- Email: [email protected]
Omics and cytokine discovery in fish: Presenting the Yellowtail kingfish (Seriola lalandi) as a case study
Accepted Manuscript Omics and cytokine discovery in fish: Presenting the yellowtail kingfish (Seriola lalandi) as a case study Gregory Jacobson, Simon Muncaster, Koen Mensink, Maria Forlenza, Nick Elliot, Grant Broomfield, Beth Signal, Steve Bird PII:
S0145-305X(17)30190-8
DOI:
10.1016/j.dci.2017.04.001
Reference:
DCI 2862
To appear in:
Developmental and Comparative Immunology
Received Date: 9 March 2017 Revised Date:
1 April 2017
Accepted Date: 1 April 2017
Please cite this article as: Jacobson, G., Muncaster, S., Mensink, K., Forlenza, M., Elliot, N., Broomfield, G., Signal, B., Bird, S., Omics and cytokine discovery in fish: Presenting the yellowtail kingfish (Seriola lalandi) as a case study, Developmental and Comparative Immunology (2017), doi: 10.1016/ j.dci.2017.04.001. This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
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Omics and cytokine discovery in fish: Presenting the Yellowtail kingfish (Seriola
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lalandi) as a case study.
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Gregory Jacobson1, Simon Muncaster2, Koen Mensink3, Maria Forlenza3, Nick Elliot1,
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Grant Broomfield1, Beth Signal1, Steve Bird1
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Engineering, University of Waikato, Private Bag 3105, Hamilton 3240 New Zealand
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Molecular Genetics, Department of Biological Sciences, School of Science and
School Applied Science, Bay of Plenty Polytechnic, 70 Windermere Dr, Poike,
Tauranga 3112 New Zealand
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University, Wageningen, The Netherlands
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Cell Biology and Immunology Group, Department of Animal Sciences, Wageningen
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Corresponding author:
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Email: [email protected], Tel: (+64) 07 838 4723
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Highlights
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• The ability to sequence genomes and transcriptomes has accelerated cytokine discovery within fish, allowing informative studies to be carried out in organisms where no genetic information existed previously.
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• Undertaking RNA-Seq on selected tissues of S. lalandi has identified a large number of cytokine and receptor genes within this species, which have been deposited within genbank.
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• The cytokine and receptor genes discovered using transcriptomic approaches can be used in future studies, to allow the effective monitoring of fish immunity with changes to its environment or in response to disease.
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Abstract A continued programme of research is essential to overcome production
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bottlenecks in any aquacultured fish species. Since the introduction of genetic and
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molecular techniques, the quality of immune research undertaken in fish has greatly
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improved. Thousands of species specific cytokine genes have been discovered,
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which can be used to conduct more sensitive studies to understand how fish
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physiology is affected by aquaculture environments or disease. Newly available
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transcriptomic technologies, make it increasingly easier to study the immunogenetics
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of farmed species for which little data exists. This paper reviews how the application
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of transcriptomic procedures such as RNA Sequencing (RNA-Seq) can advance fish
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research. As a case study, we present some preliminary findings using RNA-Seq to
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identify cytokine related genes in Seriola lalandi. These will allow in-depth
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investigations to understand the immune responses of these fish in response to
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environmental change or disease and help in the development of therapeutic
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approaches.
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1. Introduction
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Both innate and adaptive immune responses utilise regulatory proteins called
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cytokines, which are groups of small proteins (~5–20 kDa) important in cell signalling
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and modulation of an organism’s immunity. Cytokines are produced by a broad
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range of cells and each cytokine can be produced by more than one cell type, where
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they can enhance or inhibit the action of other cytokines in complex ways. They are
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important in health and disease and are important in immune responses to bacterial,
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viral, or parasitic pathogens (Barrett, 1996; Vilček, 2003). Cells of the immune
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system (macrophages, granulocytes, dendritic cells, B-cells, T-cells and mast cells)
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and endothelial cells, fibroblasts and various stromal cells secrete cytokines that bind
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to specific cellular receptors through autocrine or paracrine mechanisms. They effect
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the behaviour of the cells around them, helping to modulate the balance between
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humoral and cell-based immune responses and the maturation, growth, and
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responsiveness of particular cell populations. In mammals, there are over 100
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separate genes coding for cytokine-like activities, many with overlapping functions
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and many still needing more study. Today, the term “cytokine” encompasses
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interferons (IFN’s), the interleukins (IL’s), the tumour necrosis factor family (TNF’s),
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the transforming growth factor family (TGF’s), the colony stimulating factor family
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(CSF’s) and the chemokine family.
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The activity of cytokines have been known of and studied in mammals since 1957,
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where interferon-alpha (IFN-α) was fist investigated (Lindenmann et al., 1957).
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However, it was not until the early 1980’s that the molecular cloning of the first
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cytokines (IFNα, IL-1, IL-2 and TNFα) was achieved (Dinarello, 2007). Studies into
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the regulation of fish immunity, began in the early 1990’s (Balm et al., 1995; Jang et
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al., 1994; 1995; Verburg-van Kemenade et al., 1995) due to the importance of fish
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within the aquaculture industry and the need to understand the regulation of
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immunity, due to problems with disease. The first cytokine to be cloned in fish was
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TGF-β (Sumathy et al., 1997; Hardie et al., 1998), followed closely by IL-1β (Zou et
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al., 1999a; 1999b) and a few years later by TNF-α (Laing et al., 2001). Both of these
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were discovered using a homology-based cloning approach, where known
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particularly conserved sequence. These regions were then used to design
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degenerate oligonucleotides to enable amplification of the gene from a targeted fish
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species. This approach was limited and only worked well when the genes being
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targeted had not diverged too far during evolution. However, additional approaches
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that became available shortly after such as subtractive hybridization (Sangrador-
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Vegas et al., 2001) and expressed sequence tags (Altmann et al., 2003) also
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contributed to the discovery of cytokines in fish, helping to identify homologues of IL-
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8 and IFN-α.
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These early approaches of cytokine discovery eventually fell out of favour with the availability of expressed sequence tag (EST) sequence processing (Lindlöf, 2003)
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and the subsequent ability to sequence genomes which has become easier through
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the development of next generation sequencing (NGS) technology (van Dijk et al.,
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2014). One of the first fish genomes sequenced was from Takifugu rubripes
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(Aparicio et al., 2002) and opened up the ability to search for cytokines in fish that
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shared very little homology to their mammalian homologues. IL-2, IL-21 and IL-6
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were some of the first cytokines to be discovered (Bird et al., 2005a; 2005b), where
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conservation of synteny was found between the mammalian and fish genomes,
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allowing their discovery. More recently, the development of technologies such as
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RNA-Seq and the ability to sequence transcriptomes has transformed immune gene
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discovery and provides the ability to profile immune gene expression in organisms
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where no gene discovery had previously taken place (Dheilly et al., 2014).
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The availability of genomes and transciptomics has revolutionised cytokine
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discovery and research in fish (Figure 1). This review will look at how the application
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of new genetic technologies is aiding fish cytokine discovery and research, will use
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the Yellowtail kingfish (Seriola lalandi) as a case study to demonstrate the power of
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RNA-Seq when studying a new fish species where no cytokine genes have been
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characterised and finish by looking at the types of investigations into the roles of
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cytokines in fish immunity that are now possible in fish.
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Having information at the genetic level available within a fish species, allows
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researchers to carry out more precise investigations into the physiology of an
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organism. Recent advances within the areas of genomics and transcriptomics are
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making these types of studies more accessible to fish research, in species where no
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genetic information has been collected (Dheilly et al., 2014). Currently, there are a
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select number of fish species, where a large amount of genetic data has been
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accumulated, however this has been the result of many years of research, where the
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identification of genes took a significant amount of time (Zou et al., 2010; Secombes
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et al., 2011; Bird et al., 2015; Secombes et al., 2015; 2016). However, new
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sequencing technologies, capable of sequencing thousands to hundreds of millions
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of short DNA strands (not specifically targeting a single sequence) without a prior
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bacterial cloning and plasmid DNA isolation step, have enabled progress in the field
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of gene identification to be accelerated greatly. This began in 2005 with the invention
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of ‘massively parallel’ sequencing technologies (Mardis, 2011), with technological
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progress initially driven by projects such as the Human Genome Project, HapMap
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Project and ENCODE.
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Since the emergence of these next-generation sequencing platforms, new
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technology continues to develop at a fast pace, that improves on previous
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methodology or a new approach is developed, enabling increases in sequencing
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efficiency and lower costs. There is fierce competition between companies
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developing this technology and incentives, such as the Archon Genomics XPRIZE,
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which had a goal of sequencing 100 human genomes in 30 days for less than
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USD$1000 per genome (Kedes & Liu, 2010), have helped to push this forward. This
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progress has allowed significant advances in the area of genomics and has led to
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genome sequencing being achievable in a wider range of organisms of scientific or
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commercial interest, which includes a number of fish species. Currently, there are
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twelve fish with their entire genomes sequenced that are being assembled and
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annotated and are publically available , that includes Japanese Pufferfish, Takifugu
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rubripes (Aparicio et al., 2002), Green Spotted Puffer, Tetradon nigoviridis (Jaillon et
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al., 2004), Japanese rice fish, Oryzias latipes (Kasahara et al., 2007), Atlantic Cod,
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aculeatus (Jones et al., 2012), Zebrafish, Danio rerio (Howe et al., 2013), African
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coelacanth, Latimeria chalumnae (Amemiya et al., 2013), Southern platyfish,
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Xiphophorus maculatus (Schartl et al., 2013), Mexican cavefish, Astyanax
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mexicanus (McGaugh et al., 2014), Nile tilapia, Oreochromis niloticus (Brawand et
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al., 2014), spotted gar, Lepisosteus oculatus (Braasch et al., 2016), Amazon molly,
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Poecilia
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number of species for which the genome has or is been sequenced, but is still in
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draft form or incomplete. This includes fish such as Atlantic Salmon (Davidson et al.,
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2010), rainbow trout (Berthelot et al., 2016), channel catfish (Liu et al, 2016), large
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yellow croaker (Ao et al., 2015), Northern snakehead (Xu et al., 2017), Chinese
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clearhead icefish (Liu et al., 2017), common carp (Petit et al., 2017), Asian arowana
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(Austin et al., 2015), Asian seabass (Vij et al., 2016) and another 66 teleost species
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(Malmstrøm et al., 2017)
(http://www.ensembl.org/Poecilia_formosa/Info/Index)
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Transcriptomics is also an area of molecular biology that is beginning to have a
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large impact on fish cytokine research. Unlike the genome which is static, the
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transcriptome of a cell is dynamic and is continually changing, due to the differences
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in gene expression and the levels at which they are expressed. Techniques in this
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area are less targeted and allow the study of 1000’s of genes simultaneously,
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encoded by the genome from a specific cell, tissue or organism at a specific time or
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under a specific set of conditions. Early studies used DNA microarrays, which is a
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large collection of identified microscopic DNA spots attached to a solid surface, that
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have initially been isolated from the species of interest. These spots will hybridize to
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genes that are expressed in the sample and are detected using fluorescence,
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allowing the relative abundance of nucleic acid sequences in the sample to be
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determined. This approach has been very informative when investigating an
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immunological response of a fish and how they are affected by changes to their
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environment or disease (Tacchi et al., 2011; Tacchi et al., 2012; Pooley et al., 2013;
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Mommens et al., 2014; Trumbic et al., 2015; Cho et al., 2016) and has led to the
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understanding of actual biological pathways in the organism, rather than just
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individual gene expression. However, although a powerful approach, it has been
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limited by the lack of genetic data available in your species of interest beforehand to
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produce the microarray and it is quite a costly and technically difficult approach to
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use.
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The advances being made by NGS have had a dramatic impact on the area of
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transcriptomics and there are newly available technologies, such as RNA
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sequencing (RNA-Seq), that are able to directly sequence the majority of RNA from
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a cell or tissue sample to study all the genes being expressed at a particular point in
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time (Wang et al., 2009; Costa, 2010). The advantage of this is it does not rely on or
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is limited by the previous accumulation of genetic data and there is no requirement of
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knowing the identity of all the transcripts to be analysed prior to analysis. Along with
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cytokine discovery, this method is extremely useful for quantifying levels of gene
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expression in a sample, and can be used for comparative transcriptomics, having the
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potential to provide much more information, including isoform identification, rare
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transcript detection, and SNP detection than previous methods such as microarrays.
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RNA-Seq are newly developed and available technologies that have begun to be
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used for transcriptomic studies in a number of fish species, such as trout (Palstra et
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al., 2013), zebrafish (Hegedus et al., 2009; Ordas et al., 2011), Japanese sea bass
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(Xiang et al., 2010) and the small yellow croaker (Mu et al., 2010). The next section
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will focus on how a transcriptomics approach can be applied to a relatively
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understudied teleost species without a genome available to aid in cytokine discovery.
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3. Yellowtail kingfish (Seriola lalandi) transcriptomic approach
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3.1 Current state of kingfish aquaculture
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Over the last 15 years, there has been acquisition of genetic information in a
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variety of farmed fish species, allowing the implementation of new genetic
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technologies within aquaculture (Bostock et al., 2010; Cerdà et al., 2010; Forné et al.,
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2010; Johansen et al., 2009; Thakur et al., 2008). Before this type of information was
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available, the monitoring of fish immune responses to changes in their environments
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was very arbitrary and normally measured by physiological parameters or fish
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mortalities. With the development of genetic approaches, more focussed studies can
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now be achieved, which have allowed researchers to look in more detail and attempt
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to solve some of the important bottlenecks there are in successful production of a
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particular fish species. In some cases, findings from this research, have then been
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passed down to the fish farmers themselves, who can translate them into good
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farming practice.
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S. lalandi are widely distributed throughout the warm–temperate waters of the
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southern hemisphere. In the wild they reach 1.7 m in length and weigh 56 kg and are
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a highly prized game fish, having excellent flesh quality for a range of product
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options (such as whole fillets, sushi and the highly valued sashimi), giving it
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significant domestic and international market opportunities. Successful aquaculture
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of this species in New Zealand would provide a reliable and controlled production of
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kingfish to an ever growing market. Culture of S. lalandi is already underway in
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Australia where fingerlings are hatched and grown to maturity in sea cages (Primary
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Industries and Resources South Australia, 2011). Already in Japan there is a very
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close related species farmed, the Japanese Yellowtail (Seriola quinqueradiata),
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which forms the basis of a major aquaculture industry in Japan, where wild caught
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juveniles are grown out in sea cages. Despite an increase in demand of these
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farmed species, there is a general lack of scientific and farming knowledge and
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significant problems exist with diseases (Stephens and Savage, 2010; Burger et al.,
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2008; Kawanishi et al., 2005; 2006; Tubbs et al., 2004; Kobayashi, 2010; Mansell et
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al., 2005). Having the appropriate molecular tools to investigate and understand
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these problems will improve the current husbandry of S. lalandi and provide
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countries such as New Zealand interested in its aquaculture with the ability to
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successfully introduce this species.
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To date, limited studies have been done to determine and understand the
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genetics of S. lalandi which would provide invaluable insights into the physiology of
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this fish and allow the determination of the optimum conditions for its culture. Work
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has begun within S. quinqueradiata, where there are 2936 EST sequences from the
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spleen and kidney, available from a study into its immune response and a cDNA
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microarray has been constructed using 1001 selected EST sequences to allow
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immune gene profiling (Darawiroj et al., 2008). Currently, there are very few S.
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lalandi nucleotide sequences deposited within the National Center for Biotechnology
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Information (NCBI) nucleotide database, which include a limited number of genes for
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this species (Supplementary Table 1). Previous research into the immune
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responses of S. lalandi has relied on physiological analysis such as studying mucus
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and serum (Leef & Lee, 2008) or the concentration of blood lactate and plasma
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osmolality (Mansell et al., 2005). A greater understanding is desperately required for
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the immune system of Seriola sp. and NGS technology provides the data needed for
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a comprehensive study of these species. Using this approach will allow the
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characterization of important immune components, such as cytokines, which will
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then allow informative future studies to be carried out.
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3.2 S. lalandi transcriptomic library
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With the availability of genomes within selected fish species, gene identification
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had already accelerated and with the development of new technologies in the area of
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transcriptomics, obtaining genetic information from a new fish species is now faster
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and cheaper. There are a number of different next generation sequencing platforms
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available (Quail et al., 2012) that can be used to produce a transcriptomic database
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using RNA-Seq. However, factors such as the size of the transcriptome to be
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sequenced, how much money is available and read length are important
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considerations that will help determine which one is most suitable to use. In this
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investigation, to identify cytokine genes from S. lalandi, a high throughput next
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Merriman et al., 2012). This approach relies on the detection of hydrogen ions
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released when a complementary nucleotide is incorporated into a strand of DNA,
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leading to a lowering of pH, which is measured by a semi-conductor. This is one of
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the more recent approaches available and its main advantage over other NGS
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platforms, is its overall low cost and size of the read lengths (200-400bp).
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Wild yellowtail kingfish, S. lalandi were caught near Gannet Island, off the coast
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from Kawhia, New Zealand and the pituitary, gonad and spleen were collected and
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used to produce an RNA-Seq library (Muncaster et al., 2017). The raw IonTorrent
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transcriptome sequence libraries had 3,147,845 spleen (Supplementary Figure 1)
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and 3,646,264 gonad/pituitary (Supplementary Figure 2) reads. After undergoing
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de novo assembly using Trinity (Grabherr et al., 2011) 50,570 contigs (length range
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200 to 16046 bp) for the spleen and 13178 contigs (length range 200 to 10497 bp)
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for the gonad/pituitary were constructed. The assembled contigs were submitted to
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the webtool 'FastAnnotator' (Chen et al., 2012), where tBLASTn (Gertz et al., 2006)
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was used to translate each contig into all six reading frames and then use each one
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to search the NCBI's non-redundant protein database. Each identified protein was
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then submitted into Blast2GO (Götz et al., 2008) for the GO term annotation. The
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BLAST search was able to align 32,383 of the spleen contigs and 7898 of the
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pituitary/gonad contigs to known proteins in the non-redundant protein database.
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The GO enrichment analysis from the spleen (Figure 2) and the gonad/pituitary
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(Figure 3) transcripts identified a number involved within immune system processes.
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From these transcripts, cytokines and their receptors were able to be identified and
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submitted into Genbank.
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4. Yellowtail kingfish cytokine and cytokine receptor gene discovery
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During the last 20 years many of the immune genes known in mammalian
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organisms have been identified in teleost fish, particularly in those fish that are
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important for aquaculture, such as trout and salmon. Teleosts have been found to
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have a very similar immune cell repertoire and have many of the protein components
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important for innate and adaptive immune responses that have been characterized 11
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within mammals (Zhu, et al., 2013). This includes important receptors, involved in
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initiating innate responses such as Toll-like receptors (TLR’s) and includes members
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such as TLR-1, TLR-2, TLR-3, TLR-4, TLR-5, TLR-7, TLR-8 and TLR-9 (Pietretti &
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Wiegertjes, 2014). Important cellular markers critical for adaptive responses, such as
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T-cell receptor (TCR),
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histocompatibility complex (MHC) class I and II (Zhu, et al., 2013). In addition, a
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large number of the molecules important in regulating immunity, the cytokines have
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also been characterised and include many of the members of the interleukin (IL)
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family (Secombes et al., 2016), interferon (IFN) family (Zou et al., 2016), tumor
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necrosis factor (TNF) family (Secombes et al., 2016), chemokine family (Bird &
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Tafalla, 2015), colony stimulating factor family (Santos et al., 2006; Wang et al.,
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2008), fibroblast growth factor family (Itoh, 2007) and transforming growth factor
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family (Bobe et al., 2009) . Lastly, there have also been a number of cytokines
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discovered, that are novel to fish, with no known mammalian homologue (Husain et
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al., 2012; Hong et al., 2013; Angosto et al., 2013) and the roles of which remain to
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be determined.
CD4,
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Up until this study, the only cytokine related genes found in S. lalandi were
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partial mRNA sequences for ACKR3, CXCR4, CXCR7, IL-8, IFN-γ genes and
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complete mRNA sequences for IL-1β and TNF-α genes (Supplementary Table 1).
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This lack of information available for cytokine genes means that many previous
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studies that have investigated the effects of stress or pathogens in Seriola species
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have had to use mortality or pathological traits such as serum and mucus production,
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blood glucose and lactate concentrations, muscle pH and lactate concentrations and
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plasma osmolality to determine the physiology of the fish (Mansell et al., 2005; Leef
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& Lee, 2009; Moran et al., 2008). While this provides useful information about the
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effects of disease and stress on S. lalandi, using actual cytokine gene expression
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studies would allow a more in depth analysis of these effects at the molecular level,
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which would be particularly useful for the monitoring of immune system response to
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disease, vaccinations, stress and general fish health (Darawiroj et al., 2008).
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The spleen tissue was targeted for transcriptomics as is the main filter for
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blood-borne pathogens and antigens and is a key organ for iron metabolism and 12
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is important for the regulation of innate and adaptive immune responses locally and
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throughout the whole body (Bronte & Pittet, 2013). The pituitary and gonad
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transcriptome was available as it had been used for a study into sex differentiation.
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However, in mammals evidence exists that cytokines are produced by pituitary cells
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can mediate development, mature function, and cellular organization of the anterior
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pituitary and also directly regulate specific pituitary trophic hormone gene expression
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(Ray & Melmed, 1997). In addition, studies in fish have shown cytokine expression
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does occur in the gonads (Chaves-Pozo et al, 2008). Using the S. lalandi spleen and
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pituitary/gonad transcriptomic databases that were generated and subsequently
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searched, a significant number of cytokine genes and receptors were discovered
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(Table 1) important in both the innate and adaptive immune responses. Most of
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these assembled sequences represent partial mRNA’s that can be translated to
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provide a partial amino acid sequence, however in some cases they can code for the
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full amino acid sequence. For example, using the transcriptomic libraries it was
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possible to assemble the full open reading frame for S. lalandi TNFSF13B, also
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known as B cell activating factor belonging to the TNF family (BAFF). This is an
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important member of the TNF ligand superfamily (Schneider, 2005) and is critical for
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the survival/maturation and proliferation for peripheral B cells, (Rolink & Melchers,
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2002; Stein et al., 2002; Mackay et al., 2003; Mackay and Leung, 2006). BAFF is a
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type II transmembrane protein and is found either anchored to the cell surface or
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released in a soluble form after cleavage by a furin-like protease, with both forms
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having biological activity (Moore et al., 1999; Schneider et al., 1999). TNFSF13B has
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been characterised in a number of fish species, including zebrafish (Liang et al.,
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2010), mefugu (Ai et al., 2011), Japanese sea perch (Cui et al., 2012); grass carp
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(Pandit et al., 2013), yellow grouper (Xiao et al., 2014), miiuy coaker (Meng et al.,
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2015) and tongue sole (Sun & Sun, 2015). Multiple alignment of the predicted S.
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lalandi BAFF amino acid sequence with selected vertebrate sequences identified
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important features (Figure 4) such as a transmembrane domain, a furin protease
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cleavage site, a TNF homology domain and a conserved D-E loop (known as the
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“Flap”), which is unique to TNFSF13B and found in no other TNFSF. Interestingly,
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previous investigations had revealed that many fish species have more than one
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TNFSF13B gene present (Secombes et al., 2016), forming two very distinct groups.
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Phylogenetic analysis shows that the S. lalandi BAFF isolated is more related to the
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represented by a large number of transcripts within the spleen to enable its
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construction. Previous expression studies of this gene within healthy tissues of fish,
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showed consistently that it could be detected within immune relevant tissues, where
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highest expression was in the spleen.
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5. The use of omics in future fish cytokine research
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Currently, there are still a significant number of investigations using microarrays
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to investigate teleost immune gene responses (including cytokine gene expression)
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to disease or changes in their environment using all the gene information that has
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been discovered in a select number of species (Boltaña et al., 2017; Schaeck et al.,
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2017; Diaz et al., 2017; Kaneshige et al., 2016; Ferraresso et al., 2016; Eslamloo et
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al., 2016; Pacitti et al., 2016). Obviously these studies are in those species which
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have had a significant amount of gene discovery already carried out or a genome is
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available, however RNA-seq allows transcriptomics investigations to be carried out in
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any fish species with no previous genetic background. Recent studies, include a
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number of previously well studied species, which includes Atlantic salmon and
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rainbow trout (Morera et al., 2011; Valenzuela-Miranda et al., 2015; 2016; Núñez-
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Acuña et al., 2015; Long et al., 2015; Marancik et al., 2015; Ali et al., 2014), Three-
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spinned stickleback (Brown et al., 2016; Haase et al., 2016a; 2016b; Huang et al.,
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2016), zebrafish (Hartig et al., 2016; Li et al., 2014; Li et al., 2016; Ordas et al., 2010;
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Yang et al., 2012; Yang et al., 2015), Nile Tilapia (Wang et al., 2016a; Zhang et al.,
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2013; Zhu et al., 2015; 2017), Channel catfish (Beck et al., 2012; Li et al., 2012, Liu
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et al., 2013; Peatman et al., 2013; Sun et al., 2012), common carp (Neave et al.,
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2017; Zhang et al., 2011; Zhou et al., 2016), grass carp (Dang et al., 2016), Fugu
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(Cui et al., 2014), European seabass (Sarropoulou et al., 2012); Olive flounder
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(Hwang et al., 2017); turbot (Gao et al., 2016; Robledo et al., 2014; Ronza et al.,
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2016). But its understudied fish, such as the javelin goby (Chen et al., 2016), large
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yellow croaker (Wang et al., 2016b; Zhang et al., 2017a), fathead minnow (Wiseman
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et al., 2013), rare minnow (Wang et al., 2016c), Arctic charr (Norman et al., 2014),
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Gulf killifish (Garcia et al., 2012), bluntsnout bream (Sun et al., 2014), common roach
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(Brinkmann et al., 2016), barramundi (Xia et al., 2013; Hook et al., 2017), European
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eel (Callol et al., 2015), blue catfish (Li et al., 2013); Japanese seabass (Xiang et al.,
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Yellow-fin rockcod (Papetti et al., 2015); emerald rockcod (Gerdol et al., 2015);
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winter flounder (McElroy et al., 2015) where this approach is having a large impact.
