Qifan Zeng

Aquaculture genomics, genetics and breeding in the United States: current status, challenges, and priorities for future research

Advancing the production efficiency and profitability of aquaculture is dependent upon the ability to utilize a diverse array of genetic resources. The ultimate goals of aquaculture genomics, genetics and breeding research are to enhance aquaculture production efficiency, sustainability, product quality, and profitability in support of the commercial sector and for the benefit of consumers. In order to achieve these goals, it is important to understand the genomic structure and organization of aquaculture species, and their genomic and phenomic variations, as well as the genetic basis of traits and their interrelationships. In addition, it is also important to understand the mechanisms of regulation and evolutionary conservation at the levels of genome, transcriptome, proteome, epigenome, and systems biology. With genomic information and information between the genomes and phenomes, technologies for marker/causal mutation-assisted selection, genome selection, and genome editing can be developed for applications in aquaculture. A set of genomic tools and resources must be made available including reference genome sequences and their annotations (including coding and non-coding regulatory elements), genome-wide polymorphic markers, efficient genotyping platforms, high-density and high-resolution linkage maps, and transcriptome resources including non-coding transcripts. Genomic and genetic control of important performance and production traits, such as disease resistance, feed conversion efficiency, growth rate, processing yield, behaviour, reproductive characteristics, and tolerance to environmental stressors like low dissolved oxygen, high or low water temperature and salinity, must be understood. QTL need to be identified, validated across strains, lines and populations, and their mechanisms of control understood. Causal gene(s) need to be identified. Genetic and epigenetic regulation of important aquaculture traits need to be determined, and technologies for marker-assisted selection, causal gene/mutation-assisted selection, genome selection, and genome editing using CRISPR and other technologies must be developed, demonstrated with applicability, and application to aquaculture industries. Major progress has been made in aquaculture genomics for dozens of fish and shellfish species including the development of genetic linkage maps, physical maps, microarrays, single nucleotide polymorphism (SNP) arrays, transcriptome databases and various stages of genome reference sequences. This paper provides a general review of the current status, challenges and future research needs of aquaculture genomics, genetics, and breeding, with a focus on major aquaculture species in the United States: catfish, rainbow trout, Atlantic salmon, tilapia, striped bass, oysters, and shrimp. While the overall research priorities and the practical goals are similar across various aquaculture species, the current status in each species should dictate the next priority areas within the species. This paper is an output of the USDA Workshop for Aquaculture Genomics, Genetics, and Breeding held in late March 2016 in Auburn, Alabama, with participants from all parts of the United States.Advancing the production efficiency and profitability of aquaculture is dependent upon the ability to utilize a diverse array of genetic resources. The ultimate goals of aquaculture genomics, genetics and breeding research are to enhance aquaculture production efficiency, sustainability, product quality, and profitability in support of the commercial sector and for the benefit of consumers. In order to achieve these goals, it is important to understand the genomic structure and organization of aquaculture species, and their genomic and phenomic variations, as well as the genetic basis of traits and their interrelationships. In addition, it is also important to understand the mechanisms of regulation and evolutionary conservation at the levels of genome, transcriptome, proteome, epigenome, and systems biology. With genomic information and information between the genomes and phenomes, technologies for marker/causal mutation-assisted selection, genome selection, and genome editing can be developed for applications in aquaculture. A set of genomic tools and resources must be made available including reference genome sequences and their annotations (including coding and non-coding regulatory elements), genome-wide polymorphic markers, efficient genotyping platforms, high-density and high-resolution linkage maps, and transcriptome resources including non-coding transcripts. Genomic and genetic control of important performance and production traits, such as disease resistance, feed conversion efficiency, growth rate, processing yield, behaviour, reproductive characteristics, and tolerance to environmental stressors like low dissolved oxygen, high or low water temperature and salinity, must be understood. QTL need to be identified, validated across strains, lines and populations, and their mechanisms of control understood. Causal gene(s) need to be identified. Genetic and epigenetic regulation of important aquaculture traits need to be determined, and technologies for marker-assisted selection, causal gene/mutation-assisted selection, genome selection, and genome editing using CRISPR and other technologies must be developed, demonstrated with applicability, and application to aquaculture industries.Major progress has been made in aquaculture genomics for dozens of fish and shellfish species including the development of genetic linkage maps, physical maps, microarrays, single nucleotide polymorphism (SNP) arrays, transcriptome databases and various stages of genome reference sequences. This paper provides a general review of the current status, challenges and future research needs of aquaculture genomics, genetics, and breeding, with a focus on major aquaculture species in the United States: catfish, rainbow trout, Atlantic salmon, tilapia, striped bass, oysters, and shrimp. While the overall research priorities and the practical goals are similar across various aquaculture species, the current status in each species should dictate the next priority areas within the species. This paper is an output of the USDA Workshop for Aquaculture Genomics, Genetics, and Breeding held in late March 2016 in Auburn, Alabama, with participants from all parts of the United States.

