonize recipient gonads and resume gametogenesis, techniques for visualization, isolation, and tracing of ASGs facilitate ease of manipulation. Here we established an ASG visualization technique in two Salmo species: brown trout and Atlantic salmon. We applied a monoclonal antibody (antibody No. 95), which specifically recognizes a cell surface antigen of ASGs obtained from rainbow trout, against Salmo species. Immunohistochemistry studies showed that antibody No. 95 also specifically detected the ASGs of brown trout and Atlantic salmon. Furthermore, marker-gene analyses of brown trout testicular cells sorted by flow cytometry using Alexa flour 488-conjugated antibody No. 95 revealed that fluorescent-positive cells possessed molecular characteristics of ASGs. Finally, to investigate the transplantability of antibody-positive cells, ASGs pre-stained with antibody No. 95 were transplanted into the peritoneal cavity of rainbow trout larvae. As a result, fluorescent cells incorporated into the recipient gonads, suggesting that the visualized ASGs possess transplantability. Thus, it is expected that the ASG visualization methods developed in this study will facilitate the development of more efficiency of germ cell transplantation techniques in Salmo species. • Monoclonal Antibody recognizing type A spermatogonia (ASGs) of Salmo was selected. • Live ASGs could be visualized and isolated by the antibody labeled with Alexa 488. • ASGs visualized by the antibody could be used for germ cell transplantation. • These techniques facilitate ease of germ cell manipulation in Salmo species. [ABSTRACT FROM AUTHOR]