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Not only is it allowing the identification of cytokine genes within these species, but it
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is simultaneously allowing accurate measurements of their expression under
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different conditions.
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Interestingly with the development of NGS approaches and RNA-seq, we now
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have a good idea of the cytokine gene repertoire we should expect to find within a
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fish and transcriptomics may only appear to be helping to speed up their
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identification in a newly studied fish species. But this is not entirely true and there
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remains a lot that we can learn using this approach and justifies its use above
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platforms, such as microarray. The information obtained during RNA-seq offers an
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unbiased detection of transcripts, meaning that it can detect novel transcripts,
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duplicated genes, indels (small insertions and deletions), and single nucleotide
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variants (Hegedus et al., 2009; Robinson et al., 2012; Schunter et al., 2014; Wan et
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al., 2015; Wang et al., 2014). Experiments can also be designed to determine the
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types of non-coding transcripts and microRNAs (Desvignes et al., 2014; Jiang et al.,
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2016 ; Pauli et al., 2012; Wang et al., 2016; Yang et al., 2015), present in an
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experiment, which may play a role in cytokine gene expression. These are all things
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that have not been investigated to any great detail in relation to fish cytokine genes
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and will be easy to do now with the available and future technologies. Additionally,
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RNA-seq is much more sensitive than microarray, being able to quantify discrete,
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digital sequencing read counts instead of using gene expression measurements
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limited by background fluorescence at the low end and signal saturation at the high
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end. Lastly, the detection of rare transcripts, single transcripts per cell, or weakly
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expressed genes can easily be targeted, by increasing sequencing coverage depth.
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Each of these advantages will allow a new understanding into the cytokine genes
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expressed under different conditions within a fish species and their regulation, to a
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detail not previously possible.
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6. Conclusion
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Understanding the physiological responses of farmed fish to their surrounding environment
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transcriptomics are areas of molecular biology that are beginning to have a large
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impact on aquaculture research. These techniques can be used to help identify
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immune genes of interest, such as cytokines and their receptors, in any farmed
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species for their use in quantitative expression. Using the latest technology available,
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we have been able to identify a large number of S. lalandi cytokine genes and
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receptors involved in inflammatory and adaptive immune responses. The
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inflammatory response is an important part of innate immunity and pro-inflammatory
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cytokines have been proven especially useful when looking at immune responses of
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fish following pathogenic infection or injury (Covello et al., 2009; Lepen-Pleić et al,
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2013). Understanding adaptive immune responses in fish will be critical for
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successful therapeutic approaches to be developed, such as vaccinations
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(Brudeseth et al., 2013; Zhu et al., 2013) and is an area that still requires a lot of
446
investigation. However, this is beginning to change as new molecular tools and
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approaches are being used and developed, such as transcriptomics and the
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development of monoclonal antibodies (Alnabulsi et al., 2013). This is allowing more
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intricate investigations of a fish’s response to be monitored, and will lead to a greater
450
understanding of how vaccines perform (Gioacchini et al., 2008; von Gersdorff
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Jorgensen et al., 2008; Sarropoulou et al., 2012).
essential
to
solve
production
bottlenecks.
Genomics
and
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is
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With the use of transcriptomic approaches, researchers are able to easily identify
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cytokines and their receptors in species where little of no genetic research has
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previously been carried out, which will allow more informative studies into the
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immune responses of these fish. These studies will aim to; (1) characterise important
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cytokine related genes and identify any novel genes or transcripts present, (2)
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monitor fish health and understand their immune responses to disease, and (3)
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investigate the physiological responses of the fish to changes in their environment to
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identify optimal farming conditions for farming of fish such as S. lalandi. We expect
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this approach to help develop new technology and practises for the aquaculture of
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welfare.
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ACCEPTED MANUSCRIPT 7. References Ahn DH, Kang S, Park H (2016) Transcriptome analysis of immune response genes induced by pathogen agonists in the Antarctic bullhead notothen Notothenia coriiceps. Fish Shellfish Immunol. 55:315-22.
RI PT
Ali A, Rexroad CE, Thorgaard GH, Yao J, Salem M (2014) Characterization of the rainbow trout spleen transcriptome and identification of immune-related genes. Front Genet. 14;5:348. Alnabulsi A, Cash B, Bird S, Secombes CJ (2013) Developing tools to advance our understanding of fish immunity. Fish & Shellfish Immunology 34: 1636-37
SC
Altmann SM, Mellon MT, Distel DL, Kim CH (2003). Molecular and functional analysis of an interferon gene from the zebrafish, Danio rerio. J. Virol. 77: 1992-200.
M AN U
Amemiya CT, Alföldi J, Lee AP, Fan S, Philippe H, Maccallum I, Braasch I, Manousaki T et al. (2013) The African coelacanth genome provides insights into tetrapod evolution. Nature 496: 311-16. Angosto D, Montero J, López-Muñoz A, Alcaraz-Pérez F, Bird S, Sarropoulou E, Abellán E, Meseguer J, Sepulcre MP, Mulero V (2013) Identification and functional characterization of a new IL-1 family member, IL-1Fm2, in most evolutionarily advanced fish. Innate Immunity 20:487-500
TE D
Ao J, Mu Y, Xiang LX, Fan D, Feng M, Zhang S, Shi Q, Zhu LY, Li T, Ding Y, Nie L, Li Q, Dong WR, Jiang L, Sun B, Zhang X, Li M, Zhang HQ, Xie S, Zhu Y, Jiang X, Wang X, Mu P, Chen W, Yue Z, Wang Z, Wang J, Shao JZ, Chen X (2015) Genome sequencing of the perciform fish Larimichthys crocea provides insights into molecular and genetic mechanisms of stress adaptation. PLoS Genet. 11:e1005118.
EP
Aparicio S, Chapman J, Stupka E, Putnam N, Chia J, Dehal P et al., (2002). Wholegenome shotgun assembly and analysis of the genome of Fugu rubripes. Science, 297, 1301-1310. Austin CM, Tan MH, Croft LJ, Hammer MP, Gan HM (2015) Whole Genome Sequencing of the Asian Arowana (Scleropages formosus) Provides Insights into the Evolution of Ray-Finned Fishes. Genome Biol Evol. 7:2885-95.
AC C
464 465 466 467 468 469 470 471 472 473 474 475 476 477 478 479 480 481 482 483 484 485 486 487 488 489 490 491 492 493 494 495 496 497 498 499 500 501 502 503 504 505 506 507 508 509 510 511 512
Balm PH, van Lieshout E, Lokate J, Wendelaar Bonga SE (1995) Bacterial lipopolysaccharide (LPS) and interleukin 1 (IL-1) exert multiple physiological effects in the tilapia Oreochromis mossambicus (Teleostei). J Comp Physiol B. 165:85-92. Barrett, KE (1996) Cytokines: sources, receptors and signalling. Baillière's Clinical Gastroenterology, 10, 1-15. Beck BH, Farmer BD, Straus DL, Li C, Peatman E (2012) Putative roles for a rhamnose binding lectin in Flavobacterium columnare pathogenesis in channel catfish Ictalurus punctatus. Fish Shellfish Immunol. 33:1008-15.
18
ACCEPTED MANUSCRIPT Berthelot C, Brunet F, Chalopin D, Juanchich A, Bernard M, Noël B, Bento P, Da Silva C, Labadie K, Alberti A (2014) The rainbow trout genome provides novel insights into evolution after whole-genome duplication in vertebrates. Nat Commun. 5:3657.
RI PT
Bird S, Zou J, Kono T, Sakai M, Dijkstra JM, Secombes C. (2005a) Characterisation and expression analysis of interleukin 2 (IL-2) and IL-21 homologues in the Japanese pufferfish, Fugu rubripes, following their discovery by synteny. Immunogenetics. 56:909-23. Bird S, Zou J, Savan R, Kono T, Sakai M, Woo J, Secombes C (2005b) Characterisation and expression analysis of an interleukin 6 homologue in the Japanese pufferfish, Fugu rubripes. Dev Comp Immunol. 29:775-89.
SC
Bird S, Tafalla C (2015) Teleost chemokines and their receptors. Biology 4:756-84
M AN U
Bobe J, Nguyen T, Fostier A (2009) Ovarian function of the trout preovulatory ovary: new insights from recent gene expression studies. Comp Biochem Physiol A Mol Integr Physiol. 153:63-8. Boltaña S, Castellana B, Goetz G, Tort L, Teles M, Mulero V, Novoa B, Figueras A, Goetz FW, Gallardo-Escarate C, Planas JV, Mackenzie S (2017) Extending Immunological Profiling in the Gilthead Sea Bream, Sparus aurata, by Enriched cDNA Library Analysis, Microarray Design and Initial Studies upon the Inflammatory Response to PAMPs. Int J Mol Sci. 18: E317.
TE D
Bostock J, McAndrew B, Richards R, Jauncey K, Telfer T, Lorenzen K, Little D, Ross L, Handisyde N, Gatward I, Corner R (2010) Aquaculture: global status and trends Philosophical Transactions of the Royal Society B: Biological Sciences 365: 28972912.
EP
Braasch I, Gehrke AR, Smith JJ, Kawasaki K, Manousaki T, Pasquier J, Amores A, Desvignes T, Batzel P et al., (2016) The spotted gar genome illuminates vertebrate evolution and facilitates human-teleost comparisons. Nat Genet. 48:427-37. Brawand D, Wagner CE, Li YI, Malinsky M, Keller I, Fan S, Simakov O, Ng AY, Lim ZW, Bezault E et al., (2014) The genomic substrate for adaptive radiation in African cichlid fish. Nature. 513:375-81.
AC C
513 514 515 516 517 518 519 520 521 522 523 524 525 526 527 528 529 530 531 532 533 534 535 536 537 538 539 540 541 542 543 544 545 546 547 548 549 550 551 552 553 554 555 556 557 558 559 560 561 562
Brinkmann M, Koglin S, Eisner B, Wiseman S, Hecker M, Eichbaum K, Thalmann B, Buchinger S, Reifferscheid G, Hollert H. (2016) Characterisation of transcriptional responses to dioxins and dioxin-like contaminants in roach (Rutilus rutilus) using whole transcriptome analysis. Sci Total Environ. 541:412-23. Bronte V, Pittet MJ (2013) The spleen in local and systemic regulation of immunity. Immunity. 39:806-18. Brown M, Hablützel P, Friberg IM, Thomason AG, Stewart A, Pachebat JA, Jackson JA (2016) Seasonal immunoregulation in a naturally-occurring vertebrate. BMC Genomics. 17:369. 19
ACCEPTED MANUSCRIPT Brudeseth BE, Wiulsrød R, Fredriksen BN, Lindmo K, Løkling KE, Bordevik M, Steine N, Klevan A, Gravningen K (2013) Status and future perspectives of vaccines for industrialised fin-fish farming. Fish Shellfish Immunol. 35:1759-68.
RI PT
Burger MA, Barnes AC, Adlard RD (2008) Wildlife as reservoirs for parasites infecting commercial species: host specificity and a redescription of Kudoa amamiensis from teleost fish in Australia. Journal of Fish Diseases 31: 835-44. Callol A, Reyes-López FE, Roig FJ, Goetz G, Goetz FW, Amaro C, MacKenzie SA (2015) An Enriched European Eel Transcriptome Sheds Light upon Host-Pathogen Interactions with Vibrio vulnificus. PLoS One. 10:e0133328.
SC
Cawthorn DM, Steinman HA, Witthuhn RC (2012) Evaluation of the 16S and 12S rRNA genes as universal markers for the identification of commercial fish species in South Africa. Gene 491:40-48
M AN U
Cerdà J, Douglas S, Reith M (2010) Genomic resources for flatfish research and their applications. J. Fish Biol. 77: 1045-70. Chaves-Pozo E, Liarte S, Fernández-Alacid L, Abellán E, Meseguer J, Mulero V, García-Ayala A (2008) Pattern of expression of immune-relevant genes in the gonad of a teleost, the gilthead seabream (Sparus aurata L.). Mol Immunol. 45:2998-3011.
TE D
Chen TW, Gan RCR, Wu TH, Huang PJ, Lee CY, Chen YYM, Chen CC, Tang P (2012) FastAnnotator: an efficient transcript annotation web tool. BMC Genomics 13:Suppl 7, S9
EP
Chen QL, Luo Z, Huang C, Pan YX, Wu K (2016) De novo characterization of the liver transcriptome of javelin goby Synechogobius hasta and analysis of its transcriptomic profile following waterborne copper exposure. Fish Physiol Biochem. 42:979-94. Cho HK, Kim J, Moon JY, Nam BH, Kim YO, Kim WJ, Park JY, An CM, Cheong J, Kong HJ (2016) Microarray analysis of gene expression in olive flounder liver infected with viral haemorrhagic septicaemia virus (VHSV). Fish Shellfish Immunol. 49:66-78.
AC C
563 564 565 566 567 568 569 570 571 572 573 574 575 576 577 578 579 580 581 582 583 584 585 586 587 588 589 590 591 592 593 594 595 596 597 598 599 600 601 602 603 604 605 606 607 608 609 610 611 612
Costa V, Angelini C, De Feis I, Ciccodicola A (2010) Uncovering the complexity of transcriptomes with RNA-Seq. Journal of Biomedicine and Biotechnology 2010: 853916. Covello JM, Bird S, Morrison RN, Bridle AR, Battaglene SC, Secombes CJ, Nowak BF (2013) Isolation of RAG-1 and IgM transcripts from the striped trumpeter (Latris lineata), and their expression as markers for development of the adaptive immune response. Fish & Shellfish Immunology 34:778-88. Cui J, Liu S, Zhang B, Wang H, Sun H, Song S, Qiu X, Liu Y, Wang X, Jiang Z, Liu Z (2014) Transciptome analysis of the gill and swimbladder of Takifugu rubripes by RNA-Seq. PLoS One. 9(1):e85505. 20
ACCEPTED MANUSCRIPT Dang Y, Xu X, Shen Y, Hu M, Zhang M, Li L, Lv L, Li J (2016) Transcriptome Analysis of the Innate Immunity-Related Complement System in Spleen Tissue of Ctenopharyngodon idella Infected with Aeromonas hydrophila. PLoS One 11:e0157413.
RI PT
Darawiroj D, Kondo H, Hirono I, Aoki T (2008) Immune-related gene expression profiling of yellowtail (Seriola quinqueradiata) kidney cells stimulated with ConA and LPS using microarray analysis. Fish & Shellfish Immunology 24: 260-6. Davidson WS, Koop BF, Jones SJ, Iturra P, Vidal R, Maass A, Jonassen I, Lien S, Omholt SW (2010) Sequencing the genome of the Atlantic salmon (Salmo salar). Genome Biol. 11:403.
SC
Desvignes T, Beam MJ, Batzel P, Sydes J, Postlethwait JH (2014) Expanding the annotation of zebrafish microRNAs based on small RNA sequencing. Gene 546:3869
M AN U
Dheilly NM, Adema C, Raftos DA, Gourbal B, Grunau C, Du Pasquier L (2014) No more non-model species: the promise of next generation sequencing for comparative immunology. Dev Comp Immunol. 45:56-66.
TE D
Diaz de Cerio O, Bilbao E, Ruiz P, Pardo BG, Martínez P, Cajaraville MP, Cancio I (2017) Hepatic gene transcription profiles in turbot (Scophthalmus maximus) experimentally exposed to heavy fuel oil nº 6 and to styrene. Mar Environ Res. 123:14-24. Dinarello CA (2007) Historical insights into cytokines. Eur J Immunol. 37:S34-45.
EP
Eslamloo K, Xue X, Booman M, Smith NC, Rise ML (2016) Transcriptome profiling of the antiviral immune response in Atlantic cod macrophages. Dev Comp Immunol. 63:187-205. Felsenstein J (1985) Confidence limits on phylogenies: An approach using the bootstrap. Evolution 39: 783–791.
AC C
613 614 615 616 617 618 619 620 621 622 623 624 625 626 627 628 629 630 631 632 633 634 635 636 637 638 639 640 641 642 643 644 645 646 647 648 649 650 651 652 653 654 655 656 657 658 659 660 661
Fernández JA, Bubner EJ, Takeuchi Y, Yoshizaki G, Wang T, Cummins SF, Elizur A (2015) Primordial germ cell migration in the yellowtail kingfish (Seriola lalandi) and identification of stromal cell-derived factor 1. Gen Comp Endocrinol. 213:16-23. Ferraresso S, Bonaldo A, Parma L, Buonocore F, Scapigliati G, Gatta PP, Bargelloni L (2016) Ontogenetic onset of immune-relevant genes in the common sole (Solea solea). Fish Shellfish Immunol. 57:278-92. Forné I, Abián J, Cerdà J (2010) Fish proteome analysis: model organisms and nonsequenced species. Proteomics 10: 858-72. Gao C, Fu Q, Su B, Zhou S, Liu F, Song L, Zhang M, Ren Y, Dong X, Tan F, Li C (2016) Transcriptomic profiling revealed the signatures of intestinal barrier alteration
21
ACCEPTED MANUSCRIPT and pathogen entry in turbot (Scophthalmus maximus) following Vibrio anguillarum challenge. Dev Comp Immunol. 65:159-68. Garcia TI, Shen Y, Crawford D, Oleksiak MF, Whitehead A, Walter RB (2012) RNASeq reveals complex genetic response to Deepwater Horizon oil release in Fundulus grandis. BMC Genomics. 13:474.
RI PT
Gerdol M, Buonocore F, Scapigliati G, Pallavicini A (2015) Analysis and characterization of the head kidney transcriptome from the Antarctic fish Trematomus bernacchii (Teleostea, Notothenioidea): a source for immune relevant genes. Mar Genomics. 20:13-5.
SC
Gertz EM, Yu YK, Agarwala R, Schäffer AA, Altschul SF (2006) Composition-based statistics and translated nucleotide searches: improving the TBLASTN module of BLAST. BMC Biol. 4:41.
M AN U
Gioacchini G, Smith P, Carnevali O (2008) Effects of Ergosan on the expression of cytokine genes in the liver of juvenile rainbow trout (Oncorhynchus mykiss) exposed to enteric red mouth vaccine. Veterinary immunology and immunopathology 123:215-22 Götz S, García-Gómez JM, Terol J, Williams TD, Nagaraj SH, Nueda MJ, Robles M, Talón M, Dopazo J, Conesa A (2008) High-throughput functional annotation and data mining with the Blast2GO suite. Nucleic Acids Res. 36:3420-35.
TE D
Grabherr, M.G., Haas, B.J., Yassour, M., Levin, J.Z., Thompson, D.A., Amit, I., Adiconis, X., Fan, L., Raychowdhury, R., Zeng, Q., et al. (2011). Full-length transcriptome assembly from RNA-Seq data without a reference genome. Nature biotechnology 29:644-652.
EP
Haase D, Rieger JK, Witten A, Stoll M, Bornberg-Bauer E, Kalbe M, Reusch TB (2016a) Immunity comes first: the effect of parasite genotypes on adaptive immunity and immunization in three-spined sticklebacks. Dev Comp Immunol. 54:137-44. Haase D, Rieger JK, Witten A, Stoll M, Bornberg-Bauer E, Kalbe M, SchmidtDrewello A, Scharsack JP, Reusch TB (2016b) Comparative transcriptomics of stickleback immune gene responses upon infection by two helminth parasites, Diplostomum pseudospathaceum and Schistocephalus solidus. Zoology (Jena) 119(4):307-13
AC C
662 663 664 665 666 667 668 669 670 671 672 673 674 675 676 677 678 679 680 681 682 683 684 685 686 687 688 689 690 691 692 693 694 695 696 697 698 699 700 701 702 703 704 705 706 707 708 709 710
Hardie LJ, Laing KJ, Daniels GD, Grabowski PS, Cunningham C, Secombes CJ (1998) Isolation of the first piscine transforming growth factor beta gene: analysis reveals tissue specific expression and a potential regulatory sequence in rainbow trout (Oncorhynchus mykiss). Cytokine 10:555-63. Hartig EI, Zhu S, King BL, Coffman JA (2016) Cortisol-treated zebrafish embryos develop into pro-inflammatory adults with aberrant immune gene regulation. Biol Open. 5:1134-41.
22
ACCEPTED MANUSCRIPT Hegedus Z, Zakrzewska A, Agoston VC, Ordas A, Rácz P, Mink M, Spaink HP, Meijer AH (2009) Deep sequencing of the zebrafish transcriptome response to mycobacterium infection. Mol Immunol. 46:2918-30. Hong S, Li R, Xu Q, Secombes CJ, Wang T (2013) Two types of TNF-α exist in teleost fish: Phylogeny, expression, and bioactivity analysis of type-II TNF-α3 in rainbow trout Oncorhynchus mykiss. J. Immunol. 191:5959-72.
RI PT
Hook SE, Kroon FJ, Metcalfe S, Greenfield PA, Moncuquet P, McGrath A, Smith R, Warne MS, Turner RD, McKeown A, Westcott DA (2017) Global transcriptomic profiling in barramundi (Lates calcarifer) from rivers impacted by differing agricultural land uses. Environ Toxicol Chem. 36:103-112.
SC
Howe K, Clark MD, Torroja CF, Torrance J, Berthelot C, Muffato M, Collins JE, Humphray S et al. (2013) The zebrafish reference genome sequence and its relationship to the human genome. Nature 496: 498-503.
M AN U
Huang Y, Chain FJ, Panchal M, Eizaguirre C, Kalbe M, Lenz TL, Samonte IE, Stoll M, Bornberg-Bauer E, Reusch TB, Milinski M, Feulner PG (2016). Transcriptome profiling of immune tissues reveals habitat-specific gene expression between lake and river sticklebacks. Mol Ecol. 25:943-58. Husain M, Bird S, van Zwieten R, Secombes CJ, Wang T (2012) Cloning of the IL1β3 gene and IL-1β4 pseudogene in salmonids uncovers a second type of IL-1β gene in teleost fish. Devel. Comp. Immunol. 38:431-46.
TE D
Hwang JY, Kwon MG, Jung SH, Park MA, Kim DW, Cho WS, Park JW, Son MH (2017) RNA-Seq transcriptome analysis of the olive flounder (Paralichthys olivaceus) kidney response to vaccination with heat-inactivated viral hemorrhagic septicemia virus. Fish Shellfish Immunol. 62:221-226.
EP
Itoh N (2007) The Fgf families in humans, mice, and zebrafish: their evolutional processes and roles in development, metabolism, and disease. Biol Pharm Bull. 30:1819-25. Jaillon O, Aury JM, Brunet F, Petit JL, Stange-Thomann N, Mauceli E, Bouneau L, et al. (2004) Genome duplication in the teleost fish Tetraodon nigroviridis reveals the early vertebrate proto-karyotype. Nature 431: 946-57.
AC C
711 712 713 714 715 716 717 718 719 720 721 722 723 724 725 726 727 728 729 730 731 732 733 734 735 736 737 738 739 740 741 742 743 744 745 746 747 748 749 750 751 752 753 754 755 756 757 758 759 760
Jang SI, Hardie LJ, Secombes CJ (1994) Effects of transforming growth factor beta 1 on rainbow trout Oncorhynchus mykiss macrophage respiratory burst activity. Dev Comp Immunol. 18:315-23. Jang SI, Hardie LJ, Secombes CJ (1995) Elevation of rainbow trout Oncorhynchus mykiss macrophage respiratory burst activity with macrophage-derived supernatants. J Leukoc Biol. 57:943-7. Jeanmougin F, Thompson JD, Gouy M, Higgins DG, Gibson TJ (1998) Multiple sequence alignment with Clustal X. Trends in Biochemical Science. 23:403-5. 23
ACCEPTED MANUSCRIPT Jiang L, Liu W, Zhu A, Zhang J, Zhou J, Wu C (2016) Transcriptome analysis demonstrate widespread differential expression of long noncoding RNAs involve in Larimichthys crocea immune response. Fish Shellfish Immunol. 51:1-8.
RI PT
Johansen SD, Coucheron DH, Andreassen M, Karlsen BO, Furmanek T, Jørgensen TE, Emblem A, Breines R, Nordeide JT, Moum T, Nederbragt AJ, Stenseth NC, Jakobsen KS (2009) Large-scale sequence analyses of Atlantic cod. Nature Biotechnology 25: 263-71. Jones FC, Grabherr MG, Chan YF, Russell P, Mauceli E, Johnson J, Swofford R, Pirun M et al. (2012) The genomic basis of adaptive evolution in threespine sticklebacks. Nature 484: 55-61
SC
Jørgensen SM, Afanasyev S, Krasnov A (2008) Gene expression analyses in Atlantic salmon challenged with infectious salmon anemia virus reveal differences between individuals with early, intermediate and late mortality. BMC Genomics. 9:179.
