Regulation of Gonadotropin-Releasing Hormone (GnRH)-Receptor Gene Expression in Tilapia: Effect of GnRH and Dopamine

The present work was designed to study certain aspects of the endocrine regulation of gonadotropin-releasing hormone receptor (GnRH-R) in the pituitary of the teleost fish tilapia. A GnRH-R was cloned from the pituitary of hybrid tilapia (taGnRH-R) and was identified as a typical seven-transmembrane receptor. Northern blot analysis revealed a single GnRH-R transcript in the pituitary of approximately 2.3 kilobases. The taGnRH-R mRNA levels were significantly higher in females than in males. Injection of the salmon GnRH analog (sGnRHa; 5–50 μg/kg) increased the steady-state levels of taGnRH-R mRNA, with the highest response recorded at 25 μg/kg and at 36 h. At the higher dose of sGnRHa (50 μg/kg), taGnRH-R transcript appeared to be down-regulated. Exposure of tilapia pituitary cells in culture to graded doses (0.1–100 nM) of seabream (sbGnRH = GnRH I), chicken II (cGnRH II), or salmon GnRH (sGnRH = GnRH III) resulted in a significant increase in taGnRH-R mRNA levels. The highest levels of both LH release and taGnRH-R mRNA levels were recorded after exposure to cGnRH II and the lowest after exposure to sbGnRH. The dopamine-agonist quinpirole suppressed LH release and mRNA levels of taGnRH-R, indicating an inhibitory effect on GnRH-R synthesis. Collectively, these data provide evidence that GnRH in tilapia can up- regulate, whereas dopamine down-regulates, taGnRH-R mRNA levels

Analysis of the Central Nervous System Transcriptome of the Eastern Rock Lobster Sagmariasus verreauxi Reveals Its Putative Neuropeptidome

Neuropeptides have been discovered in many arthropod species including crustaceans. The nature of their biological function is well studied and varies from behavior modulation to physiological regulation of complex biochemical processes such as metabolism, molt and reproduction. Due to their key role in these fundamental processes, neuropeptides are often targeted for modulating these processes to align with market demands in commercially important species. We generated a comprehensive transcriptome of the eyestalk and brain of one of the few commercially important spiny lobster species in the southern Hemisphere, the Eastern rock lobster Sagmariasus verreauxi and mined it for novel neuropeptide and protein hormone-encoding transcripts. We then characterized the predicted mature hormones to verify their validity based on conserved motifs and features known from previously reported hormones. Overall, 37 transcripts which are predicted to encode mature full-length/partial peptides/proteins were identified, representing 21 peptide/protein families/subfamilies. All transcripts had high similarity to hormones that were previously characterized in other decapod crustacean species or, where absent in crustaceans, in other arthropod species. These included, in addition to other proteins previously described in crustaceans, prohormone-3 and prohormone-4 which were previously identified only in insects. A homolog of the crustacean female sex hormone (CFSH), recently found to be female-specific in brachyuran crabs was found to have the same levels of expression in both male and female eyestalks, suggesting that the CFSH female specificity is not conserved throughout decapod crustaceans. Digital gene expression showed that 24 out of the 37 transcripts presented in this study have significant changes in expression between eyestalk and brain. In some cases a trend of difference between males and females could be seen. Taken together, this study provides a comprehensive neuropeptidome of a commercially important crustacean species with novel peptides and protein hormones identified for the first time in decapods

Analysis of the distribution and structure of integrated banana streak virus DNA in a range of Musa cultivars

