Tuan Nguyen Viet

Background: The Norway lobster, Nephrops norvegicus, is economically important in European fisheries and is a key organism in local marine ecosystems. Despite multi-faceted scientific interest in this species, our current knowledge of genetic resources in this species remains very limited. Here, we generated a reference de novo transcriptome for N. norvegicus from multiple tissues in both sexes. Bioinformatic analyses were conducted to detect transcripts that were expressed exclusively in either males or females. Patterns were validated via RT-PCR.

The Australian redclaw crayfish (Cherax quadricarinatus) has recently received attention as an emerging candidate for sustainable aquaculture production in Australia and worldwide. More importantly, C. quadricarinatus serves as a good model organism for the commercially important group of decapod crustaceans as it is distributed worldwide, easy to maintain in the laboratory and its reproductive cycle has been well documented. In order to better understand the key reproduction and development regulating mechanisms in decapod crustaceans, the molecular toolkit available for model organisms such as C. quadricarinatus must be expanded. However, there has been no study undertaken to establish the C. quadricarinatus neuropeptidome. Here we report a comprehensive study of the neuropeptide genes expressed in the eyestalk in the Australian crayfish C. quadricarinatus. We characterised 53 putative neuropeptide-encoding transcripts based on key features of neuropeptides as characterised in other species. Of those, 14 neuropeptides implicated in reproduction regulation were chosen for assessment of their tissue distribution using RT-PCR. Further insights are discussed in relation to current knowledge of neuropeptides in other species and potential follow up studies. Overall, the resulting data lays the foundation for future gene-based neuroendocrinology studies in C. quadricarinatus. Neuropeptides play an important role in the regulation of both vertebrate and invertebrate reproduction 1–3. By binding to receptors in target tissues, neuropeptides elicit a number of downstream cascades including changes in secondary messengers, phosphorylation and eventually altered transcription activity which leads to a biological response 4. In silico neuropeptidome research aimed at understanding different physiological aspects of crustacean biology has been extensively studied in several crustacean species, including Scylla paramamosain 5 , Macrobrachium rosenbergii 6 , Procambarus clarkii 7 , Homarus americanus and Sagmariasus verreauxi 8,9. The crustacean eyestalk (together with the supra-oesophageal and thoracic ganglia), is known to be the primary source of expression and secretion of key factors which regulate many aspects of crustacean behaviour and physiology, including growth and reproduction 10–12. These biological processes operate under the multifaceted control of biogenic amines, neuropeptides, hormones and lipid metabolites such as methyl farnesoate and ecdys-teriods 13. Crustacean neuropeptide research has focused on a number of key factors that are related to reproduction and sexual development. For instance, the immunoreactivity of antibodies raised against gonadotropin releasing hormone (GnRH) in the central nervous system of the giant freshwater prawn M. rosenbergii suggests a vertebrate-like reproductive system in crustaceans 14. Similarly, a GnRH-like peptide was found in the white leg shrimp Litopenaeus vannamei 15 , and the primary structure of GnRH was then reported by mass spectral analysis

The striped catfish (Pangasianodon hypophthalmus) culture industry in theMekong Delta in Vietnamhas developed
rapidly over the past decade. The culture industry nowhowever, faces some significant challenges, especially related
to climate change impacts notably from predicted extensive saltwater intrusion intomany lowtopographical coastal
provinces across the Mekong Delta. This problem highlights a need for development of culture stocks that can
tolerate more saline culture environments as a response to expansion of saline water-intruded land. While a
traditional artificial selection program can potentially address this need, understanding the genomic basis of
salinity tolerance can assist development of more productive culture lines. The current study applied a
transcriptomic approach using Ion PGM technology to generate expressed sequence tag (EST) resources from the
intestine and swim bladder from striped catfish reared at a salinity level of 9 ppt which showed best growth
performance. Total sequence data generated was 467.8 Mbp, consisting of 4,116,424 reads with an average length
of 112 bp. De novo assembly was employed that generated 51,188 contigs, and allowed identification of 16,116
putative genes based on the GenBank non-redundant database. GO annotation, KEGG pathway mapping, and
functional annotation of the EST sequences recovered with a wide diversity of biological functions and processes.
In addition, more than 11,600 simple sequence repeats were also detected. This is the first comprehensive analysis
of a striped catfish transcriptome, and provides a valuable genomic resource for future selective breeding programs
and functional or evolutionary studies of genes that influence salinity tolerance in this important culture species.

Striped catfish (Pangasianodon hypophthalmus) is a commercially important freshwater fish used in inland
aquaculture in the Mekong Delta, Vietnam. The culture industry is facing a significant challenge however from
saltwater intrusion into many low topographical coastal provinces across the Mekong Delta as a result of
predicted climate change impacts. Developing genomic resources for this species can facilitate the production
of improved culture lines that can withstand raised salinity conditions, and so we have applied highthroughput
Ion Torrent sequencing of transcriptome libraries fromsix target osmoregulatory organs fromstriped
catfish as a genomic resource for use in future selection strategies. We obtained 12,177,770 reads after trimming
and processing with an average length of 97 bp. De novo assemblies were generated using CLC Genomic
Workbench, Trinity and Velvet/Oases with the best overall contig performance resulting from the CLC assembly.
De novo assembly using CLC yielded 66,451 contigswith an average length of 478 bp and N50 length of 506 bp. A
total of 37,969 contigs (57%) possessed significant similarity with proteins in the non-redundant database.
Comparative analyses revealed that a significant number of contigsmatched sequences reported in other teleost
fishes, ranging in similarity from 45.2% with Atlantic cod to 52% with zebrafish. In addition, 28,879 simple
sequence repeats (SSRs) and 55,721 single nucleotide polymorphisms (SNPs)were detected in the striped catfish
transcriptome. The sequence collection generated in the current study represents the most comprehensive
genomic resource for P. hypophthalmus available to date. Our results illustrate the utility of next-generation
sequencing as an efficient tool for constructing a large genomic database for marker development in nonmodel
species.

Increasing salinity levels in freshwater and coastal environments caused by sea level rise linked to climate change
is now recognized to be a major factor that can impact fish growth negatively, especially for freshwater teleost
species. Striped catfish (Pangasianodon hypophthalmus) is an important freshwater teleost that is now widely
farmed across the Mekong River Delta in Vietnam. Understanding the basis for tolerance and adaptation to raised
environmental salinity conditions can assist the regional culture industry to mitigate predicted impacts of climate
change across this region.
Attempt of next generation sequencing using the ion proton platform results in more than 174 million raw reads
from three tissue libraries (gill, kidney and intestine). Reads were filtered and de novo assembled using a variety
of assemblers and then clustered together to generate a combined reference transcriptome. Downstream analysis
resulted in a final reference transcriptome that contained 60,585 transcripts with an N50 of 683 bp. This resource
was further annotated using a variety of bioinformatics databases, followed by differential gene expression analysis
that resulted in 3062 transcripts that were differentially expressed in catfish samples raised under two experimental
conditions (0 and 15 ppt). A number of transcripts with a potential role in salinity tolerance were then classified
into six different functional gene categories based on their gene ontology assignments. These included; energy metabolism,
ion transportation, detoxification, signal transduction, structural organization and detoxification.
Finally, we combined the data on functional salinity tolerance genes into a hypothetical schematic model that
attempted to describe potential relationships and interactions among target genes to explain the molecular pathways
that control adaptive salinity responses in P. hypophthalmus. Our results indicate that P. hypophthalmus exhibit
predictable plastic regulatory responses to elevated salinity by means of characteristic gene expression
patterns, providing numerous candidate genes for future investigations.
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