M AN U
Kaneshige N, Jirapongpairoj W, Hirono I, Kondo H (2016) Temperature-dependent regulation of gene expression in Japanese flounder Paralichthys olivaceus kidney after Edwardsiella tarda formalin-killed cells. Fish Shellfish Immunol. 59:298-304. Kasahara M, Naruse K, Sasaki S, Nakatani Y, Qu W, Ahsan B, Yamada T, Nagayasu Y et al. (2007) The medaka draft genome and insights into vertebrate genome evolution. Nature 447: 714-19
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Kawanishi M, Kojima A, Ishihara K, Esaki H, Kijima M, Takahashi T, Suzuki S, Tamura Y (2005) Drug resistance and pulsed-field gel electrophoresis patterns of Lactococcus garvieae isolates from cultured Seriola (yellowtail, amberjack and kingfish) in Japan. Letters in Applied Microbiology 40: 322-8.
EP
Kawanishi M, Kijima M, Kojima A, Ishihara K, Esaki H, Yagyu K, Takahashi T, Suzuki S, Tamura Y (2006) Drug resistance and random amplified polymorphic DNA analysis of Photobacterium damselae ssp. piscicida isolates from cultured Seriola (yellowtail, amberjack and kingfish) in Japan. Letters in Applied Microbiology 42: 648-53.
AC C
761 762 763 764 765 766 767 768 769 770 771 772 773 774 775 776 777 778 779 780 781 782 783 784 785 786 787 788 789 790 791 792 793 794 795 796 797 798 799 800 801 802 803 804 805 806 807 808
Kedes L, Liu ET (2010) The Archon Genomics XPRIZE for whole human genome sequencing. Nature Genetics. 42: 917-918. Kobayashi T (2010) Histopathological features of cultured Japanese amberjack (Seriola quinqueradiata) with ascites occurring in the winter. Journal of Toxicologic Pathology 23: 165–169. Laing KJ, Wang T, Zou J, Holland J, Hong S, Bols N, Hirono I, Aoki T, Secombes CJ (2001) Cloning and expression analysis of rainbow trout Oncorhynchus mykiss tumour necrosis factor-alpha. Eur J Biochem 268:1315–1322
24
ACCEPTED MANUSCRIPT Langevin C, Aleksejeva E, Passoni G, Palha N, Levraud JP, Boudinot P (2013) The antiviral innate immune response in fish: evolution and conservation of the IFN system. J Mol Biol. 425:4904-20. Leef MJ, Lee PS (2009) Preliminary investigation into the killing effect of kingfish (Seriola lalandi) serum and mucus against the monogenean parasites Benedenia seriolae and Zeuxapta seriolae. Aquaculture International 17: 607-14.
RI PT
Lepen Pleić I, Secombes CJ, Bird S, Mladineo I (2014) Characterization of three proinflammatory cytokines, TNFα1, TNFα2 and IL-1β, in cage-reared Atlantic bluefin tuna Thunnus thynnus. Fish Shellfish Immunol. 36:98-112.
SC
Li C, Zhang Y, Wang R, Lu J, Nandi S, Mohanty S, Terhune J, Liu Z, Peatman E (2012) RNA-seq analysis of mucosal immune responses reveals signatures of intestinal barrier disruption and pathogen entry following Edwardsiella ictaluri infection in channel catfish, Ictalurus punctatus. Fish Shellfish Immunol. 32:816-27.
M AN U
Li C, Beck BH, Peatman E (2013) Nutritional impacts on gene expression in the surface mucosa of blue catfish (Ictalurus furcatus). Dev Comp Immunol. 44:226-34. Li F, Wang H, Liu J, Lin J, Zeng A, Ai W, Wang X, Dahlgren RA, Wang H. (2016) Immunotoxicity of β-Diketone Antibiotic Mixtures to Zebrafish (Danio rerio) by Transcriptome Analysis. PLoS One. 11(4):e0152530.
TE D
Li Z, Luo H, Li C, Huo X, Yan C, Huang X, Al-Haddawi M, Mathavan S, Gong Z (2014) Transcriptomic analysis of a transgenic zebrafish hepatocellular carcinoma model reveals a prominent role of immune responses in tumour progression and regression. Int J Cancer. 135:1564-73. Lindenmann J, Burke DC, Isaacs A. (1957) Studies on the production, mode of action and properties of interferon. Br J Exp Pathol, 38: 551-562.
EP
Lindlöf A (2003) Gene identification through large-scale EST sequence processing. Appl Bioinformatics. 2:123-9. Liu L, Li C, Su B, Beck BH, Peatman E (2013) Short-term feed deprivation alters immune status of surface mucosa in channel catfish (Ictalurus punctatus). PLoS One. 8:e74581.
AC C
809 810 811 812 813 814 815 816 817 818 819 820 821 822 823 824 825 826 827 828 829 830 831 832 833 834 835 836 837 838 839 840 841 842 843 844 845 846 847 848 849 850 851 852 853 854 855 856 857 858
Liu K, Xu D, Li J, Bian C, Duan J, Zhou Y, Zhang M, You X, You Y, Chen J, Yu H, Xu G, Fang DA, Qiang J, Jiang S, He J, Xu J, Shi Q, Zhang Z, Xu P (2017) Whole genome sequencing of Chinese clearhead icefish, Protosalanx hyalocranius. Gigascience. [Epub ahead of print] Liu Z, Liu S, Yao J, Bao L, Zhang J, Li Y, Jiang C, Sun L, Zhang Y, Zhou T et al. (2016) The channel catfish genome sequence provides insights into the evolution of scale formation in teleosts. Nat Commun. 2016;7:11757. Long M, Zhao J, Li T, Tafalla C, Zhang Q, Wang X, Gong X, Shen Z, Li A (2015) Transcriptomic and proteomic analyses of splenic immune mechanisms of rainbow 25
ACCEPTED MANUSCRIPT trout (Oncorhynchus mykiss) infected by Aeromonas salmonicida subsp. salmonicida. J Proteomics. 122:41-54. Loo GH, Schuller KA (2010) Cloning and functional characterization of a peroxiredoxin 4 from yellowtail kingfish (Seriola lalandi). Comparative Biochemistry and Physiology Part B: Biochemistry and Molecular Biology 156: 244-53.
RI PT
Malmstrøm M, Matschiner M, Tørresen OK, Jakobsen KS, Jentoft S (2017) Whole genome sequencing data and de novo draft assemblies for 66 teleost species. Sci Data. 4:160132.
SC
Mansell B, Powell MD, Ernst I, Nowak BF (2005) Effects of the gill monogenean Zeuxapta seriolae (Meserve, 1938) and treatment with hydrogen peroxide on pathophysiology of kingfish, Seriola lalandi Valenciennes, 1833. Journal of Fish Diseases 28: 253-62.
M AN U
Marancik D, Gao G, Paneru B, Ma H, Hernandez AG, Salem M, Yao J, Palti Y, Wiens GD. (2015) Whole-body transcriptome of selectively bred, resistant-, control-, and susceptible-line rainbow trout following experimental challenge with Flavobacterium psychrophilum. Front Genet. 5:453. Mardis ER (2011) A decade's perspective on DNA sequencing technology. Nature 470: 198-203.
TE D
McElroy AE, Hice LA, Frisk MG, Purcell SL, Phillips NC, Fast MD (2015) Spatial patterns in markers of contaminant exposure, glucose and glycogen metabolism, and immunological response in juvenile winter flounder (Pseudoplueronectes americanus). Comp Biochem Physiol Part D Genomics Proteomics. 2015 Jun;14:5365.
EP
McGaugh SE, Gross JB, Aken B, Blin M, Borowsky R, Chalopin D, Hinaux H, Jeffery WR, Keene A et al., (2014) The cavefish genome reveals candidate genes for eye loss. Nat Commun. 5:5307. Merriman B, Ion Torrent R&D Team, Rothberg JM (2012) Progress in ion torrent semiconductor chip based sequencing. Electrophoresis 33: 3397-417
AC C
859 860 861 862 863 864 865 866 867 868 869 870 871 872 873 874 875 876 877 878 879 880 881 882 883 884 885 886 887 888 889 890 891 892 893 894 895 896 897 898 899 900 901 902 903 904 905 906 907 908
Mommens M, Fernandes JM, Tollefsen KE, Johnston IA, Babiak I (2014) Profiling of the embryonic Atlantic halibut (Hippoglossus hippoglossus L.) transcriptome reveals maternal transcripts as potential markers of embryo quality. BMC Genomics. 15:829. Moore PA, Belvedere O, Orr A, Pieri K, LaFleur DW, Feng P, Soppet D, Charters M, Gentz R, Parmelee D, Li Y, Galperina O, Giri J, Roschke V, Nardelli B, Carrell J, Sosnovtseva S, Greenfield W, Ruben SM, Olsen HS, Fikes J, Hilbert DM (1999) BLyS: member of the tumor necrosis factor family and B lymphocyte stimulator. Science 285, 260–263 Moran D, Gwells RM, Pether SJ (2008) Low stress response exhibited by juvenile yellowtail kingfish (Seriola lalandi Valenciennes) exposed to hypercapnic conditions associated with transportation. Aquaculture Research 39: 1399-1407 26
ACCEPTED MANUSCRIPT Morera D, Roher N, Ribas L, Balasch JC, Doñate C, Callol A, Boltaña S, Roberts S, Goetz G, Goetz FW, MacKenzie SA (2011) RNA-Seq reveals an integrated immune response in nucleated erythrocytes. PLoS One 6:e26998.
RI PT
Mu Y, Ding F, Cui P, Ao J, Hu S, Chen X (2010) Transcriptome and expression profiling analysis revealed changes of multiple signaling pathways involved in immunity in the large yellow croaker during Aeromonas hydrophila infection. BMC Genomics. 11:506. Muncaster M, Kraakman K, Gibbons O, Mensink K, Forlenza M, Jacobson G, Bird, S (2017) Antimicrobial peptides within the Yellowtail Kingfish (Seriola lalandi). Dev. Comp. Immunol. (In press).
SC
Neave MJ, Sunarto A, McColl KA (2017) Transcriptomic analysis of common carp anterior kidney during Cyprinid herpesvirus 3 infection: Immunoglobulin repertoire and homologue functional divergence. Sci Rep. 7:41531.
M AN U
Nocillado JN, Biran J, Lee YY, Levavi-Sivan B, Mechaly AS, Zohar Y, Elizur A (2012) The Kiss2 receptor (Kiss2r) gene in Southern Bluefin Tuna, Thunnus maccoyii and in Yellowtail Kingfish, Seriola lalandi – functional analysis and isolation of transcript variants. Molecular and Cellular Endocrinology 362: 211-220
TE D
Nocillado JN, Zohar Y, Biran J, Levavi-Sivan B, Elizur A (2013) Chronic kisspeptin administration stimulated gonadal development in pre-pubertal male yellowtail kingfish (Seriola lalandi; Perciformes) during the breeding and non-breeding season. Gen Comp Endocrinol. 191:168-76. Norman JD, Ferguson MM, Danzmann RG (2014) An integrated transcriptomic and comparative genomic analysis of differential gene expression in Arctic charr (Salvelinus alpinus) following seawater exposure. J Exp Biol. 217:4029-42.
EP
Núñez-Acuña G, Gonçalves AT, Valenzuela-Muñoz V, Pino-Marambio J, Wadsworth S, Gallardo-Escárate C (2015) Transcriptome immunomodulation of in-feed additives in Atlantic salmon Salmo salar infested with sea lice Caligus rogercresseyi. Fish Shellfish Immunol. 47:450-60.
AC C
909 910 911 912 913 914 915 916 917 918 919 920 921 922 923 924 925 926 927 928 929 930 931 932 933 934 935 936 937 938 939 940 941 942 943 944 945 946 947 948 949 950 951 952 953 954 955 956 957 958
Ordas A, Hegedus Z, Henkel CV, Stockhammer OW, Butler D, Jansen HJ, Racz P, Mink M, Spaink HP, Meijer AH (2010) Deep sequencing of the innate immune transcriptomic response of zebrafish embryos to Salmonella infection. Fish Shellfish Immunol. 31:716-24. Pacitti D, Lawan MM, Feldmann J, Sweetman J, Wang T, Martin SA, Secombes CJ (2016) Impact of selenium supplementation on fish antiviral responses: a whole transcriptomic analysis in rainbow trout (Oncorhynchus mykiss) fed supranutritional levels of Sel-Plex®. BMC Genomics. 17:116. Page RD (1996) TreeView: an application to display phylogenetic trees on personal computers. Computer Applications in the Biosciences 12: 357–358.
27
ACCEPTED MANUSCRIPT Palstra AP, Beltran S, Burgerhout E, Brittijn SA, Magnoni LJ, Henkel CV, Jansen HJ, van den Thillart GE, Spaink HP, Planas JV (2013) Deep RNA sequencing of the skeletal muscle transcriptome in swimming fish. PLoS One 8:e53171.
RI PT
Papetti C, Harms L, Windisch HS, Frickenhaus S, Sandersfeld T, Jürgens J, Koschnick N, Knust R, Pörtner HO, Lucassen M (2015) A first insight into the spleen transcriptome of the notothenioid fish Lepidonotothen nudifrons: Resource description and functional overview. Mar Genomics. 3:237-9. Pauli A, Valen E, Lin MF, Garber M, Vastenhouw NL, Levin JZ, Fan L, Sandelin A, Rinn JL, Regev A, Schier AF (2012) Systematic identification of long noncoding RNAs expressed during zebrafish embryogenesis. Genome Res. 22:577-91.
SC
Peatman E, Li C, Peterson BC, Straus DL, Farmer BD, Beck BH (2013) Basal polarization of the mucosal compartment in Flavobacterium columnare susceptible and resistant channel catfish (Ictalurus punctatus). Mol Immunol. 56:317-27.
M AN U
Petit J, David L, Dirks R, Wiegertjes GF (2017) Genomic and transcriptomic approaches to study immunology in cyprinids: What is next? Dev Comp Immunol. [Epub ahead of print] Pietretti D, Wiegertjes GF (2014) Ligand specificities of Toll-like receptors in fish: indications from infection studies. Dev Comp Immunol. 43:205-22.
TE D
Pooley NJ, Tacchi L, Secombes CJ, Martin SA 2013. Inflammatory responses in primary muscle cell cultures in Atlantic salmon (Salmo salar). BMC Genomics 14:747 Primary Industries and Resources South Australia, 2011. PIRSA Aquaculture Marine Finfish. http://www.pir.sa.gov.au/aquaculture/aquaculture_industry/marine_finfish
EP
Quail MA, Smith M, Coupland P, Otto TD, Harris SR, Connor TR, Bertoni A, Swerdlow HP, Gu Y (2012) A tale of three next generation sequencing platforms: comparison of Ion Torrent, Pacific Biosciences and Illumina MiSeq sequencers. BMC Genomics. 13: 341. Ray D, Melmed S (1997) Pituitary cytokine and growth factor expression and action. Endocr Rev. 18:206-28.
AC C
959 960 961 962 963 964 965 966 967 968 969 970 971 972 973 974 975 976 977 978 979 980 981 982 983 984 985 986 987 988 989 990 991 992 993 994 995 996 997 998 999 1000 1001 1002 1003 1004 1005 1006 1007 1008
Reyes-Becerril M, Ascencio-Valle F, Alamillo E, Hirono I, Kondo H, Jirapongpairoj W, Angulo C (2015) Molecular cloning and comparative responses of Toll-like receptor 22 following ligands stimulation and parasitic infection in yellowtail (Seriola lalandi). Fish Shellfish Immunol. 46:323-33. Reyes-Becerril M, Ascencio-Valle F, Hirono I, Kondo H, Jirapongpairoj W, Esteban MA, Alamillo E, Angulo C (2016) TLR21's agonists in combination with Aeromonas antigens synergistically up-regulate functional TLR21 and cytokine gene expression in yellowtail leucocytes. Dev Comp Immunol. 61:107-15. Robinson N, Sahoo PK, Baranski M, Das Mahapatra K, Saha JN, Das S, Mishra Y, Das P, Barman HK, Eknath AE (2012) Expressed sequences and polymorphisms in 28
ACCEPTED MANUSCRIPT rohu carp (Labeo rohita, Hamilton) revealed by mRNA-seq. Mar Biotechnol (NY). 14:620-33. Robledo D, Ronza P, Harrison PW, Losada AP, Bermúdez R, Pardo BG, Redondo MJ, Sitjà-Bobadilla A, Quiroga MI, Martínez P (2014) RNA-seq analysis reveals significant transcriptome changes in turbot (Scophthalmus maximus) suffering severe enteromyxosis. BMC Genomics. 15:1149.
RI PT
Ronza P, Robledo D, Bermúdez R, Losada AP, Pardo BG, Sitjà-Bobadilla A, Quiroga MI, Martínez P (2016) RNA-seq analysis of early enteromyxosis in turbot (Scophthalmus maximus): new insights into parasite invasion and immune evasion strategies. Int J Parasitol. 46:507-17.
SC
Rothberg JM, Hinz W, Rearick TM, Schultz J, Mileski W, Davey M, Leamon JH, Johnson K et al., (2011) An integrated semiconductor device enabling non-optical genome sequencing. Nature. 475:348-52.
M AN U
Saitou N, Nei M (1987) The neighbor-joining method: a new method for reconstructing phylogenetic trees. Molecular Biology & Evolution 4:406-25. Sangrador-Vegas A, Lennington JB, Smith TJ (2002) Molecular cloning of an IL-8like CXC chemokine and tissue factor in rainbow trout (Oncorhynchus mykiss) by use of suppression subtractive hybridization. Cytokine. 17:66-70.
TE D
Santos MD, Yasuike M, Hirono I, Aoki T (2006) The granulocyte colony-stimulating factors (CSF3s) of fish and chicken. Immunogenetics. 58:422-32. Sarropoulou E, Galindo-Villegas J, Garcia-Alcazar A, Kasapidis P, Mulero V (2012) Characterization of european sea bass transcripts by RNA SEQ after oral vaccine against V. anguillarum. Marine Biotechnology 14: 634-42
EP
Schaeck M, Reyes-López FE, Vallejos-Vidal E, Van Cleemput J, Duchateau L, Van den Broeck W, Tort L, Decostere A (2017) Cellular and transcriptomic response to treatment with the probiotic candidate Vibrio lentus in gnotobiotic sea bass (Dicentrarchus labrax) larvae. Fish Shellfish Immunol. 63:147-156.
AC C
1009 1010 1011 1012 1013 1014 1015 1016 1017 1018 1019 1020 1021 1022 1023 1024 1025 1026 1027 1028 1029 1030 1031 1032 1033 1034 1035 1036 1037 1038 1039 1040 1041 1042 1043 1044 1045 1046 1047 1048 1049 1050 1051 1052 1053 1054 1055 1056 1057 1058
Schartl M, Walter RB, Shen Y, Garcia T, Catchen J, Amores A, Braasch I, Chalopin D, Volff JN, et al. (2013) The genome of the platyfish, Xiphophorus maculatus, provides insights into evolutionary adaptation and several complex traits. Nature Genetics 45: 567–572. Schneider P, MacKay F, Steiner V, Hofmann K, Bodmer JL, Holler N, Ambrose C, Lawton P, Bixler S, Acha-Orbea H, Valmori D, Romero P, Werner-Favre C, Zubler RH, Browning JL, Tschopp J (1999) BAFF, a novel ligand of the tumor necrosis factor family, stimulates B cell growth. J. Exp. Med. 189, 1747–1756. Schunter C, Vollmer SV, Macpherson E, Pascual M (2014) Transcriptome analyses and differential gene expression in a non-model fish species with alternative mating tactics. BMC Genomics. 15:167.
29
ACCEPTED MANUSCRIPT Secombes CJ, Wang T, Bird S (2011) The interleukins of fish. Dev. Comp. Immunol. 35:1336-1345. Secombes CJ, Zou, J, Bird S (2015) Cytokines of Cartilaginous Fish. In: Smith, Sim, Flajnik (Eds) Immunobiology of the Shark., Taylor & Francis Group Ltd. pp. 123-142
RI PT
Secombes CJ, Wang T, Bird S (2016) Vertebrate cytokines and their evolution. In: Malagoli (Ed) The Evolution of the Immune System: Conservation and Diversification. Academic Press. pp. 87-151 Star B, Nederbragt AJ, Jentoft S, Grimholt U, Malmstrøm M, Gregers TF, Rounge TB, Paulsen J, et al. (2011) The genome sequence of Atlantic cod reveals a unique immune system. Nature 477: 207-10.
SC
Stephens FJ, Savage A (2010) Two mortality events in sea-caged yellowtail kingfish Seriola lalandi Valenciennes, 1833 (Nannopercidae) from Western Australia. Australian Veterinary Journal 88: 414-6.
M AN U
Sumathy K, Desai KV, Kondaiah P (1997) Isolation of transforming growth factorbeta2 cDNA from a fish, Cyprinus carpio by RT-PCR. Gene 191:103-7. Sun F, Peatman E, Li C, Liu S, Jiang Y, Zhou Z, Liu Z (2012) Transcriptomic signatures of attachment, NF-κB suppression and IFN stimulation in the catfish gill following columnaris bacterial infection. Dev Comp Immunol. 38:169-80.
TE D
Sun S, Ge X, Xuan F, Zhu J, Yu N (2014) Nitrite-induced hepatotoxicity in Bluntsnout bream (Megalobrama amblycephala): the mechanistic insight from transcriptome to physiology analysis. Environ Toxicol Pharmacol. 37:55-65.
EP
Tacchi L, Bron JE, Taggart JB, Secombes CJ, Bickerdike R, Adler MA, Takle H, Martin SA (2011) Multiple tissue transcriptomic responses to Piscirickettsia salmonis in Atlantic salmon (Salmo salar). Physiological Genomics 43:1241-54. Tacchi L, Secombes CJ, Bickerdike R, Adler MA, Venegas C, Takle H, Martin SA (2012) Transcriptomic and physiological responses to fishmeal substitution with plant proteins in formulated feed in farmed Atlantic salmon (Salmo salar). BMC Genomics 13:363.
AC C
1059 1060 1061 1062 1063 1064 1065 1066 1067 1068 1069 1070 1071 1072 1073 1074 1075 1076 1077 1078 1079 1080 1081 1082 1083 1084 1085 1086 1087 1088 1089 1090 1091 1092 1093 1094 1095 1096 1097 1098 1099 1100 1101 1102 1103 1104 1105 1106 1107 1108
Thakur NL, Jain R, Natalio F, Hamer B, Thakur AN, Müller WE (2008) Marine molecular biology: an emerging field of biological sciences. Biotechnology Advances 26: 233-45. Trumbić Ž, Bekaert M, Taggart JB, Bron JE, Gharbi K, Mladineo I (2015) Development and validation of a mixed-tissue oligonucleotide DNA microarray for Atlantic bluefin tuna, Thunnus thynnus (Linnaeus, 1758). BMC Genomics. 16:1007 Tubbs LA, Poortenaar CW, Sewell MA, Diggles BK (2004) Effects of temperature on fecundity in vitro, egg hatching and reproductive development of Benedenia seriolae and Zeuxapta seriolae (Monogenea) parasitic on yellowtail kingfish, Seriola lalandi. International Journal of Parasitology 35: 315-27. 30
ACCEPTED MANUSCRIPT
RI PT
Valenzuela-Miranda D, Gallardo-Escárate C (2016) Novel insights into the response of Atlantic salmon (Salmo salar) to Piscirickettsia salmonis: Interplay of coding genes and lncRNAs during bacterial infection. Fish Shellfish Immunol. 59:427-438. van Dijk EL, Auger H, Jaszczyszyn Y, Thermes C (2014) Ten years of nextgeneration sequencing technology. Trends Genet. 30:418–26.
SC
Verburg-van Kemenade BM, Weyts FA, Debets R, Flik G (1995) Carp macrophages and neutrophilic granulocytes secrete an interleukin-1-like factor. Dev Comp Immunol. 19:59-70.
M AN U
Vij S, Kuhl H, Kuznetsova IS, Komissarov A, Yurchenko AA, Van Heusden P, Singh S, Thevasagayam NM, Prakki SR, Purushothaman K, Saju JM, Jiang J, Mbandi SK, et al., (2016) Chromosomal-Level Assembly of the Asian Seabass Genome Using Long Sequence Reads and Multi-layered Scaffolding. PLoS Genet. 12:e1005954. Vilček J (2003) The cytokines: an overview. In ‘The Cytokine Handbook’ Thomson AW & Lotze MT, Eds. Elsevier Ltd. pp 3-18.
TE D
von Gersdorff Jorgensen L, Nemli E, Heinecke R, Raida M, Buchmann, K (2008) Immune-relevant genes expressed in rainbow trout following immunisation with a live vaccine against Ichthyophthirius multifiliis. Diseases of Aquatic Organisms 80:189-97. Wan Q, Sua, J (2015) Transcriptome analysis provides insights into the regulatory function of alternative splicing in antiviral immunity in grass carp (Ctenopharyngodon idella). Sci Rep. 5: 12946.