The cDNA encoding the glycoprotein α (GPα) subunit of tilapia (Oreochromis mossambicus) was partially cloned using RACE-polymerase chain reaction (PCR) technique. The amplified cDNA was found to be 583 bases long, and to consist of a portion of the signal peptide, the full sequence encoding the mature peptide (94 amino acids) and the 3′ untranslated region. Northern blot analysis revealed a single band of approximately 600 bp. Alignment of the deduced amino acids of the mature protein showed that the tilapia GPα subunit shares more than 80% identity with that of other perciform fish (i.e. striped bass, sea bream and yellowfin porgy) and less than 70% with that of more taxonomically remote fish and other vertebrates. Exposure of dispersed tilapia pituitary cells to salmon gonadotropin-releasing hormone (sGnRH) elevated GPα mRNA levels via both PKC and cAMP-protein kinase A (PKA) pathways. The transcript levels were also regulated by pituitary adenylate cyclase activating polypeptide (PACAP) and neuropeptide Y (NPY), both acting through PKC and PKA pathways. Moreover, a combined treatment of PACAP or NPY with GnRH seems to have an additive effect on the GPα subunit gene transcription. These results suggest that in tilapia the expression of GPα subunit is regulated by GnRH mainly via PKC and PKA pathways. Furthermore, PACAP and NPY can elevate the GnRH-stimulated GPα subunit transcription and can directly affect the subunit mRNA levels, via the same transduction pathways

A one-phase interior point method for nonconvex optimization

The work of Wachter and Biegler suggests that infeasible-start interior point methods (IPMs) developed for linear programming cannot be adapted to nonlinear optimization without significant modification, i.e., using a two-phase or penalty method. We propose an IPM that, by careful initialization and updates of the slack variables, is guaranteed to find a first-order certificate of local infeasibility, local optimality or unboundedness of the (shifted) feasible region. Our proposed algorithm differs from other IPM methods for nonconvex programming because we reduce primal feasibility at the same rate as the barrier parameter. This gives an algorithm with more robust convergence properties and closely resembles successful algorithms from linear programming. We implement the algorithm and compare with IPOPT on a subset of CUTEst problems. Our algorithm requires a similar median number of iterations, but fails on only 9% of the problems compared with 16% for IPOPT. Experiments on infeasible variants of the CUTEst problems indicate superior performance for detecting infeasibility. The code for our implementation can be found at https://github.com/ohinder/OnePhase .Comment: fixed typo in sign of dual multiplier in KKT syste

Effects of GnRHa treatment during vitellogenesis on the reproductive physiology of thermally challenged female Atlantic salmon (Salmo salar)

Tasmanian Atlantic salmon (S. salar) broodstock can experience temperatures above 20 °C, which impairs reproductive development and inhibits ovulation. The present study investigated the prolonged use of gonadotropin releasing hormone analogue (GnRHa) during vitellogenesis as a means of maintaining endocrine function and promoting egg quality at elevated temperature in maiden and repeat spawning S. salar. GnRHa-treatment during vitellogenesis did not compensate for the negative effects of thermal challenge on the timing of ovulation, egg size, egg fertility or embryo survival in any fish maintained at 22 °C relative to 14 °C. The lack of effectiveness was reflected by the endocrine data, as plasma follicle stimulating hormone and luteinising hormone levels were not different between treated and untreated groups at 22 °C. Furthermore, plasma testosterone and E2 levels were unchanged in GnRHa-treated fish at 22 °C, and plasma levels were generally lower in both groups maintained at 22 °C relative to 14 °C. Transcription of vitellogenin, and zona pellucida B and C was not enhanced in GnRHa-treated fish relative to untreated fish at 22 °C, presumably due to observed suppression of plasma E2. These results indicate that thermal impairment of reproduction is likely to occur on multiple levels, and is difficult to overcome via hormonal manipulation

Assessing the pyloric caeca and distal gut microbiota correlation with flesh color in Atlantic salmon (Salmo salar L., 1758)