The chemokinome superfamily in channel catfish: I. CXC subfamily and their involvement in disease defense and hypoxia responses

ABSTRACT Chemokines are a superfamily of structurally related chemotactic cytokines exerting significant roles in regulating cell migration and activation. They are defined by the presence of four conserved cysteine residues and are divided into four subfamilies depending on the arrangement of the first two conserved cysteines residues: CXC, CC, C and CX3C. In this study, a complete set of 17 CXC chemokine ligand (CXCL) genes was systematically identified and characterized from channel catfish genome through data mining of existing genomic resources. Phylogenetic analysis allowed annotation of the 17 CXC chemokines. Extensive comparative genomic analyses supported their annotations and orthologies, revealing the existence of fish‐specific CXC chemokines and the expansion of CXC chemokines in the teleost genomes. The analysis of gene expression after bacterial infection indicated the CXC chemokines were expressed in a gene‐specific manner. CXCL11.3 and CXCL20.3 were expressed significantly higher in resistant fish than in susceptible fish after ESC infection, while CXCL20.2 were expressed significantly higher in resistant fish than in susceptible fish after columnaris infection. The expression of those CXC chemokines, therefore can be a useful indicator of disease resistance. A similar pattern of expression was observed between resistant and susceptible fish with biotic and abiotic stresses, ESC, columnaris and hypoxia, suggesting that high levels of expression of the majority of CXC chemokines, with exception of CXC11 and CXC20, are detrimental to the host. HIGHLIGHTS17 CXC chemokine genes were identified in channel catfish genome.The fish‐specific and the expansion of CXC chemokines in the teleost genomes were revealed.The expression of CXC chemokines showed a gene‐specific manner after bacterial infections.The expression of CXC chemokine genes showed a gene‐specific pattern under hypoxia.

Transcriptome Display During Testicular Differentiation of Channel Catfish (Ictalurus punctatus) as Revealed by RNA-Seq Analysis

ABSTRACT Channel catfish (Ictalurus punctatus) has been recognized as a dominant freshwater aquaculture species in the United States. It is also a suitable model for studying the mechanisms of sex determination and differentiation because of its sexual plasticity and exhibition of both genetic and environmental sex determination. The testicular differentiation in male channel catfish normally starts between 90 and 102 days postfertilization (dpf), while the ovarian differentiation starts early from 19 dpf. As such, efforts to better understand the postponed testicular development at the molecular level are needed. Toward that end, we conducted transcriptomic comparison of gene expression of male and female gonads at 90, 100, and 110 dpf using high-throughput RNA-Seq. Transcriptomic profiles of male gonads on 90 and 100 dpf exhibited high similarities except for a small number of significantly up-regulated genes that were involved in development of germ cell-supporting somatic cells, while drastic changes were observed during 100–110 dpf, with a group of highly up-regulated genes that were involved in germ cells development, including nanog and pou5f1. Transcriptomic comparison between testes and ovaries identified male-preferential genes, such as gsdf, cxcl12, as well as other cytokines mediated the development of the gonad into a testis. Co-expression analysis revealed highly correlated genes and potential pathways underlying germ cell differentiation and spermatogonia stem cell development. The candidate genes and pathways identified in this study set the foundation for further studies on sex determination and differentiation in catfish as well as other teleosts.