EP
1141 1142 1143 1144 1145 1146 1147 1148 1149 1150 1151 1152 1153 1154 1155 1156 1157
Valenzuela-Miranda D, Boltaña S, Cabrejos ME, Yáñez JM, Gallardo-Escárate C (2015) High-throughput transcriptome analysis of ISAV-infected Atlantic salmon Salmo salar unravels divergent immune responses associated to head-kidney, liver and gills tissues. Fish Shellfish Immunol. 45:367-77.
Wang J, Fu L, Koganti PP, Wang L, Hand JM, Ma H, Yao J (2016d) Identification and Functional Prediction of Large Intergenic Noncoding RNAs (lincRNAs) in Rainbow Trout (Oncorhynchus mykiss). Mar Biotechnol (NY). 18:271-82.
AC C
1109 1110 1111 1112 1113 1114 1115 1116 1117 1118 1119 1120 1121 1122 1123 1124 1125 1126 1127 1128 1129 1130 1131 1132 1133 1134 1135 1136 1137 1138 1139 1140
Wang L, Liu P, Wan ZY, Huang SQ, Wen YF, Lin G, Yue GH (2016a) RNA-Seq revealed the impairment of immune defence of tilapia against the infection of Streptococcus agalactiae with simulated climate warming. Fish Shellfish Immunol. 55:679-89. Wang P, Wang J, Su YQ, Mao Y, Zhang JS, Wu CW, Ke QZ, Han KH, Zheng WQ, Xu ND (2016b) Transcriptome analysis of the Larimichthys crocea liver in response to Cryptocaryon irritans. Fish Shellfish Immunol. 48:1-11. Wang T, Hanington PC, Belosevic M, Secombes CJ (2008) Two macrophage colony-stimulating factor genes exist in fish that differ in gene organization and are differentially expressed. J Immunol. 181:3310-22. 31
ACCEPTED MANUSCRIPT Wang W, Wang J, You F, Ma L, Yang X, Gao J, He Y, Qi J, Yu H, Wang Z, Wang X, Wu Z, Zhang Q (2014) Detection of alternative splice and gene duplication by RNA sequencing in Japanese flounder, Paralichthys olivaceus. G3 4: 2419–2424. Wang Z, Gerstein M, Snyder M (2009). RNA-Seq: a revolutionary tool for transcriptomics. Nat Rev Genet. 10: 57-63.
RI PT
Wang ZJ, Liu XH, Jin L, Pu DY, Huang J, Zhang YG (2016c) Transcriptome profiling analysis of rare minnow (Gobiocypris rarus) gills after waterborne cadmium exposure. Comp Biochem Physiol Part D Genomics Proteomics. 19:120-8.
SC
Ward RD, Holmes BH (2007) An analysis of nucleotide and amino acid variability in the barcode region of cytochrome c oxidase I (COX1) in fishes. Molecular Ecology Notes 7: 899-907
M AN U
Wiseman SB, He Y, Gamal-El Din M, Martin JW, Jones PD, Hecker M, Giesy JP (2013) Transcriptional responses of male fathead minnows exposed to oil sands process-affected water. Comp Biochem Physiol C Toxicol Pharmacol. 157:227-35. Xia JH, Liu P, Liu F, Lin G, Sun F, Tu R, Yue GH (2013) Analysis of stressresponsive transcriptome in the intestine of Asian seabass (Lates calcarifer) using RNA-seq. DNA Res. 20:449-60.
TE D
Xiang LX, He D, Dong WR, Zhang YW, Shao JZ (2010) Deep sequencing-based transcriptome profiling analysis of bacteria-challenged Lateolabrax japonicus reveals insight into the immune-relevant genes in marine fish. BMC Genomics. 11:472. Xu J, Bian C, Chen K, Liu G, Jiang Y, Luo Q, You X, Peng W, Li J, Huang Y, Yi Y, Dong C, Deng H, Zhang S, Zhang H, Shi Q, Xu P (2017) Draft genome of the Northern snakehead, Channa argus. Gigascience. [Epub ahead of print]
EP
Yang D, Liu Q, Yang M, Wu H, Wang Q, Xiao J, Zhang Y. (2012) RNA-seq liver transcriptome analysis reveals an activated MHC-I pathway and an inhibited MHC-II pathway at the early stage of vaccine immunization in zebrafish. BMC Genomics. 13:319.
AC C
1158 1159 1160 1161 1162 1163 1164 1165 1166 1167 1168 1169 1170 1171 1172 1173 1174 1175 1176 1177 1178 1179 1180 1181 1182 1183 1184 1185 1186 1187 1188 1189 1190 1191 1192 1193 1194 1195 1196 1197 1198 1199 1200 1201 1202 1203 1204 1205 1206 1207
Yang L, Irwin DM, He S (2015) Genome-wide identification and characterization of teleost-specific microRNAs within zebrafish. Gene. 561:181-9. Yang S, Marín-Juez R, Meijer AH, Spaink HP (2015) Common and specific downstream signaling targets controlled by Tlr2 and Tlr5 innate immune signaling in zebrafish. BMC Genomics. 16:547. Zhang R, Zhang LL, Ye X, Tian YY, Sun CF, Lu MX, Bai JJ (2013) Transcriptome profiling and digital gene expression analysis of Nile tilapia (Oreochromis niloticus) infected by Streptococcus agalactiae. Mol Biol Rep. 40:5657-68. Zhang X, Mu Y, Mu P, Ao J, Chen X (2017a) Transcriptome Analysis Reveals Comprehensive Insights into the Early Immune Response of Large Yellow Croaker 32
ACCEPTED MANUSCRIPT (Larimichthys crocea) Induced by Trivalent Bacterial Vaccine. PLoS One. 12:e0170958. Zhang X, Wen H, Wang H, Ren Y, Zhao J, Li Y (2017b) RNA-Seq analysis of salinity stress-responsive transcriptome in the liver of spotted sea bass (Lateolabrax maculatus). PLoS One. 12:e0173238.
RI PT
Zhang Y, Stupka E, Henkel CV, Jansen HJ, Spaink HP, Verbeek FJ (2011) Identification of common carp innate immune genes with whole-genome sequencing and RNA-Seq data. J Integr Bioinform. 8:169.
SC
Zhao C, Fu M, Wang C, Jiao Z, Qiu L (2016) RNA-Seq analysis of immune-relevant genes in Lateolabrax japonicus during Vibrio anguillarum infection. Fish Shellfish Immunol. 2016 52:57-64.
M AN U
Zhou Y, Yu W, Zhong H, Li J, Li H, He F, Zhou J, Tang Y, Yu J, Yu F (2016) Transcriptome analysis reveals that insulin is an immunomodulatory hormone in common carp. Fish Shellfish Immunol. 59:213-219. Zhu J, Li C, Ao Q, Tan Y, Luo Y, Guo Y, Lan G, Jiang H, Gan X (2015) Trancriptomic profiling revealed the signatures of acute immune response in tilapia (Oreochromis niloticus) following Streptococcus iniae challenge. Fish Shellfish Immunol. 46:346-53.
TE D
Zhu J, Fu Q, Ao Q, Tan Y, Luo Y, Jiang H, Li C, Gan X (2017) Transcriptomic profiling analysis of tilapia (Oreochromis niloticus) following Streptococcus agalactiae challenge. Fish Shellfish Immunol. 62:202-212. Zhu LY, Nie L, Zhu G, Xiang LX, Shao JZ (2013). Advances in research of fish immune-relevant genes: a comparative overview of innate and adaptive immunity in teleosts. Developmental and Comparative Immunology 39: 39-62.
EP
Zou J, Grabowski PS, Cunningham C, Secombes CJ (1999a) Molecular cloning of interleukin 1beta from rainbow trout Oncorhynchus mykiss reveals no evidence of an ice cut site. Cytokine 11:552-60. Zou J, Cunningham C, Secombes CJ (1999b) The rainbow trout Oncorhynchus mykiss interleukin-1 beta gene has a differ organization to mammals and undergoes incomplete splicing. Eur J Biochem. 259:901-8.
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Zou J, Bird S, Secombes CJ (2010) Antiviral sensing in teleost fish. Curr. Pharm. Des. 16:4185-93. Zou J, Castro R, Tafalla C (2016) Antiviral immunity: Origin and evolution in vertebrates. In: Malagoli (Ed) The Evolution of the Immune System: Conservation and Diversification. Academic Press. pp. 173-193
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ACCEPTED MANUSCRIPT Figure 1: Graph showing the impact that genomics and transcriptomics has had on cytokine discovery in fish over the last 20 years. The number of papers published on fish cytokine genes are shown for each year, with the discovery of a number of the early cytokines highlighted.
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Figure 2: GO enrichment analysis of the translated spleen contigs identified using tBLASTn to search the NCBI non-redundant protein database. Vertical axis represents gene ontology categories, while horizontal axis indicates the number of genes in each ontology category.
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Figure 3: GO enrichment analysis of the translated gonad and pituitary contigs identified using tBLASTn to search the NCBI non-redundant protein database. Vertical axis represents gene ontology categories, while horizontal axis indicates the number of genes in each ontology category.
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Figure 4: Multiple sequence alignment of S. lalandi BAFF with other selected vertebrate BAFF molecules using ClustalX (Jeanmougin et al., 1998). The transmembrane domain (TMD) is in bold and potential furin cleavage sites are boxed. Conserved cysteines are labelled with an arrow. The conserved TNF homology domain (THD) is highlighted in grey, with the conserved long D-E loop, known as the “Flap” highlighted in yellow. Accession numbers for BAFF are: human, Q9Y275.1; Grass carp, AGG11791.1; Zebrafish, NP_001107062.1; Mefugu, AEB69781.1; Trout, NP_001118036.1; Japanese sea perch, AEH22106.1; Chicken, NP_989658.1; Miiuy croaker, AHL44989.1; Yellow grouper, AFN70720.1; Tongue sole, XP_008326904.1; Yellowtail kingfish, ????.
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Figure 5: Phylogenetic analysis of S. lalandi BAFF (highlighted with a red box) with selected teleost, reptile, bird, amphibian and cartilaginous fish BAFF (TNFSF13B) and APRIL (TNFSF13) amino acid sequences. Accession numbers of each sequence are included in the figure. A different colour is used to indicate the clear clustering of sequences into separate groups. Analysis was performed using the neighbour-joining method (Saitou & Nei, 1987), the tree drawn using the TreeView program v1.6.1 (Page, 1996) and confidence limits added (Felsenstein, 1985).
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Supplementary Figure 1: Graph showing the distribution of the 3,147,845 contigs obtained from the spleen RNA-Seq run. The contig lengths sequenced and their frequency are plotted. Supplementary Figure 2: Graph showing the distribution of the 3,646,264 contigs obtained from the spleen RNA-Seq run. The contig lengths sequenced and their frequency are plotted.
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Table 1: Seriola lalandi cytokine and receptor genes obtained using RNA-Seq
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CXCL9 CXCL10 CXCL14 CCL3 CCL18 CCL19 CCL20 CCL25
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Chemokine Family
Accession No.
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Ligand IL-2 IL-4 IL-6 IL-7 IL-10 IL-11 IL-12 p35 IL-12 p40 IL-15 IL-16 IL-18 IL-34 EBI3
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Interleukin (IL) Family
Receptor IL-1Racp IL-1R Type I IL-1R Type II IL-2Rb IL-4Ra IL-5Ra IL-6Rb IL-7Ra IL-10Ra IL-12Rb2 IL-17RA IL-17RB IL-17RE IL-18Racp IL-20Ra IL-21R IL-31Ra γc (CD132) beta-c Glycoprotein 130 CXCR1 CXCR3 CXCR4 CXCR5 CCR1 CCR2 CCR3 CCR4 CCR7 CCR9 XCR1
Accession No.
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Table 1: Seriola lalandi cytokine and receptor genes obtained using RNA-Seq
TNF-a1 TNF-a2 TNF-b2 TNFSF6 TNFSF12 TNFSF13B TNFSF14
Interferon (IFN) Family
IFN-g
Fibroblast growth factor (FGF) Family
FGF-2 FGF-7
Colony stimulating factor (CSF) Family
M-CSF
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Transforming growth factor (TGF) Family
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Tumour necrosis factor (TNF) Family
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TGF-b
Leukemia inhibitory factor (LIF) Monocyte chemotactic protein (MCP) MCP1 Macrophage migration inhibitory factor (MMIF) MMIF Ciliary neurotrophic factor (CNF)
TNFRSF1A TNFRSF1B TNFRSF2 TNFRSF3 TNFRSF5 TNFRSF6 TNFRSF9 TNFRSF10B TNFRSF13B TNFRSF14 TNFRSF16 TNFRSF19 TNFRSF21 TNFRSF26 IFN-gR IFN-a/bR2 FGF-R1
M-CSFR G-CSFR TGF-bR3 LIFR
CNFR
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Figure 1
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Figure 4 TMD MDDSTEREQSRLTSCLKKREEMKLKECVSILPRKESPSVRSSKDGKLLAATLLLA-LLSCCLTVVSFY ---------MKSVDCVH---VIQQKDTASSPSG--PPGAASGTTGLFSVTFLWLAMLLSSCLAAVSLY ------------------------MAAEDVGPSRGER---RRL--PWLFLVLVVAAITSSSLSVISLY ------------------------MPAEDVGPGRGER---RRL--SWLFLVLVAAAITSSSLSVISLY -------------------MASAGPNPEGGRPASRQESGGRRL--SWLVLLLTLAAVTSSSLSALSLY -----------------------MAVLAGAKPGTGQRAGEGRL--SWPVFLLTLAAVTSSSLSALSLY -----------------------MAALAGFESGTGPRTGERRL--SWPVFLLTLVAVTSSSLSALSLY -------------------MVPPMAVSAGGKSLAKQRTGKVRSSWFSPVVLLTLAAVTSSSLSALSLY -------------------MGP---VRVGLEAGSGQRAGEGSP--SWPVVLLTLVAISSSFLSAVSLY -------------------MGPAMAALAGVEAGTGQRAGEGRL--SWPVFLLTLAAVTSSSFSALSLY -------------------MGPAMAVLAGVKPGTGQQAGGGRL--SWPVFLLTLAAVTCSSLSALSVY . . * . : .. ::.:*.*
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Human Chicken Grass_carp Zebrafish Trout Japanese_sea_perch Yellow_grouper Tounge_sole Mefugu Miiuy_croaker Yellowtail_kingfish
QVAALQGDLASLRAELQG-----HHAEKLPAGAGAPKAG------LEEAPAVTAGLKIFEPPAPG HAITLKTELEALRSELIYRVRARSPLEQPPVSPGDKKAGASVSSFLQVSAAGARQENRLPGPSPA HVLALQAEVEGLRAEVVR-----KREEQSGMLEEPMNGA-------EKLTHQQEHEEIKDRIEYL HVLALKAEVEGLRAEVAR-----KREEQSGMLDEPVNEA-------EKQTHR---EEGGDSIEYL HLLALRAEVEELRSEVFR-----RREEQQ-EARHGETLQ-------QMSSRARRSSPDHP----QLMALRAEVEGLKSEVCR-----RREEGQ-DAKQASQTE-------NIGSRRNIQEPL------QLVALRAEVEGLKSEVCR-----RREEGQ-EAKHARQTE-------NIS-RRSSQEPL------QLMALRAEVEGLRSEVGR-----RREEGEFKIKCESQSE-------SINLRR------------QLLALRAEVDALRSEVGR-----TREYGQ-RAQHASQMA-------NVSSWRSSQEVRG-----QLMALRAEVEGLKSEVVR-----RREESQ-EVKHASQAD-------NMSSRRSIHEPL------QLVALRAEVEGLKSEVCR-----RREEGQ-EAKHGGQTE-------NISSRRSNQEPL------: :*: :: *::*: .
Human Chicken Grass_carp Zebrafish Trout Japanese_sea_perch Yellow_grouper Tounge_sole Mefugu Miiuy_croaker Yellowtail_kingfish
EGNSSQ-------NSRNKRAVQGPEETVTQDCLQLIADSETPTIQKGS----YTFVPWLLSFKRG ESFQTEI--WDRNRNRGRRSIVNAEETVLQACLQLIADSKSDIQQKDD----SSIVPWLLSFKRG QQTEMDDTTTDRVAVSKRSLGHVSNKAESQACLQMMADNRKKTFQKEFALELCTAIPWHVGLKRG HQADMD-ITTDPSVMSKRSLSHAPNKAEPQPCLQMMADNKKKTFQKEFAFDYCTAIPWQVGLKRG HPPDPQ---PGLSFVRKRSVGTGTENSVSQPCLQMLADSNRKTFQKEFALEPYTGIPWQAGLRRG HQPESQ---HALPLIRKRRQVSVTEALVSQPCLQLLANTTRKLFRKDISSMPHIGIPWQAGLRRG HQPGTQ---DALTLTRNRRLVSGSETLVSQPCLQLLANSSRKTFRKEYKSEPHTGIPWQAGLRRG HQAEAQ---HAFSLIRKRRMVSEPQTIVSQSCLQLLANDKRETYRKEFDLEPHTGIPWQTGLERG RRPGSP---HAFLSLRRQKRLAGTDTLVSQPCLQMLANSSRTTFRKELTSGPHTGIPWKSGLRRG HQPGSP---HAFALIKKRSLASDSEASVSQPCLQLLANSSRKTFGKEFESGPHTGIPWQSGLKRG QQPESQ---HASTLLRKRRVVAGADTLVSQPCLQLLANRKRTTFSKELESTLYTGIPWQTGLRRG . : .: * ***::*: * :** .:.**
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D-E loop SALEEKENKILVKETGYFFIY-----GQVLYTDKTYAMGHLIQRKKVHVFGDELSLVTLFRCIQN TALEEQGNKIVIKETGYFFIY-----GQVLYTDTTFAMGHLIQRKKAHVFGDDLSLVTLFRCIQN SALEEEQGTILIKEEGFFFIY-----SQVYYTDSTFAMGHIVIRIKKNVVGDESQHVVLFRCIQS SALEEEQGTILVKEEGFFFIYSQLSFSQVYYTDPTFAMGHIVIRIKKNVVGNESQHVVLFRCIQS SALEAESDSILVREEGYYFVY-----SQVYYMDTTFAMGHVVIRKKRNVVGDEAQHVTLFRCIQN SALELFRDRILVNEEGYYFVY-----SQVYYMDSTFAMGHVVIRWKKNVVGDEPQYVFLFRCIQN SALEADRDCMLVREEGFYFVY-----GQVYYIDSTFAMGHVVIRRKRNVVGDEPQSVILFRCIQN SSLKQDGDTMVVQEEGFYFVY-----SQVYYMDRTFAMGHVVIRRKRNVVGDEPQFVVLFRCIQS SALEADGDSILVGEEGFYFVY-----SQVYYMDSIFAMGHVVIRRKRTVVGDETPEVILFRCIQN SALEPDGDSILVREEGFYFVY-----SQIYYMDSTFAMGHVVIRRKRNVVGDEDPCVILFRCIQS SALEAEGDRILVRQEGFFFVY-----SQVFYMDSTFAMGHVVIRWKSNVVGNDDPFAVLFRCIQS ::*: . ::: : *::*:* .*: * * :****:: * * *.*:: . ******.
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Human Chicken Grass_carp Zebrafish Trout Japanese_sea_perch Yellow_grouper Tounge_sole Mefugu Miiuy_croaker Yellowtail_kingfish
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Human Chicken Grass_carp Zebrafish Trout Japanese_sea_perch Yellow_grouper Tounge_sole Mefugu Miiuy_croaker Yellowtail_kingfish
Human Chicken Grass_carp Zebrafish Trout Japanese_sea_perch Yellow_grouper Tounge_sole Mefugu Miiuy_croaker Yellowtail_kingfish
MPETLPNNSCYSAGIAKLEEGDELQLAIPRENAQISLDGDVTFFGALKLL MPQSYPNNSCYTAGIAKLEEGDELQLTIPRRRAKISLDGDGTFFGAVRLL MNRVNHFNTCYTGGVVKLDSGDRLELLIPRTHANISLDGDSTFLGAIKLA MNRVNHYNTCYTGGVVKLDSGDKLDLLIPRANANISLEGDATFLGAIKLA MNPVYPYNTCYTGGIVKLEVGDSVELLIPRSTAKVSLDGDSTFLGAVRLA MNTDHPYNTCYTGGIVKLELGDHLELLIPRSTANVSLDGDSTFLGAVRLG MNPVHPYNTCFTGGIVKLEAGDHLELLIPRSTANVSLYGDATFLGAVKLA MNDTHPYNTCYTGGVVKLEVGDHLELLIPRSTANVSLDGDATFMGAFKLV MNPVYPFNTCYTGGIVKLKRGDHLELLIPRSTASVSLDEDSTFLGAIKLG MNPVYPYNTCYTGGIVKLEAGDHLELLIPRSTANVSLDGDATFLGAVKLA MNPVFPFNTCYTGGIVKLEVGDHLELLIPPFTANVSLDGDVTFLGAVKLA * *:*::.*:.**. ** ::* ** *.:** * **:**.:*
Figure 5
ACCEPTED MANUSCRIPT NORTHERN PIKE TNFSF13 XP 010868771.1 SALMON TNFSF13 NP 001135076.1 TROUT TNFSF13 NP 001118143.1 ATLANTIC HERRING TNFSF13 XP 012690225.1 GRASS CARP TNFSF13 AGO01886.1
TNFSF13
ZEBRAFISH TNFSF13 NP 001161936.1 XENOPUS TNFSF13 XP 004919464.1 73
HUMAN TNFSF13B BAE16556.1 GREEN ANOLE TNFSF13 XP 008120421.1
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AFRICAN LUNGFISH TNFSF13 AKL90490.1 COELACANTH TNF13 XP 005999827.1 COELACANTH TNF13B.2 XP 005997217.1
SPOTTED GAR TNFSF13B.2 XP 006632891.1 TROUT TNFSF13B.2 ABC84584.1
FUGU TNFSF13B.2 XP 003970507.1
AMAZON MOLLY TNFSF13B.2 XP 007548545.1
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GUPPY TNFSF13B.2 XP 008418124.1
PLATYFISH TNFSF13B.2 XP 005795434.1 TILAPIA TNFSF13B.2 XP 003445926.1
Group II TNFSF13B
STICKLEBACK TNFSF13B.2 AAY27077.1 BICOLOR DAMSELFISH TNFSF13B.2 XP 008279538.1
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YELLOW CROAKER TNFSF13B.2 XP 010734493.1 MIIUY CROAKER TNFSF13B.2 AHL44990.1 BAMBOO SHARK TNFSF13B.2 ADZ54859.1 ELEPHANT SHARK TNFSF13B.2 JK934351.1 AFRICAN LUNGFISH TNFSF13B AKL90491.1
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COELACANTH TNF13B.1 XP 005997065.1 ELEPHANT SHARK TNFSF13B.1 SCAFF29
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SPOTTED CATSHARK TNFSF13B.1 CDG23444.1 SPINY DOGFISH TNFSF13B.1 CCD04084.1 XENOPUS TNFSF13B XP 004912429.1
HUMAN TNFSF13B AAH20674.1
51
CHICKEN TNFSF13B AAM90951.2 GREEN ANOLE TNFSF13B XP 003215395.2
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SPOTTED GAR TNFSF13B.1 XP 006639318.1 SMELT TNFSF13B.1 ACO08866.1
MEXICAN TETRA TNFSF13B.1 XP 007228002.1
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ZEBRAFISH TNFSF13B.1 NP 001107062.1
73
53
GOLDFISH TNFSF13B.1 AEG47359.1 GRASS CARP TNFSF13B.1 AGG11791.1 TROUT TNFSF13B.1B CDQ92381.1
SALMON TNFSF13B.1 NP 001135232.1 TROUT TNFSF13B.1A NP 001118036.1 TONGUE SOLE TNFSF13B.1 XP 008326904.1 TILAPIA TNFSF13B.1 NP 001276352.1
47
MEDAKA TNFSF13B.1 XP 011488237.1 MUMMICHOG TNFSF13B.1 XP 012730561.1 SOUTHERN PLATYFISH TNFSF13B.1 XP 005798137.1
31
69
GUPPY TNFSF13B.1 XP 008399683.1 AMAZON MOLLY TNFSF13B.1 XP 007555419.1
BICOLOR DAMSELFISH TNFSF13B.1 XP 008297202.1 YELLOWTAIL KINGFISH TNFSF13B.1 12 10
0.1
BLACK ROCKCOD TNFSF13B.1 XP 010789927.1 14 23
YELLOW GROUPER TNFSF13B.1 AFN70720.1 MIIUY CROAKER TNFSF13B.1 AHL44989.1 FUGU TNFSF13B.1 XP 003961760.1 JAPANESE SEA BASS TNFSF13B.1 AEH22106.1
Group I TNFSF13B
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Figure 2
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S0145-305X(17)30190-8
DOI:
10.1016/j.dci.2017.04.001
Reference:
DCI 2862
To appear in:
Developmental and Comparative Immunology
Received Date: 9 March 2017 Revised Date:
1 April 2017
Accepted Date: 1 April 2017
Please cite this article as: Jacobson, G., Muncaster, S., Mensink, K., Forlenza, M., Elliot, N., Broomfield, G., Signal, B., Bird, S., Omics and cytokine discovery in fish: Presenting the yellowtail kingfish (Seriola lalandi) as a case study, Developmental and Comparative Immunology (2017), doi: 10.1016/ j.dci.2017.04.001. This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
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Omics and cytokine discovery in fish: Presenting the Yellowtail kingfish (Seriola
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lalandi) as a case study.