The Atlantic salmon (Salmo salar L., 1758) is a temperate fish species native to the northern Atlantic Ocean. The distinctive pink–red flesh color (i.e., pigmentation) significantly affects the market price. Flesh paleness leads to customer dissatisfaction, a loss of competitiveness, a drop in product value and, consequently, severe economic losses. This work extends our knowledge on salmonid carotenoid dynamics to include the interaction between the gut microbiota and flesh color. A significant association between the flesh color and abundance of specific bacterial communities in the gut microbiota suggests that color may be affected either by seeding resilient beneficial bacteria or by inhibiting the negative effect of pathogenic bacteria. We sampled 96 fish, which covered all phenotypes of flesh color, including the average color and the evenness of color of different areas of the fillet, at both the distal intestine and the pyloric caeca of each individual, followed by 16S rRNA sequencing at the V3-V4 region. The microbiota profiles of these two gut regions were significantly different; however, there was a consistency in the microbiota, which correlated with the flesh color. Moreover, the pyloric caeca microbiota also showed high correlation with the evenness of the flesh color (beta diversity index, PERMANOVA, p = 0.002). The results from the pyloric caeca indicate that Carnobacterium, a group belonging to the lactic acid bacteria, is strongly related to the flesh color and the evenness of the color between the flesh areas

Applying the Power of Transcriptomics: Understanding Male Sexual Development in Decapod Crustacea

The decapod Crustacea are the most species-rich order of the Crustacea and include some of the most charismatic and highly valued commercial species. Thus the decapods draw a significant research interest in relation to aquaculture, as well as gaining a broader understanding of these species’ biology. However, the diverse physiology of the group considered with the lack of a model species have presented an obstacle for comparative analyses. In reflection of this, the recent integration of comparative transcriptomics has rapidly advanced our understanding of key regulatory pathways and developmental phenomena, an example being our understanding of sexual development. We discuss our work in the Eastern spiny lobster, Sagmariasus verreauxi, in the context of what is currently known about male sexual development in the decapods, highlighting the importance of transcriptomic techniques in achieving our recent advancements. We describe the progression made in our understanding of male sexual differentiation and maturation, as mediated by the insulin-like androgenic gland hormone (IAG), integrating the role of regulatory binding proteins (IGFBPs), a tyrosine kinase insulin receptor (TKIR), as well as the upstream effect of neuroendocrine hormones (GIH and MIH). We then consider the less well understood mechanism of male sex determination, with an emphasis on what we believe to be the key regulatory factors, the Dsx- and mab-3-related transcription factors (Dmrts). Finally, we discuss the function of the antennal gland (AnG) in sexual development, relating to the emergence of male-biased upregulation in the AnG in later sexual maturation and the sexually dimorphic expression of two key genes Sv-TKIR and Sv-Dmrt1. We then present the AnG as a case study to illustrate how comparative transcriptomic techniques can be applied to guide preliminary analyses, like the hypothesis that the AnG may function in pheromone biosynthesis. In summary, we describe the power of transcriptomics in facilitating the progress made in our understanding of male sexual development, as illustrated by the commercial decapod species, S. verreauxi. Considering future directions, we suggest that the integration of multiple omics-based techniques offers the most powerful tool to ensure we continue to piece together the biology of the important group of decapod Crustacea

Deploying new generation sequencing for the study of flesh color depletion in Atlantic Salmon (Salmo salar)

The flesh pigmentation of farmed Atlantic salmon is formed by accumulation of carotenoids derived from commercial diets. In the salmon gastrointestinal system, the hindgut is considered critical in the processes of carotenoids uptake and metabolism. In Tasmania, flesh color depletion can noticeably affect farmed Atlantic salmon at different levels of severity following extremely hot summers. In this study, RNA sequencing (RNA-Seq) was performed to investigate the reduction in flesh pigmentation. Library preparation is a key step that significantly impacts the effectiveness of RNA sequencing (RNA-Seq) experiments. Besides the commonly used whole transcript RNA-Seq method, the 3’ mRNA-Seq method is being applied widely, owing to its reduced cost, enabling more repeats to be sequenced at the expense of lower resolution. Therefore, the output of the Illumina TruSeq kit (whole transcript RNA-Seq) and the Lexogen QuantSeq kit (3’ mRNA-Seq) was analyzed to identify genes in the Atlantic salmon hindgut that are differentially expressed (DEGs) between two flesh color phenotypes