Development of a 690 K SNP array in catfish and its application for genetic mapping and validation of the reference genome sequence

Single nucleotide polymorphisms (SNPs) are capable of providing the highest level of genome coverage for genomic and genetic analysis because of their abundance and relatively even distribution in the genome. Such a capacity, however, cannot be achieved without an efficient genotyping platform such as SNP arrays. In this work, we developed a high-density SNP array with 690,662 unique SNPs (herein 690 K array) that were relatively evenly distributed across the entire genome, and covered 98.6% of the reference genome sequence. Here we also report linkage mapping using the 690 K array, which allowed mapping of over 250,000 SNPs on the linkage map, the highest marker density among all the constructed linkage maps. These markers were mapped to 29 linkage groups (LGs) with 30,591 unique marker positions. This linkage map anchored 1,602 scaffolds of the reference genome sequence to LGs, accounting for over 97% of the total genome assembly. A total of 1,007 previously unmapped scaffolds were placed to LGs, allowing validation and in few instances correction of the reference genome sequence assembly. This linkage map should serve as a valuable resource for various genetic and genomic analyses, especially for GWAS and QTL mapping for genes associated with economically important traits.

The chemokinome superfamily: II. The 64 CC chemokines in channel catfish and their involvement in disease and hypoxia responses

&NA; Chemokines are a superfamily of structurally related chemotactic cytokines exerting significant roles in regulating cell migration and activation. Based on the arrangement of the first four cysteine residues, they are classified into CC, CXC, C and CX3C subfamilies. In this study, a complete set of 64 CC chemokine ligand (CCL) genes was systematically identified, annotated, and characterized from the channel catfish genome. Extensive phylogenetic and comparative genomic analyses supported their annotations, allowing establishment of their orthologies, revealing fish‐specific CC chemokines and the expansion of CC chemokines in the teleost genomes through lineage‐specific tandem duplications. With 64 genes, the channel catfish genome harbors the largest numbers of CC chemokines among all the genomes characterized to date, however, they fall into 11 distinct CC chemokine groups. Analysis of gene expression after bacterial infections indicated that the CC chemokines were regulated in a gene‐specific and time‐dependent manner. While only one member of CCL19 (CCL19a.1) was significantly up‐regulated after Edwardsiella ictaluri infection, all CCL19 members (CCL19a.1, CCL19a.2 and CCL19b) were significantly induced after Flavobacterium columnare infection. In addition, CCL19a.1, CCL19a.2 and CCL19b were also drastically up‐regulated in ESC‐susceptible fish, but not in resistant fish, suggesting potential significant roles of CCL19 in catfish immune responses. High expression levels of certain CC appeared to be correlated with susceptibility to diseases and intolerance to hypoxia. HighlightsA total of 64 CC chemokine genes were systematically identified and characterized in channel catfish genome.The fish‐specific CC chemokines and the expansion of CC chemokines in the teleost genomes were revealed and identified.CCL19 might play significant roles in catfish immune activities.High expression levels of certain CC appeared to be correlated with susceptibility to diseases and intolerance to hypoxia.

Analysis of apolipoprotein genes and their involvement in disease response of channel catfish after bacterial infection.