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Gregory Jacobson1, Simon Muncaster2, Koen Mensink3, Maria Forlenza3, Nick Elliot1,
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Grant Broomfield1, Beth Signal1, Steve Bird1
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Engineering, University of Waikato, Private Bag 3105, Hamilton 3240 New Zealand
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Molecular Genetics, Department of Biological Sciences, School of Science and
School Applied Science, Bay of Plenty Polytechnic, 70 Windermere Dr, Poike,
Tauranga 3112 New Zealand
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University, Wageningen, The Netherlands
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Cell Biology and Immunology Group, Department of Animal Sciences, Wageningen
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Corresponding author:
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Email: [email protected], Tel: (+64) 07 838 4723
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Highlights
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• The ability to sequence genomes and transcriptomes has accelerated cytokine discovery within fish, allowing informative studies to be carried out in organisms where no genetic information existed previously.
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• Undertaking RNA-Seq on selected tissues of S. lalandi has identified a large number of cytokine and receptor genes within this species, which have been deposited within genbank.
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• The cytokine and receptor genes discovered using transcriptomic approaches can be used in future studies, to allow the effective monitoring of fish immunity with changes to its environment or in response to disease.
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Abstract A continued programme of research is essential to overcome production
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bottlenecks in any aquacultured fish species. Since the introduction of genetic and
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molecular techniques, the quality of immune research undertaken in fish has greatly
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improved. Thousands of species specific cytokine genes have been discovered,
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which can be used to conduct more sensitive studies to understand how fish
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physiology is affected by aquaculture environments or disease. Newly available
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transcriptomic technologies, make it increasingly easier to study the immunogenetics
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of farmed species for which little data exists. This paper reviews how the application
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of transcriptomic procedures such as RNA Sequencing (RNA-Seq) can advance fish
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research. As a case study, we present some preliminary findings using RNA-Seq to
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identify cytokine related genes in Seriola lalandi. These will allow in-depth
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investigations to understand the immune responses of these fish in response to
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environmental change or disease and help in the development of therapeutic
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approaches.
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1. Introduction
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Both innate and adaptive immune responses utilise regulatory proteins called
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cytokines, which are groups of small proteins (~5–20 kDa) important in cell signalling
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and modulation of an organism’s immunity. Cytokines are produced by a broad
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range of cells and each cytokine can be produced by more than one cell type, where
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they can enhance or inhibit the action of other cytokines in complex ways. They are
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important in health and disease and are important in immune responses to bacterial,
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viral, or parasitic pathogens (Barrett, 1996; Vilček, 2003). Cells of the immune
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system (macrophages, granulocytes, dendritic cells, B-cells, T-cells and mast cells)
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and endothelial cells, fibroblasts and various stromal cells secrete cytokines that bind
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to specific cellular receptors through autocrine or paracrine mechanisms. They effect
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the behaviour of the cells around them, helping to modulate the balance between
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humoral and cell-based immune responses and the maturation, growth, and
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responsiveness of particular cell populations. In mammals, there are over 100
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separate genes coding for cytokine-like activities, many with overlapping functions
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and many still needing more study. Today, the term “cytokine” encompasses
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interferons (IFN’s), the interleukins (IL’s), the tumour necrosis factor family (TNF’s),
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the transforming growth factor family (TGF’s), the colony stimulating factor family
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(CSF’s) and the chemokine family.
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The activity of cytokines have been known of and studied in mammals since 1957,
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where interferon-alpha (IFN-α) was fist investigated (Lindenmann et al., 1957).
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However, it was not until the early 1980’s that the molecular cloning of the first
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cytokines (IFNα, IL-1, IL-2 and TNFα) was achieved (Dinarello, 2007). Studies into
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the regulation of fish immunity, began in the early 1990’s (Balm et al., 1995; Jang et
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al., 1994; 1995; Verburg-van Kemenade et al., 1995) due to the importance of fish
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within the aquaculture industry and the need to understand the regulation of
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immunity, due to problems with disease. The first cytokine to be cloned in fish was
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TGF-β (Sumathy et al., 1997; Hardie et al., 1998), followed closely by IL-1β (Zou et
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al., 1999a; 1999b) and a few years later by TNF-α (Laing et al., 2001). Both of these
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were discovered using a homology-based cloning approach, where known
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particularly conserved sequence. These regions were then used to design
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degenerate oligonucleotides to enable amplification of the gene from a targeted fish
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species. This approach was limited and only worked well when the genes being
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targeted had not diverged too far during evolution. However, additional approaches
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that became available shortly after such as subtractive hybridization (Sangrador-
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Vegas et al., 2001) and expressed sequence tags (Altmann et al., 2003) also
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contributed to the discovery of cytokines in fish, helping to identify homologues of IL-
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8 and IFN-α.
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These early approaches of cytokine discovery eventually fell out of favour with the availability of expressed sequence tag (EST) sequence processing (Lindlöf, 2003)
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and the subsequent ability to sequence genomes which has become easier through
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the development of next generation sequencing (NGS) technology (van Dijk et al.,
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2014). One of the first fish genomes sequenced was from Takifugu rubripes
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(Aparicio et al., 2002) and opened up the ability to search for cytokines in fish that
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shared very little homology to their mammalian homologues. IL-2, IL-21 and IL-6
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were some of the first cytokines to be discovered (Bird et al., 2005a; 2005b), where
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conservation of synteny was found between the mammalian and fish genomes,
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allowing their discovery. More recently, the development of technologies such as
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RNA-Seq and the ability to sequence transcriptomes has transformed immune gene
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discovery and provides the ability to profile immune gene expression in organisms
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where no gene discovery had previously taken place (Dheilly et al., 2014).
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The availability of genomes and transciptomics has revolutionised cytokine
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discovery and research in fish (Figure 1). This review will look at how the application
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of new genetic technologies is aiding fish cytokine discovery and research, will use
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the Yellowtail kingfish (Seriola lalandi) as a case study to demonstrate the power of
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RNA-Seq when studying a new fish species where no cytokine genes have been
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characterised and finish by looking at the types of investigations into the roles of
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cytokines in fish immunity that are now possible in fish.
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Having information at the genetic level available within a fish species, allows
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researchers to carry out more precise investigations into the physiology of an
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organism. Recent advances within the areas of genomics and transcriptomics are
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making these types of studies more accessible to fish research, in species where no
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genetic information has been collected (Dheilly et al., 2014). Currently, there are a
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select number of fish species, where a large amount of genetic data has been
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accumulated, however this has been the result of many years of research, where the
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identification of genes took a significant amount of time (Zou et al., 2010; Secombes
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et al., 2011; Bird et al., 2015; Secombes et al., 2015; 2016). However, new
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sequencing technologies, capable of sequencing thousands to hundreds of millions
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of short DNA strands (not specifically targeting a single sequence) without a prior
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bacterial cloning and plasmid DNA isolation step, have enabled progress in the field
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of gene identification to be accelerated greatly. This began in 2005 with the invention
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of ‘massively parallel’ sequencing technologies (Mardis, 2011), with technological
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progress initially driven by projects such as the Human Genome Project, HapMap
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Project and ENCODE.
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Since the emergence of these next-generation sequencing platforms, new
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technology continues to develop at a fast pace, that improves on previous
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methodology or a new approach is developed, enabling increases in sequencing
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efficiency and lower costs. There is fierce competition between companies
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developing this technology and incentives, such as the Archon Genomics XPRIZE,
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which had a goal of sequencing 100 human genomes in 30 days for less than
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USD$1000 per genome (Kedes & Liu, 2010), have helped to push this forward. This
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progress has allowed significant advances in the area of genomics and has led to
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genome sequencing being achievable in a wider range of organisms of scientific or
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commercial interest, which includes a number of fish species. Currently, there are
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twelve fish with their entire genomes sequenced that are being assembled and
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annotated and are publically available , that includes Japanese Pufferfish, Takifugu
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rubripes (Aparicio et al., 2002), Green Spotted Puffer, Tetradon nigoviridis (Jaillon et
150
al., 2004), Japanese rice fish, Oryzias latipes (Kasahara et al., 2007), Atlantic Cod,
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aculeatus (Jones et al., 2012), Zebrafish, Danio rerio (Howe et al., 2013), African
153
coelacanth, Latimeria chalumnae (Amemiya et al., 2013), Southern platyfish,
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Xiphophorus maculatus (Schartl et al., 2013), Mexican cavefish, Astyanax
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mexicanus (McGaugh et al., 2014), Nile tilapia, Oreochromis niloticus (Brawand et
156
al., 2014), spotted gar, Lepisosteus oculatus (Braasch et al., 2016), Amazon molly,
157
Poecilia
158
number of species for which the genome has or is been sequenced, but is still in
159
draft form or incomplete. This includes fish such as Atlantic Salmon (Davidson et al.,
160
2010), rainbow trout (Berthelot et al., 2016), channel catfish (Liu et al, 2016), large
161
yellow croaker (Ao et al., 2015), Northern snakehead (Xu et al., 2017), Chinese
162
clearhead icefish (Liu et al., 2017), common carp (Petit et al., 2017), Asian arowana
163
(Austin et al., 2015), Asian seabass (Vij et al., 2016) and another 66 teleost species
164
(Malmstrøm et al., 2017)
(http://www.ensembl.org/Poecilia_formosa/Info/Index)
with
a
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Transcriptomics is also an area of molecular biology that is beginning to have a
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large impact on fish cytokine research. Unlike the genome which is static, the
168
transcriptome of a cell is dynamic and is continually changing, due to the differences
169
in gene expression and the levels at which they are expressed. Techniques in this
170
area are less targeted and allow the study of 1000’s of genes simultaneously,
171
encoded by the genome from a specific cell, tissue or organism at a specific time or
172
under a specific set of conditions. Early studies used DNA microarrays, which is a
173
large collection of identified microscopic DNA spots attached to a solid surface, that
174
have initially been isolated from the species of interest. These spots will hybridize to
175
genes that are expressed in the sample and are detected using fluorescence,
176
allowing the relative abundance of nucleic acid sequences in the sample to be
177
determined. This approach has been very informative when investigating an
178
immunological response of a fish and how they are affected by changes to their
179
environment or disease (Tacchi et al., 2011; Tacchi et al., 2012; Pooley et al., 2013;
180
Mommens et al., 2014; Trumbic et al., 2015; Cho et al., 2016) and has led to the
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understanding of actual biological pathways in the organism, rather than just
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individual gene expression. However, although a powerful approach, it has been
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limited by the lack of genetic data available in your species of interest beforehand to
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produce the microarray and it is quite a costly and technically difficult approach to
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use.
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The advances being made by NGS have had a dramatic impact on the area of
188
transcriptomics and there are newly available technologies, such as RNA
189
sequencing (RNA-Seq), that are able to directly sequence the majority of RNA from
190
a cell or tissue sample to study all the genes being expressed at a particular point in
191
time (Wang et al., 2009; Costa, 2010). The advantage of this is it does not rely on or
192
is limited by the previous accumulation of genetic data and there is no requirement of
193
knowing the identity of all the transcripts to be analysed prior to analysis. Along with
194
cytokine discovery, this method is extremely useful for quantifying levels of gene
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expression in a sample, and can be used for comparative transcriptomics, having the
196
potential to provide much more information, including isoform identification, rare
197
transcript detection, and SNP detection than previous methods such as microarrays.
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RNA-Seq are newly developed and available technologies that have begun to be
199
used for transcriptomic studies in a number of fish species, such as trout (Palstra et
200
al., 2013), zebrafish (Hegedus et al., 2009; Ordas et al., 2011), Japanese sea bass
201
(Xiang et al., 2010) and the small yellow croaker (Mu et al., 2010). The next section
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will focus on how a transcriptomics approach can be applied to a relatively
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understudied teleost species without a genome available to aid in cytokine discovery.
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3. Yellowtail kingfish (Seriola lalandi) transcriptomic approach
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3.1 Current state of kingfish aquaculture
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Over the last 15 years, there has been acquisition of genetic information in a
210
variety of farmed fish species, allowing the implementation of new genetic
211
technologies within aquaculture (Bostock et al., 2010; Cerdà et al., 2010; Forné et al.,
212
2010; Johansen et al., 2009; Thakur et al., 2008). Before this type of information was
213
available, the monitoring of fish immune responses to changes in their environments
214
was very arbitrary and normally measured by physiological parameters or fish
215
mortalities. With the development of genetic approaches, more focussed studies can
216
now be achieved, which have allowed researchers to look in more detail and attempt
217
to solve some of the important bottlenecks there are in successful production of a
218
particular fish species. In some cases, findings from this research, have then been
219
passed down to the fish farmers themselves, who can translate them into good
220
farming practice.
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S. lalandi are widely distributed throughout the warm–temperate waters of the
223
southern hemisphere. In the wild they reach 1.7 m in length and weigh 56 kg and are
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a highly prized game fish, having excellent flesh quality for a range of product
225
options (such as whole fillets, sushi and the highly valued sashimi), giving it
226
significant domestic and international market opportunities. Successful aquaculture
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of this species in New Zealand would provide a reliable and controlled production of
228
kingfish to an ever growing market. Culture of S. lalandi is already underway in
229
Australia where fingerlings are hatched and grown to maturity in sea cages (Primary
230
Industries and Resources South Australia, 2011). Already in Japan there is a very
231
close related species farmed, the Japanese Yellowtail (Seriola quinqueradiata),
232
which forms the basis of a major aquaculture industry in Japan, where wild caught
233
juveniles are grown out in sea cages. Despite an increase in demand of these
234
farmed species, there is a general lack of scientific and farming knowledge and
235
significant problems exist with diseases (Stephens and Savage, 2010; Burger et al.,
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2008; Kawanishi et al., 2005; 2006; Tubbs et al., 2004; Kobayashi, 2010; Mansell et
237
al., 2005). Having the appropriate molecular tools to investigate and understand
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these problems will improve the current husbandry of S. lalandi and provide
239
countries such as New Zealand interested in its aquaculture with the ability to
240
successfully introduce this species.
241
To date, limited studies have been done to determine and understand the
243
genetics of S. lalandi which would provide invaluable insights into the physiology of
244
this fish and allow the determination of the optimum conditions for its culture. Work
245
has begun within S. quinqueradiata, where there are 2936 EST sequences from the
246
spleen and kidney, available from a study into its immune response and a cDNA
247
microarray has been constructed using 1001 selected EST sequences to allow
248
immune gene profiling (Darawiroj et al., 2008). Currently, there are very few S.
249
lalandi nucleotide sequences deposited within the National Center for Biotechnology
250
Information (NCBI) nucleotide database, which include a limited number of genes for
251
this species (Supplementary Table 1). Previous research into the immune
252
responses of S. lalandi has relied on physiological analysis such as studying mucus
253
and serum (Leef & Lee, 2008) or the concentration of blood lactate and plasma
254
osmolality (Mansell et al., 2005). A greater understanding is desperately required for
255
the immune system of Seriola sp. and NGS technology provides the data needed for
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a comprehensive study of these species. Using this approach will allow the
257
characterization of important immune components, such as cytokines, which will
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then allow informative future studies to be carried out.
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3.2 S. lalandi transcriptomic library
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With the availability of genomes within selected fish species, gene identification
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had already accelerated and with the development of new technologies in the area of
263
transcriptomics, obtaining genetic information from a new fish species is now faster
264
and cheaper. There are a number of different next generation sequencing platforms
265
available (Quail et al., 2012) that can be used to produce a transcriptomic database
266
using RNA-Seq. However, factors such as the size of the transcriptome to be
267
sequenced, how much money is available and read length are important
268
considerations that will help determine which one is most suitable to use. In this
269
investigation, to identify cytokine genes from S. lalandi, a high throughput next
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271
Merriman et al., 2012). This approach relies on the detection of hydrogen ions
272
released when a complementary nucleotide is incorporated into a strand of DNA,
273
leading to a lowering of pH, which is measured by a semi-conductor. This is one of
274
the more recent approaches available and its main advantage over other NGS
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platforms, is its overall low cost and size of the read lengths (200-400bp).
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Wild yellowtail kingfish, S. lalandi were caught near Gannet Island, off the coast
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from Kawhia, New Zealand and the pituitary, gonad and spleen were collected and
279
used to produce an RNA-Seq library (Muncaster et al., 2017). The raw IonTorrent
280
transcriptome sequence libraries had 3,147,845 spleen (Supplementary Figure 1)
281
and 3,646,264 gonad/pituitary (Supplementary Figure 2) reads. After undergoing
282
de novo assembly using Trinity (Grabherr et al., 2011) 50,570 contigs (length range
283
200 to 16046 bp) for the spleen and 13178 contigs (length range 200 to 10497 bp)
284
for the gonad/pituitary were constructed. The assembled contigs were submitted to
285
the webtool 'FastAnnotator' (Chen et al., 2012), where tBLASTn (Gertz et al., 2006)
286
was used to translate each contig into all six reading frames and then use each one
287
to search the NCBI's non-redundant protein database. Each identified protein was
288
then submitted into Blast2GO (Götz et al., 2008) for the GO term annotation. The
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BLAST search was able to align 32,383 of the spleen contigs and 7898 of the
290
pituitary/gonad contigs to known proteins in the non-redundant protein database.
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The GO enrichment analysis from the spleen (Figure 2) and the gonad/pituitary
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(Figure 3) transcripts identified a number involved within immune system processes.
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From these transcripts, cytokines and their receptors were able to be identified and
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submitted into Genbank.
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4. Yellowtail kingfish cytokine and cytokine receptor gene discovery
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During the last 20 years many of the immune genes known in mammalian
298
organisms have been identified in teleost fish, particularly in those fish that are
299
important for aquaculture, such as trout and salmon. Teleosts have been found to
300
have a very similar immune cell repertoire and have many of the protein components
301
important for innate and adaptive immune responses that have been characterized 11
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within mammals (Zhu, et al., 2013). This includes important receptors, involved in
303
initiating innate responses such as Toll-like receptors (TLR’s) and includes members
304
such as TLR-1, TLR-2, TLR-3, TLR-4, TLR-5, TLR-7, TLR-8 and TLR-9 (Pietretti &
305
Wiegertjes, 2014). Important cellular markers critical for adaptive responses, such as
306
T-cell receptor (TCR),
307
histocompatibility complex (MHC) class I and II (Zhu, et al., 2013). In addition, a
308
large number of the molecules important in regulating immunity, the cytokines have
309
also been characterised and include many of the members of the interleukin (IL)
310
family (Secombes et al., 2016), interferon (IFN) family (Zou et al., 2016), tumor
311
necrosis factor (TNF) family (Secombes et al., 2016), chemokine family (Bird &
312
Tafalla, 2015), colony stimulating factor family (Santos et al., 2006; Wang et al.,
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2008), fibroblast growth factor family (Itoh, 2007) and transforming growth factor
314
family (Bobe et al., 2009) . Lastly, there have also been a number of cytokines
315
discovered, that are novel to fish, with no known mammalian homologue (Husain et
316
al., 2012; Hong et al., 2013; Angosto et al., 2013) and the roles of which remain to
317
be determined.
CD4,
CD8,
immunoglobulin (Ig) and
major
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Up until this study, the only cytokine related genes found in S. lalandi were
319
partial mRNA sequences for ACKR3, CXCR4, CXCR7, IL-8, IFN-γ genes and
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complete mRNA sequences for IL-1β and TNF-α genes (Supplementary Table 1).
321
This lack of information available for cytokine genes means that many previous
322
studies that have investigated the effects of stress or pathogens in Seriola species
323
have had to use mortality or pathological traits such as serum and mucus production,
324
blood glucose and lactate concentrations, muscle pH and lactate concentrations and
325
plasma osmolality to determine the physiology of the fish (Mansell et al., 2005; Leef
326
& Lee, 2009; Moran et al., 2008). While this provides useful information about the
327
effects of disease and stress on S. lalandi, using actual cytokine gene expression
328
studies would allow a more in depth analysis of these effects at the molecular level,
329
which would be particularly useful for the monitoring of immune system response to
330
disease, vaccinations, stress and general fish health (Darawiroj et al., 2008).
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The spleen tissue was targeted for transcriptomics as is the main filter for
333
blood-borne pathogens and antigens and is a key organ for iron metabolism and 12
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335
is important for the regulation of innate and adaptive immune responses locally and
336
throughout the whole body (Bronte & Pittet, 2013). The pituitary and gonad
337
transcriptome was available as it had been used for a study into sex differentiation.
338
However, in mammals evidence exists that cytokines are produced by pituitary cells
339
can mediate development, mature function, and cellular organization of the anterior
340
pituitary and also directly regulate specific pituitary trophic hormone gene expression
341
(Ray & Melmed, 1997). In addition, studies in fish have shown cytokine expression
342
does occur in the gonads (Chaves-Pozo et al, 2008). Using the S. lalandi spleen and
343
pituitary/gonad transcriptomic databases that were generated and subsequently
344
searched, a significant number of cytokine genes and receptors were discovered
345
(Table 1) important in both the innate and adaptive immune responses. Most of
346
these assembled sequences represent partial mRNA’s that can be translated to
347
provide a partial amino acid sequence, however in some cases they can code for the
348
full amino acid sequence. For example, using the transcriptomic libraries it was
349
possible to assemble the full open reading frame for S. lalandi TNFSF13B, also
350
known as B cell activating factor belonging to the TNF family (BAFF). This is an
351
important member of the TNF ligand superfamily (Schneider, 2005) and is critical for
352
the survival/maturation and proliferation for peripheral B cells, (Rolink & Melchers,
353
2002; Stein et al., 2002; Mackay et al., 2003; Mackay and Leung, 2006). BAFF is a
354
type II transmembrane protein and is found either anchored to the cell surface or
355
released in a soluble form after cleavage by a furin-like protease, with both forms
356
having biological activity (Moore et al., 1999; Schneider et al., 1999). TNFSF13B has
357
been characterised in a number of fish species, including zebrafish (Liang et al.,
358
2010), mefugu (Ai et al., 2011), Japanese sea perch (Cui et al., 2012); grass carp
359
(Pandit et al., 2013), yellow grouper (Xiao et al., 2014), miiuy coaker (Meng et al.,
360
2015) and tongue sole (Sun & Sun, 2015). Multiple alignment of the predicted S.
361
lalandi BAFF amino acid sequence with selected vertebrate sequences identified
362
important features (Figure 4) such as a transmembrane domain, a furin protease
363
cleavage site, a TNF homology domain and a conserved D-E loop (known as the
364
“Flap”), which is unique to TNFSF13B and found in no other TNFSF. Interestingly,
365
previous investigations had revealed that many fish species have more than one
366
TNFSF13B gene present (Secombes et al., 2016), forming two very distinct groups.
367
Phylogenetic analysis shows that the S. lalandi BAFF isolated is more related to the
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369
represented by a large number of transcripts within the spleen to enable its
370
construction. Previous expression studies of this gene within healthy tissues of fish,
371
showed consistently that it could be detected within immune relevant tissues, where
372
highest expression was in the spleen.
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5. The use of omics in future fish cytokine research
375
Currently, there are still a significant number of investigations using microarrays
376
to investigate teleost immune gene responses (including cytokine gene expression)
377
to disease or changes in their environment using all the gene information that has
378
been discovered in a select number of species (Boltaña et al., 2017; Schaeck et al.,
379
2017; Diaz et al., 2017; Kaneshige et al., 2016; Ferraresso et al., 2016; Eslamloo et
380
al., 2016; Pacitti et al., 2016). Obviously these studies are in those species which
381
have had a significant amount of gene discovery already carried out or a genome is
382
available, however RNA-seq allows transcriptomics investigations to be carried out in
383
any fish species with no previous genetic background. Recent studies, include a
384
number of previously well studied species, which includes Atlantic salmon and
385
rainbow trout (Morera et al., 2011; Valenzuela-Miranda et al., 2015; 2016; Núñez-
386
Acuña et al., 2015; Long et al., 2015; Marancik et al., 2015; Ali et al., 2014), Three-
387
spinned stickleback (Brown et al., 2016; Haase et al., 2016a; 2016b; Huang et al.,
388
2016), zebrafish (Hartig et al., 2016; Li et al., 2014; Li et al., 2016; Ordas et al., 2010;
389
Yang et al., 2012; Yang et al., 2015), Nile Tilapia (Wang et al., 2016a; Zhang et al.,
390
2013; Zhu et al., 2015; 2017), Channel catfish (Beck et al., 2012; Li et al., 2012, Liu
391
et al., 2013; Peatman et al., 2013; Sun et al., 2012), common carp (Neave et al.,
392
2017; Zhang et al., 2011; Zhou et al., 2016), grass carp (Dang et al., 2016), Fugu
393
(Cui et al., 2014), European seabass (Sarropoulou et al., 2012); Olive flounder
394
(Hwang et al., 2017); turbot (Gao et al., 2016; Robledo et al., 2014; Ronza et al.,
395
2016). But its understudied fish, such as the javelin goby (Chen et al., 2016), large
396
yellow croaker (Wang et al., 2016b; Zhang et al., 2017a), fathead minnow (Wiseman
397
et al., 2013), rare minnow (Wang et al., 2016c), Arctic charr (Norman et al., 2014),
398
Gulf killifish (Garcia et al., 2012), bluntsnout bream (Sun et al., 2014), common roach
399
(Brinkmann et al., 2016), barramundi (Xia et al., 2013; Hook et al., 2017), European
400
eel (Callol et al., 2015), blue catfish (Li et al., 2013); Japanese seabass (Xiang et al.,
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402
Yellow-fin rockcod (Papetti et al., 2015); emerald rockcod (Gerdol et al., 2015);
403
winter flounder (McElroy et al., 2015) where this approach is having a large impact.
