DENUGREC · Development of genomic tools for assessing nutrition, growth and reproduction issues in farmed crustacean species

Development of genomic tools for assessing nutrition, growth and reproduction issues in farmed crustacean species

In crustaceans, high-throughput sequencing technology (usually referred to as Next Generation Sequencing; NGS), in particular sequencing of the entire transcriptome (referred to as RNA sequencing; RNA-seq) has been proved useful in rapidly identifying the transcript sequences encoding for proteins that are at the core of developmental biology, growth reproduction, nutrition, circadian rhythms and immune and stress response and in some cases, propel the elucidation of pathways and novel groups of genes. DeNuGReC project have synthetized transcriptomes of 11 decapod crustacean species, mainly of aquaculture interest but also ecological to improve the conservation of the species in wild and maintain biodiversity. The advancement of data analysis methodologies in RNA-Seq were lectured in the Institute of Marine Sciences (CSIC) of Barcelona to researchers and early stage PhD students in the field of marine biology and aquaculture with little to no knowledge of RNA-Seq technology but planning to explore (and apply) high throughput sequencing technologies and bioinformatics methods in their research. The main second research objective of DeNuGReC was to use transcriptomic data to provide predictive tools for assessing metabolic pathways in the main metabolic processes. The specific objectives achieved during DeNuGReC were: 1. Transcriptome sequences. Transcriptomes of several tissues (testes, ovary, vas deferens, hepatopancreas, brain, eyestalk and thoracic ganglia, stomach, abdominal muscle and claw, cuticle antennal glands) and development stages (adult males and females, juveniles and larvae) of 11 different crustacean species were generated. 2. Optimal guideline for RNA-seq projects. Applications and opportunities in non-model decapod crustacean species were discussed. 3. Reproduction – Ovarian maturation. Gene sequences related to reproductive performance and gonadal maturation were identified in the banana prawn F. merguiensis and N. norvegicus transcriptome. Potential genes to be identified as the Gonad Stimulation Factor (GSF) has been proposed for crustaceans. 4. Reproduction – Gender issues. Among the most highly expressed genes between males and females of N. norvegicus were serine threonine protein kinase (STPK) in testis and vitellogenin (Vg) in female hepatopancreas. 5. Reproduction – Regulation. To compare the C. quadricarinatus and N. norvegicus neuropeptidomes, in particularly the DEG related to reproduction regulation. 6. Reproduction – Social dominance hierarchy. In males of Macrobrachium rosenbergii, a social dominance hierarchy among mature individuals is present where dominant males suppress the growth of subordinate males of the same age class. The DGE between this type of males could facilitate their adaptation to changing external environmental conditions. 7. Sexual differentiation. The insulin-like androgenic gland hormone (IAG) responsible for male sexual differentiation was characterized and expressed in the Eastern spiny lobster S. verreauxi, redclaw crayfish C. quadricarinatus and giant freshwater prawn M. rosenbergii. The first gene detected in a decapod crustacean encoding an insulin-like receptor was identified and characterized in the prawn M. rosenbergii (Mr-IR). 8. Growth – Metamorphosis. The metamorphosis process could account for a considerably high percentile of transcribed sequences over a short period of time, with the genome encoding for different life forms. Such important changes in expression patterns for a high number of genes poses a challenge for accurately assign each gene to a function. 9. Circadian rhythms. Clock gene homologs such as timeless, bmal1, period and cryptochrome 2 were expressed in the eyestalks of the Norway lobster N. norvegicus. 10. Immunological response. Shrimps (F. merguiensis) infected with hepatopancreatic parvo-like virus (HPV) reported differentially expressed genes involved in the immune response of crustaceans, including pattern recognition proteins, antimicrobial peptides, components of the prophenoloxidase system, and antiviral activity. 11. Evolution and adaptation. A comparative transcriptomics approach among divergent lineages of P. australiensis and their hybrid offspring has identified candidate genes that are potentially involved in the process of speciation.

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