404
Not only is it allowing the identification of cytokine genes within these species, but it
405
is simultaneously allowing accurate measurements of their expression under
406
different conditions.
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Interestingly with the development of NGS approaches and RNA-seq, we now
408
have a good idea of the cytokine gene repertoire we should expect to find within a
409
fish and transcriptomics may only appear to be helping to speed up their
410
identification in a newly studied fish species. But this is not entirely true and there
411
remains a lot that we can learn using this approach and justifies its use above
412
platforms, such as microarray. The information obtained during RNA-seq offers an
413
unbiased detection of transcripts, meaning that it can detect novel transcripts,
414
duplicated genes, indels (small insertions and deletions), and single nucleotide
415
variants (Hegedus et al., 2009; Robinson et al., 2012; Schunter et al., 2014; Wan et
416
al., 2015; Wang et al., 2014). Experiments can also be designed to determine the
417
types of non-coding transcripts and microRNAs (Desvignes et al., 2014; Jiang et al.,
418
2016 ; Pauli et al., 2012; Wang et al., 2016; Yang et al., 2015), present in an
419
experiment, which may play a role in cytokine gene expression. These are all things
420
that have not been investigated to any great detail in relation to fish cytokine genes
421
and will be easy to do now with the available and future technologies. Additionally,
422
RNA-seq is much more sensitive than microarray, being able to quantify discrete,
423
digital sequencing read counts instead of using gene expression measurements
424
limited by background fluorescence at the low end and signal saturation at the high
425
end. Lastly, the detection of rare transcripts, single transcripts per cell, or weakly
426
expressed genes can easily be targeted, by increasing sequencing coverage depth.
427
Each of these advantages will allow a new understanding into the cytokine genes
428
expressed under different conditions within a fish species and their regulation, to a
429
detail not previously possible.
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6. Conclusion
432
Understanding the physiological responses of farmed fish to their surrounding environment
434
transcriptomics are areas of molecular biology that are beginning to have a large
435
impact on aquaculture research. These techniques can be used to help identify
436
immune genes of interest, such as cytokines and their receptors, in any farmed
437
species for their use in quantitative expression. Using the latest technology available,
438
we have been able to identify a large number of S. lalandi cytokine genes and
439
receptors involved in inflammatory and adaptive immune responses. The
440
inflammatory response is an important part of innate immunity and pro-inflammatory
441
cytokines have been proven especially useful when looking at immune responses of
442
fish following pathogenic infection or injury (Covello et al., 2009; Lepen-Pleić et al,
443
2013). Understanding adaptive immune responses in fish will be critical for
444
successful therapeutic approaches to be developed, such as vaccinations
445
(Brudeseth et al., 2013; Zhu et al., 2013) and is an area that still requires a lot of
446
investigation. However, this is beginning to change as new molecular tools and
447
approaches are being used and developed, such as transcriptomics and the
448
development of monoclonal antibodies (Alnabulsi et al., 2013). This is allowing more
449
intricate investigations of a fish’s response to be monitored, and will lead to a greater
450
understanding of how vaccines perform (Gioacchini et al., 2008; von Gersdorff
451
Jorgensen et al., 2008; Sarropoulou et al., 2012).
essential
to
solve
production
bottlenecks.
Genomics
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With the use of transcriptomic approaches, researchers are able to easily identify
454
cytokines and their receptors in species where little of no genetic research has
455
previously been carried out, which will allow more informative studies into the
456
immune responses of these fish. These studies will aim to; (1) characterise important
457
cytokine related genes and identify any novel genes or transcripts present, (2)
458
monitor fish health and understand their immune responses to disease, and (3)
459
investigate the physiological responses of the fish to changes in their environment to
460
identify optimal farming conditions for farming of fish such as S. lalandi. We expect
461
this approach to help develop new technology and practises for the aquaculture of
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welfare.
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ACCEPTED MANUSCRIPT 7. References Ahn DH, Kang S, Park H (2016) Transcriptome analysis of immune response genes induced by pathogen agonists in the Antarctic bullhead notothen Notothenia coriiceps. Fish Shellfish Immunol. 55:315-22.
RI PT
Ali A, Rexroad CE, Thorgaard GH, Yao J, Salem M (2014) Characterization of the rainbow trout spleen transcriptome and identification of immune-related genes. Front Genet. 14;5:348. Alnabulsi A, Cash B, Bird S, Secombes CJ (2013) Developing tools to advance our understanding of fish immunity. Fish & Shellfish Immunology 34: 1636-37
SC
Altmann SM, Mellon MT, Distel DL, Kim CH (2003). Molecular and functional analysis of an interferon gene from the zebrafish, Danio rerio. J. Virol. 77: 1992-200.
M AN U
Amemiya CT, Alföldi J, Lee AP, Fan S, Philippe H, Maccallum I, Braasch I, Manousaki T et al. (2013) The African coelacanth genome provides insights into tetrapod evolution. Nature 496: 311-16. Angosto D, Montero J, López-Muñoz A, Alcaraz-Pérez F, Bird S, Sarropoulou E, Abellán E, Meseguer J, Sepulcre MP, Mulero V (2013) Identification and functional characterization of a new IL-1 family member, IL-1Fm2, in most evolutionarily advanced fish. Innate Immunity 20:487-500
TE D
Ao J, Mu Y, Xiang LX, Fan D, Feng M, Zhang S, Shi Q, Zhu LY, Li T, Ding Y, Nie L, Li Q, Dong WR, Jiang L, Sun B, Zhang X, Li M, Zhang HQ, Xie S, Zhu Y, Jiang X, Wang X, Mu P, Chen W, Yue Z, Wang Z, Wang J, Shao JZ, Chen X (2015) Genome sequencing of the perciform fish Larimichthys crocea provides insights into molecular and genetic mechanisms of stress adaptation. PLoS Genet. 11:e1005118.
EP
Aparicio S, Chapman J, Stupka E, Putnam N, Chia J, Dehal P et al., (2002). Wholegenome shotgun assembly and analysis of the genome of Fugu rubripes. Science, 297, 1301-1310. Austin CM, Tan MH, Croft LJ, Hammer MP, Gan HM (2015) Whole Genome Sequencing of the Asian Arowana (Scleropages formosus) Provides Insights into the Evolution of Ray-Finned Fishes. Genome Biol Evol. 7:2885-95.
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Balm PH, van Lieshout E, Lokate J, Wendelaar Bonga SE (1995) Bacterial lipopolysaccharide (LPS) and interleukin 1 (IL-1) exert multiple physiological effects in the tilapia Oreochromis mossambicus (Teleostei). J Comp Physiol B. 165:85-92. Barrett, KE (1996) Cytokines: sources, receptors and signalling. Baillière's Clinical Gastroenterology, 10, 1-15. Beck BH, Farmer BD, Straus DL, Li C, Peatman E (2012) Putative roles for a rhamnose binding lectin in Flavobacterium columnare pathogenesis in channel catfish Ictalurus punctatus. Fish Shellfish Immunol. 33:1008-15.
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ACCEPTED MANUSCRIPT Berthelot C, Brunet F, Chalopin D, Juanchich A, Bernard M, Noël B, Bento P, Da Silva C, Labadie K, Alberti A (2014) The rainbow trout genome provides novel insights into evolution after whole-genome duplication in vertebrates. Nat Commun. 5:3657.
RI PT
Bird S, Zou J, Kono T, Sakai M, Dijkstra JM, Secombes C. (2005a) Characterisation and expression analysis of interleukin 2 (IL-2) and IL-21 homologues in the Japanese pufferfish, Fugu rubripes, following their discovery by synteny. Immunogenetics. 56:909-23. Bird S, Zou J, Savan R, Kono T, Sakai M, Woo J, Secombes C (2005b) Characterisation and expression analysis of an interleukin 6 homologue in the Japanese pufferfish, Fugu rubripes. Dev Comp Immunol. 29:775-89.
SC
Bird S, Tafalla C (2015) Teleost chemokines and their receptors. Biology 4:756-84
M AN U
Bobe J, Nguyen T, Fostier A (2009) Ovarian function of the trout preovulatory ovary: new insights from recent gene expression studies. Comp Biochem Physiol A Mol Integr Physiol. 153:63-8. Boltaña S, Castellana B, Goetz G, Tort L, Teles M, Mulero V, Novoa B, Figueras A, Goetz FW, Gallardo-Escarate C, Planas JV, Mackenzie S (2017) Extending Immunological Profiling in the Gilthead Sea Bream, Sparus aurata, by Enriched cDNA Library Analysis, Microarray Design and Initial Studies upon the Inflammatory Response to PAMPs. Int J Mol Sci. 18: E317.
TE D
Bostock J, McAndrew B, Richards R, Jauncey K, Telfer T, Lorenzen K, Little D, Ross L, Handisyde N, Gatward I, Corner R (2010) Aquaculture: global status and trends Philosophical Transactions of the Royal Society B: Biological Sciences 365: 28972912.
EP
Braasch I, Gehrke AR, Smith JJ, Kawasaki K, Manousaki T, Pasquier J, Amores A, Desvignes T, Batzel P et al., (2016) The spotted gar genome illuminates vertebrate evolution and facilitates human-teleost comparisons. Nat Genet. 48:427-37. Brawand D, Wagner CE, Li YI, Malinsky M, Keller I, Fan S, Simakov O, Ng AY, Lim ZW, Bezault E et al., (2014) The genomic substrate for adaptive radiation in African cichlid fish. Nature. 513:375-81.
AC C
513 514 515 516 517 518 519 520 521 522 523 524 525 526 527 528 529 530 531 532 533 534 535 536 537 538 539 540 541 542 543 544 545 546 547 548 549 550 551 552 553 554 555 556 557 558 559 560 561 562
Brinkmann M, Koglin S, Eisner B, Wiseman S, Hecker M, Eichbaum K, Thalmann B, Buchinger S, Reifferscheid G, Hollert H. (2016) Characterisation of transcriptional responses to dioxins and dioxin-like contaminants in roach (Rutilus rutilus) using whole transcriptome analysis. Sci Total Environ. 541:412-23. Bronte V, Pittet MJ (2013) The spleen in local and systemic regulation of immunity. Immunity. 39:806-18. Brown M, Hablützel P, Friberg IM, Thomason AG, Stewart A, Pachebat JA, Jackson JA (2016) Seasonal immunoregulation in a naturally-occurring vertebrate. BMC Genomics. 17:369. 19
ACCEPTED MANUSCRIPT Brudeseth BE, Wiulsrød R, Fredriksen BN, Lindmo K, Løkling KE, Bordevik M, Steine N, Klevan A, Gravningen K (2013) Status and future perspectives of vaccines for industrialised fin-fish farming. Fish Shellfish Immunol. 35:1759-68.
RI PT
Burger MA, Barnes AC, Adlard RD (2008) Wildlife as reservoirs for parasites infecting commercial species: host specificity and a redescription of Kudoa amamiensis from teleost fish in Australia. Journal of Fish Diseases 31: 835-44. Callol A, Reyes-López FE, Roig FJ, Goetz G, Goetz FW, Amaro C, MacKenzie SA (2015) An Enriched European Eel Transcriptome Sheds Light upon Host-Pathogen Interactions with Vibrio vulnificus. PLoS One. 10:e0133328.
SC
Cawthorn DM, Steinman HA, Witthuhn RC (2012) Evaluation of the 16S and 12S rRNA genes as universal markers for the identification of commercial fish species in South Africa. Gene 491:40-48
M AN U
Cerdà J, Douglas S, Reith M (2010) Genomic resources for flatfish research and their applications. J. Fish Biol. 77: 1045-70. Chaves-Pozo E, Liarte S, Fernández-Alacid L, Abellán E, Meseguer J, Mulero V, García-Ayala A (2008) Pattern of expression of immune-relevant genes in the gonad of a teleost, the gilthead seabream (Sparus aurata L.). Mol Immunol. 45:2998-3011.
TE D
Chen TW, Gan RCR, Wu TH, Huang PJ, Lee CY, Chen YYM, Chen CC, Tang P (2012) FastAnnotator: an efficient transcript annotation web tool. BMC Genomics 13:Suppl 7, S9
EP
Chen QL, Luo Z, Huang C, Pan YX, Wu K (2016) De novo characterization of the liver transcriptome of javelin goby Synechogobius hasta and analysis of its transcriptomic profile following waterborne copper exposure. Fish Physiol Biochem. 42:979-94. Cho HK, Kim J, Moon JY, Nam BH, Kim YO, Kim WJ, Park JY, An CM, Cheong J, Kong HJ (2016) Microarray analysis of gene expression in olive flounder liver infected with viral haemorrhagic septicaemia virus (VHSV). Fish Shellfish Immunol. 49:66-78.
AC C
563 564 565 566 567 568 569 570 571 572 573 574 575 576 577 578 579 580 581 582 583 584 585 586 587 588 589 590 591 592 593 594 595 596 597 598 599 600 601 602 603 604 605 606 607 608 609 610 611 612
Costa V, Angelini C, De Feis I, Ciccodicola A (2010) Uncovering the complexity of transcriptomes with RNA-Seq. Journal of Biomedicine and Biotechnology 2010: 853916. Covello JM, Bird S, Morrison RN, Bridle AR, Battaglene SC, Secombes CJ, Nowak BF (2013) Isolation of RAG-1 and IgM transcripts from the striped trumpeter (Latris lineata), and their expression as markers for development of the adaptive immune response. Fish & Shellfish Immunology 34:778-88. Cui J, Liu S, Zhang B, Wang H, Sun H, Song S, Qiu X, Liu Y, Wang X, Jiang Z, Liu Z (2014) Transciptome analysis of the gill and swimbladder of Takifugu rubripes by RNA-Seq. PLoS One. 9(1):e85505. 20
ACCEPTED MANUSCRIPT Dang Y, Xu X, Shen Y, Hu M, Zhang M, Li L, Lv L, Li J (2016) Transcriptome Analysis of the Innate Immunity-Related Complement System in Spleen Tissue of Ctenopharyngodon idella Infected with Aeromonas hydrophila. PLoS One 11:e0157413.
RI PT
Darawiroj D, Kondo H, Hirono I, Aoki T (2008) Immune-related gene expression profiling of yellowtail (Seriola quinqueradiata) kidney cells stimulated with ConA and LPS using microarray analysis. Fish & Shellfish Immunology 24: 260-6. Davidson WS, Koop BF, Jones SJ, Iturra P, Vidal R, Maass A, Jonassen I, Lien S, Omholt SW (2010) Sequencing the genome of the Atlantic salmon (Salmo salar). Genome Biol. 11:403.
SC
Desvignes T, Beam MJ, Batzel P, Sydes J, Postlethwait JH (2014) Expanding the annotation of zebrafish microRNAs based on small RNA sequencing. Gene 546:3869
M AN U
Dheilly NM, Adema C, Raftos DA, Gourbal B, Grunau C, Du Pasquier L (2014) No more non-model species: the promise of next generation sequencing for comparative immunology. Dev Comp Immunol. 45:56-66.
TE D
Diaz de Cerio O, Bilbao E, Ruiz P, Pardo BG, Martínez P, Cajaraville MP, Cancio I (2017) Hepatic gene transcription profiles in turbot (Scophthalmus maximus) experimentally exposed to heavy fuel oil nº 6 and to styrene. Mar Environ Res. 123:14-24. Dinarello CA (2007) Historical insights into cytokines. Eur J Immunol. 37:S34-45.
EP
Eslamloo K, Xue X, Booman M, Smith NC, Rise ML (2016) Transcriptome profiling of the antiviral immune response in Atlantic cod macrophages. Dev Comp Immunol. 63:187-205. Felsenstein J (1985) Confidence limits on phylogenies: An approach using the bootstrap. Evolution 39: 783–791.
AC C
613 614 615 616 617 618 619 620 621 622 623 624 625 626 627 628 629 630 631 632 633 634 635 636 637 638 639 640 641 642 643 644 645 646 647 648 649 650 651 652 653 654 655 656 657 658 659 660 661
Fernández JA, Bubner EJ, Takeuchi Y, Yoshizaki G, Wang T, Cummins SF, Elizur A (2015) Primordial germ cell migration in the yellowtail kingfish (Seriola lalandi) and identification of stromal cell-derived factor 1. Gen Comp Endocrinol. 213:16-23. Ferraresso S, Bonaldo A, Parma L, Buonocore F, Scapigliati G, Gatta PP, Bargelloni L (2016) Ontogenetic onset of immune-relevant genes in the common sole (Solea solea). Fish Shellfish Immunol. 57:278-92. Forné I, Abián J, Cerdà J (2010) Fish proteome analysis: model organisms and nonsequenced species. Proteomics 10: 858-72. Gao C, Fu Q, Su B, Zhou S, Liu F, Song L, Zhang M, Ren Y, Dong X, Tan F, Li C (2016) Transcriptomic profiling revealed the signatures of intestinal barrier alteration
21
ACCEPTED MANUSCRIPT and pathogen entry in turbot (Scophthalmus maximus) following Vibrio anguillarum challenge. Dev Comp Immunol. 65:159-68. Garcia TI, Shen Y, Crawford D, Oleksiak MF, Whitehead A, Walter RB (2012) RNASeq reveals complex genetic response to Deepwater Horizon oil release in Fundulus grandis. BMC Genomics. 13:474.
RI PT
Gerdol M, Buonocore F, Scapigliati G, Pallavicini A (2015) Analysis and characterization of the head kidney transcriptome from the Antarctic fish Trematomus bernacchii (Teleostea, Notothenioidea): a source for immune relevant genes. Mar Genomics. 20:13-5.
SC
Gertz EM, Yu YK, Agarwala R, Schäffer AA, Altschul SF (2006) Composition-based statistics and translated nucleotide searches: improving the TBLASTN module of BLAST. BMC Biol. 4:41.
M AN U
Gioacchini G, Smith P, Carnevali O (2008) Effects of Ergosan on the expression of cytokine genes in the liver of juvenile rainbow trout (Oncorhynchus mykiss) exposed to enteric red mouth vaccine. Veterinary immunology and immunopathology 123:215-22 Götz S, García-Gómez JM, Terol J, Williams TD, Nagaraj SH, Nueda MJ, Robles M, Talón M, Dopazo J, Conesa A (2008) High-throughput functional annotation and data mining with the Blast2GO suite. Nucleic Acids Res. 36:3420-35.
TE D
Grabherr, M.G., Haas, B.J., Yassour, M., Levin, J.Z., Thompson, D.A., Amit, I., Adiconis, X., Fan, L., Raychowdhury, R., Zeng, Q., et al. (2011). Full-length transcriptome assembly from RNA-Seq data without a reference genome. Nature biotechnology 29:644-652.
EP
Haase D, Rieger JK, Witten A, Stoll M, Bornberg-Bauer E, Kalbe M, Reusch TB (2016a) Immunity comes first: the effect of parasite genotypes on adaptive immunity and immunization in three-spined sticklebacks. Dev Comp Immunol. 54:137-44. Haase D, Rieger JK, Witten A, Stoll M, Bornberg-Bauer E, Kalbe M, SchmidtDrewello A, Scharsack JP, Reusch TB (2016b) Comparative transcriptomics of stickleback immune gene responses upon infection by two helminth parasites, Diplostomum pseudospathaceum and Schistocephalus solidus. Zoology (Jena) 119(4):307-13
AC C
662 663 664 665 666 667 668 669 670 671 672 673 674 675 676 677 678 679 680 681 682 683 684 685 686 687 688 689 690 691 692 693 694 695 696 697 698 699 700 701 702 703 704 705 706 707 708 709 710
Hardie LJ, Laing KJ, Daniels GD, Grabowski PS, Cunningham C, Secombes CJ (1998) Isolation of the first piscine transforming growth factor beta gene: analysis reveals tissue specific expression and a potential regulatory sequence in rainbow trout (Oncorhynchus mykiss). Cytokine 10:555-63. Hartig EI, Zhu S, King BL, Coffman JA (2016) Cortisol-treated zebrafish embryos develop into pro-inflammatory adults with aberrant immune gene regulation. Biol Open. 5:1134-41.
22
ACCEPTED MANUSCRIPT Hegedus Z, Zakrzewska A, Agoston VC, Ordas A, Rácz P, Mink M, Spaink HP, Meijer AH (2009) Deep sequencing of the zebrafish transcriptome response to mycobacterium infection. Mol Immunol. 46:2918-30. Hong S, Li R, Xu Q, Secombes CJ, Wang T (2013) Two types of TNF-α exist in teleost fish: Phylogeny, expression, and bioactivity analysis of type-II TNF-α3 in rainbow trout Oncorhynchus mykiss. J. Immunol. 191:5959-72.
RI PT
Hook SE, Kroon FJ, Metcalfe S, Greenfield PA, Moncuquet P, McGrath A, Smith R, Warne MS, Turner RD, McKeown A, Westcott DA (2017) Global transcriptomic profiling in barramundi (Lates calcarifer) from rivers impacted by differing agricultural land uses. Environ Toxicol Chem. 36:103-112.
SC
Howe K, Clark MD, Torroja CF, Torrance J, Berthelot C, Muffato M, Collins JE, Humphray S et al. (2013) The zebrafish reference genome sequence and its relationship to the human genome. Nature 496: 498-503.
M AN U
Huang Y, Chain FJ, Panchal M, Eizaguirre C, Kalbe M, Lenz TL, Samonte IE, Stoll M, Bornberg-Bauer E, Reusch TB, Milinski M, Feulner PG (2016). Transcriptome profiling of immune tissues reveals habitat-specific gene expression between lake and river sticklebacks. Mol Ecol. 25:943-58. Husain M, Bird S, van Zwieten R, Secombes CJ, Wang T (2012) Cloning of the IL1β3 gene and IL-1β4 pseudogene in salmonids uncovers a second type of IL-1β gene in teleost fish. Devel. Comp. Immunol. 38:431-46.
TE D
Hwang JY, Kwon MG, Jung SH, Park MA, Kim DW, Cho WS, Park JW, Son MH (2017) RNA-Seq transcriptome analysis of the olive flounder (Paralichthys olivaceus) kidney response to vaccination with heat-inactivated viral hemorrhagic septicemia virus. Fish Shellfish Immunol. 62:221-226.
EP
Itoh N (2007) The Fgf families in humans, mice, and zebrafish: their evolutional processes and roles in development, metabolism, and disease. Biol Pharm Bull. 30:1819-25. Jaillon O, Aury JM, Brunet F, Petit JL, Stange-Thomann N, Mauceli E, Bouneau L, et al. (2004) Genome duplication in the teleost fish Tetraodon nigroviridis reveals the early vertebrate proto-karyotype. Nature 431: 946-57.
AC C
711 712 713 714 715 716 717 718 719 720 721 722 723 724 725 726 727 728 729 730 731 732 733 734 735 736 737 738 739 740 741 742 743 744 745 746 747 748 749 750 751 752 753 754 755 756 757 758 759 760
Jang SI, Hardie LJ, Secombes CJ (1994) Effects of transforming growth factor beta 1 on rainbow trout Oncorhynchus mykiss macrophage respiratory burst activity. Dev Comp Immunol. 18:315-23. Jang SI, Hardie LJ, Secombes CJ (1995) Elevation of rainbow trout Oncorhynchus mykiss macrophage respiratory burst activity with macrophage-derived supernatants. J Leukoc Biol. 57:943-7. Jeanmougin F, Thompson JD, Gouy M, Higgins DG, Gibson TJ (1998) Multiple sequence alignment with Clustal X. Trends in Biochemical Science. 23:403-5. 23
ACCEPTED MANUSCRIPT Jiang L, Liu W, Zhu A, Zhang J, Zhou J, Wu C (2016) Transcriptome analysis demonstrate widespread differential expression of long noncoding RNAs involve in Larimichthys crocea immune response. Fish Shellfish Immunol. 51:1-8.
RI PT
Johansen SD, Coucheron DH, Andreassen M, Karlsen BO, Furmanek T, Jørgensen TE, Emblem A, Breines R, Nordeide JT, Moum T, Nederbragt AJ, Stenseth NC, Jakobsen KS (2009) Large-scale sequence analyses of Atlantic cod. Nature Biotechnology 25: 263-71. Jones FC, Grabherr MG, Chan YF, Russell P, Mauceli E, Johnson J, Swofford R, Pirun M et al. (2012) The genomic basis of adaptive evolution in threespine sticklebacks. Nature 484: 55-61
SC
Jørgensen SM, Afanasyev S, Krasnov A (2008) Gene expression analyses in Atlantic salmon challenged with infectious salmon anemia virus reveal differences between individuals with early, intermediate and late mortality. BMC Genomics. 9:179.
M AN U
Kaneshige N, Jirapongpairoj W, Hirono I, Kondo H (2016) Temperature-dependent regulation of gene expression in Japanese flounder Paralichthys olivaceus kidney after Edwardsiella tarda formalin-killed cells. Fish Shellfish Immunol. 59:298-304. Kasahara M, Naruse K, Sasaki S, Nakatani Y, Qu W, Ahsan B, Yamada T, Nagayasu Y et al. (2007) The medaka draft genome and insights into vertebrate genome evolution. Nature 447: 714-19
TE D
Kawanishi M, Kojima A, Ishihara K, Esaki H, Kijima M, Takahashi T, Suzuki S, Tamura Y (2005) Drug resistance and pulsed-field gel electrophoresis patterns of Lactococcus garvieae isolates from cultured Seriola (yellowtail, amberjack and kingfish) in Japan. Letters in Applied Microbiology 40: 322-8.
EP
Kawanishi M, Kijima M, Kojima A, Ishihara K, Esaki H, Yagyu K, Takahashi T, Suzuki S, Tamura Y (2006) Drug resistance and random amplified polymorphic DNA analysis of Photobacterium damselae ssp. piscicida isolates from cultured Seriola (yellowtail, amberjack and kingfish) in Japan. Letters in Applied Microbiology 42: 648-53.
AC C
761 762 763 764 765 766 767 768 769 770 771 772 773 774 775 776 777 778 779 780 781 782 783 784 785 786 787 788 789 790 791 792 793 794 795 796 797 798 799 800 801 802 803 804 805 806 807 808
Kedes L, Liu ET (2010) The Archon Genomics XPRIZE for whole human genome sequencing. Nature Genetics. 42: 917-918. Kobayashi T (2010) Histopathological features of cultured Japanese amberjack (Seriola quinqueradiata) with ascites occurring in the winter. Journal of Toxicologic Pathology 23: 165–169. Laing KJ, Wang T, Zou J, Holland J, Hong S, Bols N, Hirono I, Aoki T, Secombes CJ (2001) Cloning and expression analysis of rainbow trout Oncorhynchus mykiss tumour necrosis factor-alpha. Eur J Biochem 268:1315–1322
24
ACCEPTED MANUSCRIPT Langevin C, Aleksejeva E, Passoni G, Palha N, Levraud JP, Boudinot P (2013) The antiviral innate immune response in fish: evolution and conservation of the IFN system. J Mol Biol. 425:4904-20. Leef MJ, Lee PS (2009) Preliminary investigation into the killing effect of kingfish (Seriola lalandi) serum and mucus against the monogenean parasites Benedenia seriolae and Zeuxapta seriolae. Aquaculture International 17: 607-14.
RI PT
Lepen Pleić I, Secombes CJ, Bird S, Mladineo I (2014) Characterization of three proinflammatory cytokines, TNFα1, TNFα2 and IL-1β, in cage-reared Atlantic bluefin tuna Thunnus thynnus. Fish Shellfish Immunol. 36:98-112.
SC
Li C, Zhang Y, Wang R, Lu J, Nandi S, Mohanty S, Terhune J, Liu Z, Peatman E (2012) RNA-seq analysis of mucosal immune responses reveals signatures of intestinal barrier disruption and pathogen entry following Edwardsiella ictaluri infection in channel catfish, Ictalurus punctatus. Fish Shellfish Immunol. 32:816-27.
M AN U
Li C, Beck BH, Peatman E (2013) Nutritional impacts on gene expression in the surface mucosa of blue catfish (Ictalurus furcatus). Dev Comp Immunol. 44:226-34. Li F, Wang H, Liu J, Lin J, Zeng A, Ai W, Wang X, Dahlgren RA, Wang H. (2016) Immunotoxicity of β-Diketone Antibiotic Mixtures to Zebrafish (Danio rerio) by Transcriptome Analysis. PLoS One. 11(4):e0152530.
TE D
Li Z, Luo H, Li C, Huo X, Yan C, Huang X, Al-Haddawi M, Mathavan S, Gong Z (2014) Transcriptomic analysis of a transgenic zebrafish hepatocellular carcinoma model reveals a prominent role of immune responses in tumour progression and regression. Int J Cancer. 135:1564-73. Lindenmann J, Burke DC, Isaacs A. (1957) Studies on the production, mode of action and properties of interferon. Br J Exp Pathol, 38: 551-562.
EP
Lindlöf A (2003) Gene identification through large-scale EST sequence processing. Appl Bioinformatics. 2:123-9. Liu L, Li C, Su B, Beck BH, Peatman E (2013) Short-term feed deprivation alters immune status of surface mucosa in channel catfish (Ictalurus punctatus). PLoS One. 8:e74581.
AC C
809 810 811 812 813 814 815 816 817 818 819 820 821 822 823 824 825 826 827 828 829 830 831 832 833 834 835 836 837 838 839 840 841 842 843 844 845 846 847 848 849 850 851 852 853 854 855 856 857 858
Liu K, Xu D, Li J, Bian C, Duan J, Zhou Y, Zhang M, You X, You Y, Chen J, Yu H, Xu G, Fang DA, Qiang J, Jiang S, He J, Xu J, Shi Q, Zhang Z, Xu P (2017) Whole genome sequencing of Chinese clearhead icefish, Protosalanx hyalocranius. Gigascience. [Epub ahead of print] Liu Z, Liu S, Yao J, Bao L, Zhang J, Li Y, Jiang C, Sun L, Zhang Y, Zhou T et al. (2016) The channel catfish genome sequence provides insights into the evolution of scale formation in teleosts. Nat Commun. 2016;7:11757. Long M, Zhao J, Li T, Tafalla C, Zhang Q, Wang X, Gong X, Shen Z, Li A (2015) Transcriptomic and proteomic analyses of splenic immune mechanisms of rainbow 25
ACCEPTED MANUSCRIPT trout (Oncorhynchus mykiss) infected by Aeromonas salmonicida subsp. salmonicida. J Proteomics. 122:41-54. Loo GH, Schuller KA (2010) Cloning and functional characterization of a peroxiredoxin 4 from yellowtail kingfish (Seriola lalandi). Comparative Biochemistry and Physiology Part B: Biochemistry and Molecular Biology 156: 244-53.
RI PT
Malmstrøm M, Matschiner M, Tørresen OK, Jakobsen KS, Jentoft S (2017) Whole genome sequencing data and de novo draft assemblies for 66 teleost species. Sci Data. 4:160132.
SC
Mansell B, Powell MD, Ernst I, Nowak BF (2005) Effects of the gill monogenean Zeuxapta seriolae (Meserve, 1938) and treatment with hydrogen peroxide on pathophysiology of kingfish, Seriola lalandi Valenciennes, 1833. Journal of Fish Diseases 28: 253-62.
M AN U
Marancik D, Gao G, Paneru B, Ma H, Hernandez AG, Salem M, Yao J, Palti Y, Wiens GD. (2015) Whole-body transcriptome of selectively bred, resistant-, control-, and susceptible-line rainbow trout following experimental challenge with Flavobacterium psychrophilum. Front Genet. 5:453. Mardis ER (2011) A decade's perspective on DNA sequencing technology. Nature 470: 198-203.
TE D
McElroy AE, Hice LA, Frisk MG, Purcell SL, Phillips NC, Fast MD (2015) Spatial patterns in markers of contaminant exposure, glucose and glycogen metabolism, and immunological response in juvenile winter flounder (Pseudoplueronectes americanus). Comp Biochem Physiol Part D Genomics Proteomics. 2015 Jun;14:5365.
EP
McGaugh SE, Gross JB, Aken B, Blin M, Borowsky R, Chalopin D, Hinaux H, Jeffery WR, Keene A et al., (2014) The cavefish genome reveals candidate genes for eye loss. Nat Commun. 5:5307. Merriman B, Ion Torrent R&D Team, Rothberg JM (2012) Progress in ion torrent semiconductor chip based sequencing. Electrophoresis 33: 3397-417
AC C
859 860 861 862 863 864 865 866 867 868 869 870 871 872 873 874 875 876 877 878 879 880 881 882 883 884 885 886 887 888 889 890 891 892 893 894 895 896 897 898 899 900 901 902 903 904 905 906 907 908
Mommens M, Fernandes JM, Tollefsen KE, Johnston IA, Babiak I (2014) Profiling of the embryonic Atlantic halibut (Hippoglossus hippoglossus L.) transcriptome reveals maternal transcripts as potential markers of embryo quality. BMC Genomics. 15:829. Moore PA, Belvedere O, Orr A, Pieri K, LaFleur DW, Feng P, Soppet D, Charters M, Gentz R, Parmelee D, Li Y, Galperina O, Giri J, Roschke V, Nardelli B, Carrell J, Sosnovtseva S, Greenfield W, Ruben SM, Olsen HS, Fikes J, Hilbert DM (1999) BLyS: member of the tumor necrosis factor family and B lymphocyte stimulator. Science 285, 260–263 Moran D, Gwells RM, Pether SJ (2008) Low stress response exhibited by juvenile yellowtail kingfish (Seriola lalandi Valenciennes) exposed to hypercapnic conditions associated with transportation. Aquaculture Research 39: 1399-1407 26
ACCEPTED MANUSCRIPT Morera D, Roher N, Ribas L, Balasch JC, Doñate C, Callol A, Boltaña S, Roberts S, Goetz G, Goetz FW, MacKenzie SA (2011) RNA-Seq reveals an integrated immune response in nucleated erythrocytes. PLoS One 6:e26998.
RI PT
Mu Y, Ding F, Cui P, Ao J, Hu S, Chen X (2010) Transcriptome and expression profiling analysis revealed changes of multiple signaling pathways involved in immunity in the large yellow croaker during Aeromonas hydrophila infection. BMC Genomics. 11:506. Muncaster M, Kraakman K, Gibbons O, Mensink K, Forlenza M, Jacobson G, Bird, S (2017) Antimicrobial peptides within the Yellowtail Kingfish (Seriola lalandi). Dev. Comp. Immunol. (In press).
SC
Neave MJ, Sunarto A, McColl KA (2017) Transcriptomic analysis of common carp anterior kidney during Cyprinid herpesvirus 3 infection: Immunoglobulin repertoire and homologue functional divergence. Sci Rep. 7:41531.
M AN U
Nocillado JN, Biran J, Lee YY, Levavi-Sivan B, Mechaly AS, Zohar Y, Elizur A (2012) The Kiss2 receptor (Kiss2r) gene in Southern Bluefin Tuna, Thunnus maccoyii and in Yellowtail Kingfish, Seriola lalandi – functional analysis and isolation of transcript variants. Molecular and Cellular Endocrinology 362: 211-220
TE D
Nocillado JN, Zohar Y, Biran J, Levavi-Sivan B, Elizur A (2013) Chronic kisspeptin administration stimulated gonadal development in pre-pubertal male yellowtail kingfish (Seriola lalandi; Perciformes) during the breeding and non-breeding season. Gen Comp Endocrinol. 191:168-76. Norman JD, Ferguson MM, Danzmann RG (2014) An integrated transcriptomic and comparative genomic analysis of differential gene expression in Arctic charr (Salvelinus alpinus) following seawater exposure. J Exp Biol. 217:4029-42.
EP
Núñez-Acuña G, Gonçalves AT, Valenzuela-Muñoz V, Pino-Marambio J, Wadsworth S, Gallardo-Escárate C (2015) Transcriptome immunomodulation of in-feed additives in Atlantic salmon Salmo salar infested with sea lice Caligus rogercresseyi. Fish Shellfish Immunol. 47:450-60.
AC C
909 910 911 912 913 914 915 916 917 918 919 920 921 922 923 924 925 926 927 928 929 930 931 932 933 934 935 936 937 938 939 940 941 942 943 944 945 946 947 948 949 950 951 952 953 954 955 956 957 958
Ordas A, Hegedus Z, Henkel CV, Stockhammer OW, Butler D, Jansen HJ, Racz P, Mink M, Spaink HP, Meijer AH (2010) Deep sequencing of the innate immune transcriptomic response of zebrafish embryos to Salmonella infection. Fish Shellfish Immunol. 31:716-24. Pacitti D, Lawan MM, Feldmann J, Sweetman J, Wang T, Martin SA, Secombes CJ (2016) Impact of selenium supplementation on fish antiviral responses: a whole transcriptomic analysis in rainbow trout (Oncorhynchus mykiss) fed supranutritional levels of Sel-Plex®. BMC Genomics. 17:116. Page RD (1996) TreeView: an application to display phylogenetic trees on personal computers. Computer Applications in the Biosciences 12: 357–358.
27
ACCEPTED MANUSCRIPT Palstra AP, Beltran S, Burgerhout E, Brittijn SA, Magnoni LJ, Henkel CV, Jansen HJ, van den Thillart GE, Spaink HP, Planas JV (2013) Deep RNA sequencing of the skeletal muscle transcriptome in swimming fish. PLoS One 8:e53171.
RI PT
Papetti C, Harms L, Windisch HS, Frickenhaus S, Sandersfeld T, Jürgens J, Koschnick N, Knust R, Pörtner HO, Lucassen M (2015) A first insight into the spleen transcriptome of the notothenioid fish Lepidonotothen nudifrons: Resource description and functional overview. Mar Genomics. 3:237-9. Pauli A, Valen E, Lin MF, Garber M, Vastenhouw NL, Levin JZ, Fan L, Sandelin A, Rinn JL, Regev A, Schier AF (2012) Systematic identification of long noncoding RNAs expressed during zebrafish embryogenesis. Genome Res. 22:577-91.
SC
Peatman E, Li C, Peterson BC, Straus DL, Farmer BD, Beck BH (2013) Basal polarization of the mucosal compartment in Flavobacterium columnare susceptible and resistant channel catfish (Ictalurus punctatus). Mol Immunol. 56:317-27.
M AN U
Petit J, David L, Dirks R, Wiegertjes GF (2017) Genomic and transcriptomic approaches to study immunology in cyprinids: What is next? Dev Comp Immunol. [Epub ahead of print] Pietretti D, Wiegertjes GF (2014) Ligand specificities of Toll-like receptors in fish: indications from infection studies. Dev Comp Immunol. 43:205-22.
TE D
Pooley NJ, Tacchi L, Secombes CJ, Martin SA 2013. Inflammatory responses in primary muscle cell cultures in Atlantic salmon (Salmo salar). BMC Genomics 14:747 Primary Industries and Resources South Australia, 2011. PIRSA Aquaculture Marine Finfish. http://www.pir.sa.gov.au/aquaculture/aquaculture_industry/marine_finfish
EP
Quail MA, Smith M, Coupland P, Otto TD, Harris SR, Connor TR, Bertoni A, Swerdlow HP, Gu Y (2012) A tale of three next generation sequencing platforms: comparison of Ion Torrent, Pacific Biosciences and Illumina MiSeq sequencers. BMC Genomics. 13: 341. Ray D, Melmed S (1997) Pituitary cytokine and growth factor expression and action. Endocr Rev. 18:206-28.
AC C
959 960 961 962 963 964 965 966 967 968 969 970 971 972 973 974 975 976 977 978 979 980 981 982 983 984 985 986 987 988 989 990 991 992 993 994 995 996 997 998 999 1000 1001 1002 1003 1004 1005 1006 1007 1008
Reyes-Becerril M, Ascencio-Valle F, Alamillo E, Hirono I, Kondo H, Jirapongpairoj W, Angulo C (2015) Molecular cloning and comparative responses of Toll-like receptor 22 following ligands stimulation and parasitic infection in yellowtail (Seriola lalandi). Fish Shellfish Immunol. 46:323-33. Reyes-Becerril M, Ascencio-Valle F, Hirono I, Kondo H, Jirapongpairoj W, Esteban MA, Alamillo E, Angulo C (2016) TLR21's agonists in combination with Aeromonas antigens synergistically up-regulate functional TLR21 and cytokine gene expression in yellowtail leucocytes. Dev Comp Immunol. 61:107-15. Robinson N, Sahoo PK, Baranski M, Das Mahapatra K, Saha JN, Das S, Mishra Y, Das P, Barman HK, Eknath AE (2012) Expressed sequences and polymorphisms in 28
ACCEPTED MANUSCRIPT rohu carp (Labeo rohita, Hamilton) revealed by mRNA-seq. Mar Biotechnol (NY). 14:620-33. Robledo D, Ronza P, Harrison PW, Losada AP, Bermúdez R, Pardo BG, Redondo MJ, Sitjà-Bobadilla A, Quiroga MI, Martínez P (2014) RNA-seq analysis reveals significant transcriptome changes in turbot (Scophthalmus maximus) suffering severe enteromyxosis. BMC Genomics. 15:1149.
RI PT
Ronza P, Robledo D, Bermúdez R, Losada AP, Pardo BG, Sitjà-Bobadilla A, Quiroga MI, Martínez P (2016) RNA-seq analysis of early enteromyxosis in turbot (Scophthalmus maximus): new insights into parasite invasion and immune evasion strategies. Int J Parasitol. 46:507-17.
SC
Rothberg JM, Hinz W, Rearick TM, Schultz J, Mileski W, Davey M, Leamon JH, Johnson K et al., (2011) An integrated semiconductor device enabling non-optical genome sequencing. Nature. 475:348-52.
M AN U
Saitou N, Nei M (1987) The neighbor-joining method: a new method for reconstructing phylogenetic trees. Molecular Biology & Evolution 4:406-25. Sangrador-Vegas A, Lennington JB, Smith TJ (2002) Molecular cloning of an IL-8like CXC chemokine and tissue factor in rainbow trout (Oncorhynchus mykiss) by use of suppression subtractive hybridization. Cytokine. 17:66-70.
TE D
Santos MD, Yasuike M, Hirono I, Aoki T (2006) The granulocyte colony-stimulating factors (CSF3s) of fish and chicken. Immunogenetics. 58:422-32. Sarropoulou E, Galindo-Villegas J, Garcia-Alcazar A, Kasapidis P, Mulero V (2012) Characterization of european sea bass transcripts by RNA SEQ after oral vaccine against V. anguillarum. Marine Biotechnology 14: 634-42
EP
Schaeck M, Reyes-López FE, Vallejos-Vidal E, Van Cleemput J, Duchateau L, Van den Broeck W, Tort L, Decostere A (2017) Cellular and transcriptomic response to treatment with the probiotic candidate Vibrio lentus in gnotobiotic sea bass (Dicentrarchus labrax) larvae. Fish Shellfish Immunol. 63:147-156.
AC C
1009 1010 1011 1012 1013 1014 1015 1016 1017 1018 1019 1020 1021 1022 1023 1024 1025 1026 1027 1028 1029 1030 1031 1032 1033 1034 1035 1036 1037 1038 1039 1040 1041 1042 1043 1044 1045 1046 1047 1048 1049 1050 1051 1052 1053 1054 1055 1056 1057 1058
Schartl M, Walter RB, Shen Y, Garcia T, Catchen J, Amores A, Braasch I, Chalopin D, Volff JN, et al. (2013) The genome of the platyfish, Xiphophorus maculatus, provides insights into evolutionary adaptation and several complex traits. Nature Genetics 45: 567–572. Schneider P, MacKay F, Steiner V, Hofmann K, Bodmer JL, Holler N, Ambrose C, Lawton P, Bixler S, Acha-Orbea H, Valmori D, Romero P, Werner-Favre C, Zubler RH, Browning JL, Tschopp J (1999) BAFF, a novel ligand of the tumor necrosis factor family, stimulates B cell growth. J. Exp. Med. 189, 1747–1756. Schunter C, Vollmer SV, Macpherson E, Pascual M (2014) Transcriptome analyses and differential gene expression in a non-model fish species with alternative mating tactics. BMC Genomics. 15:167.
29
ACCEPTED MANUSCRIPT Secombes CJ, Wang T, Bird S (2011) The interleukins of fish. Dev. Comp. Immunol. 35:1336-1345. Secombes CJ, Zou, J, Bird S (2015) Cytokines of Cartilaginous Fish. In: Smith, Sim, Flajnik (Eds) Immunobiology of the Shark., Taylor & Francis Group Ltd. pp. 123-142
RI PT
Secombes CJ, Wang T, Bird S (2016) Vertebrate cytokines and their evolution. In: Malagoli (Ed) The Evolution of the Immune System: Conservation and Diversification. Academic Press. pp. 87-151 Star B, Nederbragt AJ, Jentoft S, Grimholt U, Malmstrøm M, Gregers TF, Rounge TB, Paulsen J, et al. (2011) The genome sequence of Atlantic cod reveals a unique immune system. Nature 477: 207-10.
SC
Stephens FJ, Savage A (2010) Two mortality events in sea-caged yellowtail kingfish Seriola lalandi Valenciennes, 1833 (Nannopercidae) from Western Australia. Australian Veterinary Journal 88: 414-6.
M AN U
Sumathy K, Desai KV, Kondaiah P (1997) Isolation of transforming growth factorbeta2 cDNA from a fish, Cyprinus carpio by RT-PCR. Gene 191:103-7. Sun F, Peatman E, Li C, Liu S, Jiang Y, Zhou Z, Liu Z (2012) Transcriptomic signatures of attachment, NF-κB suppression and IFN stimulation in the catfish gill following columnaris bacterial infection. Dev Comp Immunol. 38:169-80.
TE D
Sun S, Ge X, Xuan F, Zhu J, Yu N (2014) Nitrite-induced hepatotoxicity in Bluntsnout bream (Megalobrama amblycephala): the mechanistic insight from transcriptome to physiology analysis. Environ Toxicol Pharmacol. 37:55-65.
EP
Tacchi L, Bron JE, Taggart JB, Secombes CJ, Bickerdike R, Adler MA, Takle H, Martin SA (2011) Multiple tissue transcriptomic responses to Piscirickettsia salmonis in Atlantic salmon (Salmo salar). Physiological Genomics 43:1241-54. Tacchi L, Secombes CJ, Bickerdike R, Adler MA, Venegas C, Takle H, Martin SA (2012) Transcriptomic and physiological responses to fishmeal substitution with plant proteins in formulated feed in farmed Atlantic salmon (Salmo salar). BMC Genomics 13:363.
AC C
1059 1060 1061 1062 1063 1064 1065 1066 1067 1068 1069 1070 1071 1072 1073 1074 1075 1076 1077 1078 1079 1080 1081 1082 1083 1084 1085 1086 1087 1088 1089 1090 1091 1092 1093 1094 1095 1096 1097 1098 1099 1100 1101 1102 1103 1104 1105 1106 1107 1108
Thakur NL, Jain R, Natalio F, Hamer B, Thakur AN, Müller WE (2008) Marine molecular biology: an emerging field of biological sciences. Biotechnology Advances 26: 233-45. Trumbić Ž, Bekaert M, Taggart JB, Bron JE, Gharbi K, Mladineo I (2015) Development and validation of a mixed-tissue oligonucleotide DNA microarray for Atlantic bluefin tuna, Thunnus thynnus (Linnaeus, 1758). BMC Genomics. 16:1007 Tubbs LA, Poortenaar CW, Sewell MA, Diggles BK (2004) Effects of temperature on fecundity in vitro, egg hatching and reproductive development of Benedenia seriolae and Zeuxapta seriolae (Monogenea) parasitic on yellowtail kingfish, Seriola lalandi. International Journal of Parasitology 35: 315-27. 30
ACCEPTED MANUSCRIPT
RI PT
Valenzuela-Miranda D, Gallardo-Escárate C (2016) Novel insights into the response of Atlantic salmon (Salmo salar) to Piscirickettsia salmonis: Interplay of coding genes and lncRNAs during bacterial infection. Fish Shellfish Immunol. 59:427-438. van Dijk EL, Auger H, Jaszczyszyn Y, Thermes C (2014) Ten years of nextgeneration sequencing technology. Trends Genet. 30:418–26.
SC
Verburg-van Kemenade BM, Weyts FA, Debets R, Flik G (1995) Carp macrophages and neutrophilic granulocytes secrete an interleukin-1-like factor. Dev Comp Immunol. 19:59-70.
M AN U
Vij S, Kuhl H, Kuznetsova IS, Komissarov A, Yurchenko AA, Van Heusden P, Singh S, Thevasagayam NM, Prakki SR, Purushothaman K, Saju JM, Jiang J, Mbandi SK, et al., (2016) Chromosomal-Level Assembly of the Asian Seabass Genome Using Long Sequence Reads and Multi-layered Scaffolding. PLoS Genet. 12:e1005954. Vilček J (2003) The cytokines: an overview. In ‘The Cytokine Handbook’ Thomson AW & Lotze MT, Eds. Elsevier Ltd. pp 3-18.
TE D
von Gersdorff Jorgensen L, Nemli E, Heinecke R, Raida M, Buchmann, K (2008) Immune-relevant genes expressed in rainbow trout following immunisation with a live vaccine against Ichthyophthirius multifiliis. Diseases of Aquatic Organisms 80:189-97. Wan Q, Sua, J (2015) Transcriptome analysis provides insights into the regulatory function of alternative splicing in antiviral immunity in grass carp (Ctenopharyngodon idella). Sci Rep. 5: 12946.
EP
1141 1142 1143 1144 1145 1146 1147 1148 1149 1150 1151 1152 1153 1154 1155 1156 1157
Valenzuela-Miranda D, Boltaña S, Cabrejos ME, Yáñez JM, Gallardo-Escárate C (2015) High-throughput transcriptome analysis of ISAV-infected Atlantic salmon Salmo salar unravels divergent immune responses associated to head-kidney, liver and gills tissues. Fish Shellfish Immunol. 45:367-77.
Wang J, Fu L, Koganti PP, Wang L, Hand JM, Ma H, Yao J (2016d) Identification and Functional Prediction of Large Intergenic Noncoding RNAs (lincRNAs) in Rainbow Trout (Oncorhynchus mykiss). Mar Biotechnol (NY). 18:271-82.
AC C
1109 1110 1111 1112 1113 1114 1115 1116 1117 1118 1119 1120 1121 1122 1123 1124 1125 1126 1127 1128 1129 1130 1131 1132 1133 1134 1135 1136 1137 1138 1139 1140
Wang L, Liu P, Wan ZY, Huang SQ, Wen YF, Lin G, Yue GH (2016a) RNA-Seq revealed the impairment of immune defence of tilapia against the infection of Streptococcus agalactiae with simulated climate warming. Fish Shellfish Immunol. 55:679-89. Wang P, Wang J, Su YQ, Mao Y, Zhang JS, Wu CW, Ke QZ, Han KH, Zheng WQ, Xu ND (2016b) Transcriptome analysis of the Larimichthys crocea liver in response to Cryptocaryon irritans. Fish Shellfish Immunol. 48:1-11. Wang T, Hanington PC, Belosevic M, Secombes CJ (2008) Two macrophage colony-stimulating factor genes exist in fish that differ in gene organization and are differentially expressed. J Immunol. 181:3310-22. 31
ACCEPTED MANUSCRIPT Wang W, Wang J, You F, Ma L, Yang X, Gao J, He Y, Qi J, Yu H, Wang Z, Wang X, Wu Z, Zhang Q (2014) Detection of alternative splice and gene duplication by RNA sequencing in Japanese flounder, Paralichthys olivaceus. G3 4: 2419–2424. Wang Z, Gerstein M, Snyder M (2009). RNA-Seq: a revolutionary tool for transcriptomics. Nat Rev Genet. 10: 57-63.
RI PT
Wang ZJ, Liu XH, Jin L, Pu DY, Huang J, Zhang YG (2016c) Transcriptome profiling analysis of rare minnow (Gobiocypris rarus) gills after waterborne cadmium exposure. Comp Biochem Physiol Part D Genomics Proteomics. 19:120-8.
SC
Ward RD, Holmes BH (2007) An analysis of nucleotide and amino acid variability in the barcode region of cytochrome c oxidase I (COX1) in fishes. Molecular Ecology Notes 7: 899-907
M AN U
Wiseman SB, He Y, Gamal-El Din M, Martin JW, Jones PD, Hecker M, Giesy JP (2013) Transcriptional responses of male fathead minnows exposed to oil sands process-affected water. Comp Biochem Physiol C Toxicol Pharmacol. 157:227-35. Xia JH, Liu P, Liu F, Lin G, Sun F, Tu R, Yue GH (2013) Analysis of stressresponsive transcriptome in the intestine of Asian seabass (Lates calcarifer) using RNA-seq. DNA Res. 20:449-60.
TE D
Xiang LX, He D, Dong WR, Zhang YW, Shao JZ (2010) Deep sequencing-based transcriptome profiling analysis of bacteria-challenged Lateolabrax japonicus reveals insight into the immune-relevant genes in marine fish. BMC Genomics. 11:472. Xu J, Bian C, Chen K, Liu G, Jiang Y, Luo Q, You X, Peng W, Li J, Huang Y, Yi Y, Dong C, Deng H, Zhang S, Zhang H, Shi Q, Xu P (2017) Draft genome of the Northern snakehead, Channa argus. Gigascience. [Epub ahead of print]
EP
Yang D, Liu Q, Yang M, Wu H, Wang Q, Xiao J, Zhang Y. (2012) RNA-seq liver transcriptome analysis reveals an activated MHC-I pathway and an inhibited MHC-II pathway at the early stage of vaccine immunization in zebrafish. BMC Genomics. 13:319.
AC C
1158 1159 1160 1161 1162 1163 1164 1165 1166 1167 1168 1169 1170 1171 1172 1173 1174 1175 1176 1177 1178 1179 1180 1181 1182 1183 1184 1185 1186 1187 1188 1189 1190 1191 1192 1193 1194 1195 1196 1197 1198 1199 1200 1201 1202 1203 1204 1205 1206 1207
Yang L, Irwin DM, He S (2015) Genome-wide identification and characterization of teleost-specific microRNAs within zebrafish. Gene. 561:181-9. Yang S, Marín-Juez R, Meijer AH, Spaink HP (2015) Common and specific downstream signaling targets controlled by Tlr2 and Tlr5 innate immune signaling in zebrafish. BMC Genomics. 16:547. Zhang R, Zhang LL, Ye X, Tian YY, Sun CF, Lu MX, Bai JJ (2013) Transcriptome profiling and digital gene expression analysis of Nile tilapia (Oreochromis niloticus) infected by Streptococcus agalactiae. Mol Biol Rep. 40:5657-68. Zhang X, Mu Y, Mu P, Ao J, Chen X (2017a) Transcriptome Analysis Reveals Comprehensive Insights into the Early Immune Response of Large Yellow Croaker 32
ACCEPTED MANUSCRIPT (Larimichthys crocea) Induced by Trivalent Bacterial Vaccine. PLoS One. 12:e0170958. Zhang X, Wen H, Wang H, Ren Y, Zhao J, Li Y (2017b) RNA-Seq analysis of salinity stress-responsive transcriptome in the liver of spotted sea bass (Lateolabrax maculatus). PLoS One. 12:e0173238.
RI PT
Zhang Y, Stupka E, Henkel CV, Jansen HJ, Spaink HP, Verbeek FJ (2011) Identification of common carp innate immune genes with whole-genome sequencing and RNA-Seq data. J Integr Bioinform. 8:169.
SC
Zhao C, Fu M, Wang C, Jiao Z, Qiu L (2016) RNA-Seq analysis of immune-relevant genes in Lateolabrax japonicus during Vibrio anguillarum infection. Fish Shellfish Immunol. 2016 52:57-64.
M AN U
Zhou Y, Yu W, Zhong H, Li J, Li H, He F, Zhou J, Tang Y, Yu J, Yu F (2016) Transcriptome analysis reveals that insulin is an immunomodulatory hormone in common carp. Fish Shellfish Immunol. 59:213-219. Zhu J, Li C, Ao Q, Tan Y, Luo Y, Guo Y, Lan G, Jiang H, Gan X (2015) Trancriptomic profiling revealed the signatures of acute immune response in tilapia (Oreochromis niloticus) following Streptococcus iniae challenge. Fish Shellfish Immunol. 46:346-53.
TE D
Zhu J, Fu Q, Ao Q, Tan Y, Luo Y, Jiang H, Li C, Gan X (2017) Transcriptomic profiling analysis of tilapia (Oreochromis niloticus) following Streptococcus agalactiae challenge. Fish Shellfish Immunol. 62:202-212. Zhu LY, Nie L, Zhu G, Xiang LX, Shao JZ (2013). Advances in research of fish immune-relevant genes: a comparative overview of innate and adaptive immunity in teleosts. Developmental and Comparative Immunology 39: 39-62.
EP
Zou J, Grabowski PS, Cunningham C, Secombes CJ (1999a) Molecular cloning of interleukin 1beta from rainbow trout Oncorhynchus mykiss reveals no evidence of an ice cut site. Cytokine 11:552-60. Zou J, Cunningham C, Secombes CJ (1999b) The rainbow trout Oncorhynchus mykiss interleukin-1 beta gene has a differ organization to mammals and undergoes incomplete splicing. Eur J Biochem. 259:901-8.
AC C
1208 1209 1210 1211 1212 1213 1214 1215 1216 1217 1218 1219 1220 1221 1222 1223 1224 1225 1226 1227 1228 1229 1230 1231 1232 1233 1234 1235 1236 1237 1238 1239 1240 1241 1242 1243 1244 1245 1246 1247 1248 1249 1250 1251 1252 1253
Zou J, Bird S, Secombes CJ (2010) Antiviral sensing in teleost fish. Curr. Pharm. Des. 16:4185-93. Zou J, Castro R, Tafalla C (2016) Antiviral immunity: Origin and evolution in vertebrates. In: Malagoli (Ed) The Evolution of the Immune System: Conservation and Diversification. Academic Press. pp. 173-193
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8. Acknowledgements This study was funded by a University of Waikato Research Trust Contestable Fund
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ACCEPTED MANUSCRIPT Figure 1: Graph showing the impact that genomics and transcriptomics has had on cytokine discovery in fish over the last 20 years. The number of papers published on fish cytokine genes are shown for each year, with the discovery of a number of the early cytokines highlighted.
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Figure 2: GO enrichment analysis of the translated spleen contigs identified using tBLASTn to search the NCBI non-redundant protein database. Vertical axis represents gene ontology categories, while horizontal axis indicates the number of genes in each ontology category.
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Figure 3: GO enrichment analysis of the translated gonad and pituitary contigs identified using tBLASTn to search the NCBI non-redundant protein database. Vertical axis represents gene ontology categories, while horizontal axis indicates the number of genes in each ontology category.
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Figure 4: Multiple sequence alignment of S. lalandi BAFF with other selected vertebrate BAFF molecules using ClustalX (Jeanmougin et al., 1998). The transmembrane domain (TMD) is in bold and potential furin cleavage sites are boxed. Conserved cysteines are labelled with an arrow. The conserved TNF homology domain (THD) is highlighted in grey, with the conserved long D-E loop, known as the “Flap” highlighted in yellow. Accession numbers for BAFF are: human, Q9Y275.1; Grass carp, AGG11791.1; Zebrafish, NP_001107062.1; Mefugu, AEB69781.1; Trout, NP_001118036.1; Japanese sea perch, AEH22106.1; Chicken, NP_989658.1; Miiuy croaker, AHL44989.1; Yellow grouper, AFN70720.1; Tongue sole, XP_008326904.1; Yellowtail kingfish, ????.
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Figure 5: Phylogenetic analysis of S. lalandi BAFF (highlighted with a red box) with selected teleost, reptile, bird, amphibian and cartilaginous fish BAFF (TNFSF13B) and APRIL (TNFSF13) amino acid sequences. Accession numbers of each sequence are included in the figure. A different colour is used to indicate the clear clustering of sequences into separate groups. Analysis was performed using the neighbour-joining method (Saitou & Nei, 1987), the tree drawn using the TreeView program v1.6.1 (Page, 1996) and confidence limits added (Felsenstein, 1985).
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Supplementary Figure 1: Graph showing the distribution of the 3,147,845 contigs obtained from the spleen RNA-Seq run. The contig lengths sequenced and their frequency are plotted. Supplementary Figure 2: Graph showing the distribution of the 3,646,264 contigs obtained from the spleen RNA-Seq run. The contig lengths sequenced and their frequency are plotted.
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Table 1: Seriola lalandi cytokine and receptor genes obtained using RNA-Seq
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CXCL9 CXCL10 CXCL14 CCL3 CCL18 CCL19 CCL20 CCL25
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Chemokine Family
Accession No.
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Ligand IL-2 IL-4 IL-6 IL-7 IL-10 IL-11 IL-12 p35 IL-12 p40 IL-15 IL-16 IL-18 IL-34 EBI3
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Interleukin (IL) Family
Receptor IL-1Racp IL-1R Type I IL-1R Type II IL-2Rb IL-4Ra IL-5Ra IL-6Rb IL-7Ra IL-10Ra IL-12Rb2 IL-17RA IL-17RB IL-17RE IL-18Racp IL-20Ra IL-21R IL-31Ra γc (CD132) beta-c Glycoprotein 130 CXCR1 CXCR3 CXCR4 CXCR5 CCR1 CCR2 CCR3 CCR4 CCR7 CCR9 XCR1
Accession No.
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Table 1: Seriola lalandi cytokine and receptor genes obtained using RNA-Seq
TNF-a1 TNF-a2 TNF-b2 TNFSF6 TNFSF12 TNFSF13B TNFSF14
Interferon (IFN) Family
IFN-g
Fibroblast growth factor (FGF) Family
FGF-2 FGF-7
Colony stimulating factor (CSF) Family
M-CSF
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Transforming growth factor (TGF) Family
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Tumour necrosis factor (TNF) Family
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ACKR4
TGF-b
Leukemia inhibitory factor (LIF) Monocyte chemotactic protein (MCP) MCP1 Macrophage migration inhibitory factor (MMIF) MMIF Ciliary neurotrophic factor (CNF)
TNFRSF1A TNFRSF1B TNFRSF2 TNFRSF3 TNFRSF5 TNFRSF6 TNFRSF9 TNFRSF10B TNFRSF13B TNFRSF14 TNFRSF16 TNFRSF19 TNFRSF21 TNFRSF26 IFN-gR IFN-a/bR2 FGF-R1
M-CSFR G-CSFR TGF-bR3 LIFR
CNFR
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Figure 1
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Figure 4 TMD MDDSTEREQSRLTSCLKKREEMKLKECVSILPRKESPSVRSSKDGKLLAATLLLA-LLSCCLTVVSFY ---------MKSVDCVH---VIQQKDTASSPSG--PPGAASGTTGLFSVTFLWLAMLLSSCLAAVSLY ------------------------MAAEDVGPSRGER---RRL--PWLFLVLVVAAITSSSLSVISLY ------------------------MPAEDVGPGRGER---RRL--SWLFLVLVAAAITSSSLSVISLY -------------------MASAGPNPEGGRPASRQESGGRRL--SWLVLLLTLAAVTSSSLSALSLY -----------------------MAVLAGAKPGTGQRAGEGRL--SWPVFLLTLAAVTSSSLSALSLY -----------------------MAALAGFESGTGPRTGERRL--SWPVFLLTLVAVTSSSLSALSLY -------------------MVPPMAVSAGGKSLAKQRTGKVRSSWFSPVVLLTLAAVTSSSLSALSLY -------------------MGP---VRVGLEAGSGQRAGEGSP--SWPVVLLTLVAISSSFLSAVSLY -------------------MGPAMAALAGVEAGTGQRAGEGRL--SWPVFLLTLAAVTSSSFSALSLY -------------------MGPAMAVLAGVKPGTGQQAGGGRL--SWPVFLLTLAAVTCSSLSALSVY . . * . : .. ::.:*.*
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Human Chicken Grass_carp Zebrafish Trout Japanese_sea_perch Yellow_grouper Tounge_sole Mefugu Miiuy_croaker Yellowtail_kingfish
QVAALQGDLASLRAELQG-----HHAEKLPAGAGAPKAG------LEEAPAVTAGLKIFEPPAPG HAITLKTELEALRSELIYRVRARSPLEQPPVSPGDKKAGASVSSFLQVSAAGARQENRLPGPSPA HVLALQAEVEGLRAEVVR-----KREEQSGMLEEPMNGA-------EKLTHQQEHEEIKDRIEYL HVLALKAEVEGLRAEVAR-----KREEQSGMLDEPVNEA-------EKQTHR---EEGGDSIEYL HLLALRAEVEELRSEVFR-----RREEQQ-EARHGETLQ-------QMSSRARRSSPDHP----QLMALRAEVEGLKSEVCR-----RREEGQ-DAKQASQTE-------NIGSRRNIQEPL------QLVALRAEVEGLKSEVCR-----RREEGQ-EAKHARQTE-------NIS-RRSSQEPL------QLMALRAEVEGLRSEVGR-----RREEGEFKIKCESQSE-------SINLRR------------QLLALRAEVDALRSEVGR-----TREYGQ-RAQHASQMA-------NVSSWRSSQEVRG-----QLMALRAEVEGLKSEVVR-----RREESQ-EVKHASQAD-------NMSSRRSIHEPL------QLVALRAEVEGLKSEVCR-----RREEGQ-EAKHGGQTE-------NISSRRSNQEPL------: :*: :: *::*: .
Human Chicken Grass_carp Zebrafish Trout Japanese_sea_perch Yellow_grouper Tounge_sole Mefugu Miiuy_croaker Yellowtail_kingfish
EGNSSQ-------NSRNKRAVQGPEETVTQDCLQLIADSETPTIQKGS----YTFVPWLLSFKRG ESFQTEI--WDRNRNRGRRSIVNAEETVLQACLQLIADSKSDIQQKDD----SSIVPWLLSFKRG QQTEMDDTTTDRVAVSKRSLGHVSNKAESQACLQMMADNRKKTFQKEFALELCTAIPWHVGLKRG HQADMD-ITTDPSVMSKRSLSHAPNKAEPQPCLQMMADNKKKTFQKEFAFDYCTAIPWQVGLKRG HPPDPQ---PGLSFVRKRSVGTGTENSVSQPCLQMLADSNRKTFQKEFALEPYTGIPWQAGLRRG HQPESQ---HALPLIRKRRQVSVTEALVSQPCLQLLANTTRKLFRKDISSMPHIGIPWQAGLRRG HQPGTQ---DALTLTRNRRLVSGSETLVSQPCLQLLANSSRKTFRKEYKSEPHTGIPWQAGLRRG HQAEAQ---HAFSLIRKRRMVSEPQTIVSQSCLQLLANDKRETYRKEFDLEPHTGIPWQTGLERG RRPGSP---HAFLSLRRQKRLAGTDTLVSQPCLQMLANSSRTTFRKELTSGPHTGIPWKSGLRRG HQPGSP---HAFALIKKRSLASDSEASVSQPCLQLLANSSRKTFGKEFESGPHTGIPWQSGLKRG QQPESQ---HASTLLRKRRVVAGADTLVSQPCLQLLANRKRTTFSKELESTLYTGIPWQTGLRRG . : .: * ***::*: * :** .:.**
M AN U
TE D
D-E loop SALEEKENKILVKETGYFFIY-----GQVLYTDKTYAMGHLIQRKKVHVFGDELSLVTLFRCIQN TALEEQGNKIVIKETGYFFIY-----GQVLYTDTTFAMGHLIQRKKAHVFGDDLSLVTLFRCIQN SALEEEQGTILIKEEGFFFIY-----SQVYYTDSTFAMGHIVIRIKKNVVGDESQHVVLFRCIQS SALEEEQGTILVKEEGFFFIYSQLSFSQVYYTDPTFAMGHIVIRIKKNVVGNESQHVVLFRCIQS SALEAESDSILVREEGYYFVY-----SQVYYMDTTFAMGHVVIRKKRNVVGDEAQHVTLFRCIQN SALELFRDRILVNEEGYYFVY-----SQVYYMDSTFAMGHVVIRWKKNVVGDEPQYVFLFRCIQN SALEADRDCMLVREEGFYFVY-----GQVYYIDSTFAMGHVVIRRKRNVVGDEPQSVILFRCIQN SSLKQDGDTMVVQEEGFYFVY-----SQVYYMDRTFAMGHVVIRRKRNVVGDEPQFVVLFRCIQS SALEADGDSILVGEEGFYFVY-----SQVYYMDSIFAMGHVVIRRKRTVVGDETPEVILFRCIQN SALEPDGDSILVREEGFYFVY-----SQIYYMDSTFAMGHVVIRRKRNVVGDEDPCVILFRCIQS SALEAEGDRILVRQEGFFFVY-----SQVFYMDSTFAMGHVVIRWKSNVVGNDDPFAVLFRCIQS ::*: . ::: : *::*:* .*: * * :****:: * * *.*:: . ******.
AC C
EP
Human Chicken Grass_carp Zebrafish Trout Japanese_sea_perch Yellow_grouper Tounge_sole Mefugu Miiuy_croaker Yellowtail_kingfish
SC
Human Chicken Grass_carp Zebrafish Trout Japanese_sea_perch Yellow_grouper Tounge_sole Mefugu Miiuy_croaker Yellowtail_kingfish
Human Chicken Grass_carp Zebrafish Trout Japanese_sea_perch Yellow_grouper Tounge_sole Mefugu Miiuy_croaker Yellowtail_kingfish
MPETLPNNSCYSAGIAKLEEGDELQLAIPRENAQISLDGDVTFFGALKLL MPQSYPNNSCYTAGIAKLEEGDELQLTIPRRRAKISLDGDGTFFGAVRLL MNRVNHFNTCYTGGVVKLDSGDRLELLIPRTHANISLDGDSTFLGAIKLA MNRVNHYNTCYTGGVVKLDSGDKLDLLIPRANANISLEGDATFLGAIKLA MNPVYPYNTCYTGGIVKLEVGDSVELLIPRSTAKVSLDGDSTFLGAVRLA MNTDHPYNTCYTGGIVKLELGDHLELLIPRSTANVSLDGDSTFLGAVRLG MNPVHPYNTCFTGGIVKLEAGDHLELLIPRSTANVSLYGDATFLGAVKLA MNDTHPYNTCYTGGVVKLEVGDHLELLIPRSTANVSLDGDATFMGAFKLV MNPVYPFNTCYTGGIVKLKRGDHLELLIPRSTASVSLDEDSTFLGAIKLG MNPVYPYNTCYTGGIVKLEAGDHLELLIPRSTANVSLDGDATFLGAVKLA MNPVFPFNTCYTGGIVKLEVGDHLELLIPPFTANVSLDGDVTFLGAVKLA * *:*::.*:.**. ** ::* ** *.:** * **:**.:*
Figure 5
ACCEPTED MANUSCRIPT NORTHERN PIKE TNFSF13 XP 010868771.1 SALMON TNFSF13 NP 001135076.1 TROUT TNFSF13 NP 001118143.1 ATLANTIC HERRING TNFSF13 XP 012690225.1 GRASS CARP TNFSF13 AGO01886.1
TNFSF13
ZEBRAFISH TNFSF13 NP 001161936.1 XENOPUS TNFSF13 XP 004919464.1 73
HUMAN TNFSF13B BAE16556.1 GREEN ANOLE TNFSF13 XP 008120421.1
RI PT
AFRICAN LUNGFISH TNFSF13 AKL90490.1 COELACANTH TNF13 XP 005999827.1 COELACANTH TNF13B.2 XP 005997217.1
SPOTTED GAR TNFSF13B.2 XP 006632891.1 TROUT TNFSF13B.2 ABC84584.1
FUGU TNFSF13B.2 XP 003970507.1
AMAZON MOLLY TNFSF13B.2 XP 007548545.1
SC
GUPPY TNFSF13B.2 XP 008418124.1
PLATYFISH TNFSF13B.2 XP 005795434.1 TILAPIA TNFSF13B.2 XP 003445926.1
Group II TNFSF13B
STICKLEBACK TNFSF13B.2 AAY27077.1 BICOLOR DAMSELFISH TNFSF13B.2 XP 008279538.1
M AN U
68 61
YELLOW CROAKER TNFSF13B.2 XP 010734493.1 MIIUY CROAKER TNFSF13B.2 AHL44990.1 BAMBOO SHARK TNFSF13B.2 ADZ54859.1 ELEPHANT SHARK TNFSF13B.2 JK934351.1 AFRICAN LUNGFISH TNFSF13B AKL90491.1
53
COELACANTH TNF13B.1 XP 005997065.1 ELEPHANT SHARK TNFSF13B.1 SCAFF29
TE D
SPOTTED CATSHARK TNFSF13B.1 CDG23444.1 SPINY DOGFISH TNFSF13B.1 CCD04084.1 XENOPUS TNFSF13B XP 004912429.1
HUMAN TNFSF13B AAH20674.1
51
CHICKEN TNFSF13B AAM90951.2 GREEN ANOLE TNFSF13B XP 003215395.2
EP
SPOTTED GAR TNFSF13B.1 XP 006639318.1 SMELT TNFSF13B.1 ACO08866.1
MEXICAN TETRA TNFSF13B.1 XP 007228002.1
AC C
ZEBRAFISH TNFSF13B.1 NP 001107062.1
73
53
GOLDFISH TNFSF13B.1 AEG47359.1 GRASS CARP TNFSF13B.1 AGG11791.1 TROUT TNFSF13B.1B CDQ92381.1
SALMON TNFSF13B.1 NP 001135232.1 TROUT TNFSF13B.1A NP 001118036.1 TONGUE SOLE TNFSF13B.1 XP 008326904.1 TILAPIA TNFSF13B.1 NP 001276352.1
47
MEDAKA TNFSF13B.1 XP 011488237.1 MUMMICHOG TNFSF13B.1 XP 012730561.1 SOUTHERN PLATYFISH TNFSF13B.1 XP 005798137.1
31
69
GUPPY TNFSF13B.1 XP 008399683.1 AMAZON MOLLY TNFSF13B.1 XP 007555419.1
BICOLOR DAMSELFISH TNFSF13B.1 XP 008297202.1 YELLOWTAIL KINGFISH TNFSF13B.1 12 10
0.1
BLACK ROCKCOD TNFSF13B.1 XP 010789927.1 14 23
YELLOW GROUPER TNFSF13B.1 AFN70720.1 MIIUY CROAKER TNFSF13B.1 AHL44989.1 FUGU TNFSF13B.1 XP 003961760.1 JAPANESE SEA BASS TNFSF13B.1 AEH22106.1
Group I TNFSF13B
ACCEPTED MANUSCRIPT
AC C
EP
TE D
M AN U
SC
RI PT
Figure 2
ACCEPTED MANUSCRIPT
AC C
EP
TE D
M AN U
SC
RI PT
Figure 3