ABSTRACT Apolipoproteins are protein component of plasma lipoproteins. They exert crucial roles in lipoprotein metabolism and serve as enzyme cofactors, receptor ligands, and lipid transfer carriers in mammals. In teleosts, apolipoproteins are also involved in diverse processes including embryonic and ontogenic development, liver and digestive system organogenesis, and innate immunity. In this study, we identified a set of 19 apolipoprotein genes in channel catfish (Ictalurus punctatus). Phylogenetic analysis and syntenic analysis were conducted to determine their identities and evolutionary relationships. The expression signatures of apolipoproteins in channel catfish were determined in healthy tissues and after infections with two major bacterial pathogens, Edwardsiella ictaluri and Flavobacterium columnare. In healthy channel catfish, most apolipoprotein genes exhibited tissue‐specific expression patterns in channel catfish. After ESC and columnaris infections, 5 and 7 apolipoprotein genes were differentially expressed respectively, which presented a pathogen‐specific and time‐dependent pattern of regulation. After ESC infection, three exchangeable apolipoproteins (apoA‐IB, apoC‐I, and apoE‐B) were suppressed in catfish intestine, while two nonexchangeable apolipoproteins (apoB‐A and apoB‐B) were slightly up‐regulated. After columnaris infection, apoB‐B, apoD‐B, and apoE‐A were significantly down‐regulated in catfish gill, while apoF, apoL‐IV, apoO‐like, and apo‐14 kDa showed significantly up‐regulation. Taken together, these results suggested that apolipoprotein genes may play significant roles in innate immune responses to bacterial pathogens in channel catfish. HighlightsA complete set of 19 apolipoprotein genes were identified in channel catfish.The 19 apolipoprotein genes were annotated by phylogenetic and syntenic analysis.Differentially expressed apolipoprotein genes were identified after bacterial infections.

The CC and CXC chemokine receptors in channel catfish (Ictalurus punctatus) and their involvement in disease and hypoxia responses

Chemokines are vital regulators of cell mobilization for immune surveillance, inflammation, and development. Chemokines signal through binding to their receptors that are a superfamily of seven-transmembrane domain G-coupled receptors. Recently, a complete repertoire of both CC and CXC chemokines have been identified in channel catfish, but nothing is known about their receptors. In this study, a set of 29 CC chemokine receptor (CCR) genes and 8 CXC chemokine receptor (CXCR) genes were identified and annotated from the channel catfish genome. Extensive phylogenetic and comparative genomic analyses were conducted to annotate these genes, revealing fish-specific CC chemokine receptors, and lineage-specific tandem duplications of chemokine receptors in the teleost genomes. With 29 genes, the channel catfish genome harbors the largest numbers of CC chemokine receptors among all the genomes characterized. Analysis of gene expression after bacterial infections indicated that the chemokine receptors were regulated in a gene-specific manner. Most differentially expressed chemokine receptors were up-regulated after Edwardsiella ictaluri and Flavobacterium columnare infection. Among which, CXCR3 and CXCR4 were observed to participate in immune responses to both bacterial infections, indicating their potential roles in catfish immune activities. In addition, CXCR3.2 was significantly up-regulated in ESC-susceptible fish, and CXCR4b was mildly induced in ESC-resistant fish, further supporting the significant roles of CXCR3 and CXCR4 in catfish immune responses. CXCR4b and CCR9a were both up-regulated not only after bacterial infection, but also after hypoxia stress, providing the linkage between bacterial infection and low oxygen stresses. These results should be valuable for comparative immunological studies and provide insights into their roles in disease and stress responses.

Comparative transcriptome analysis reveals conserved branching morphogenesis related genes involved in chamber formation of catfish swimbladder

The swimbladder is an internal gas-filled organ in teleosts. Its major function is to regulate buoyancy. The swimbladder exhibits great variation in size, shape, and number of compartments or chambers among teleosts. However, genomic control of swimbladder variation is unknown. Channel catfish ( Ictalurus punctatus), blue catfish ( Ictalurus furcatus), and their F1 hybrids of female channel catfish × male blue catfish (C × B hybrid catfish) provide a good model in which to investigate the swimbladder morphology, because channel catfish possess a single-chambered swimbladder, whereas blue catfish possess a bichambered swimbladder; C × B hybrid catfish possess a bichambered swimbladder but with a significantly reduced posterior chamber. Here we determined the transcriptional profiles of swimbladder from channel catfish, blue catfish, and C × B hybrid catfish. We examined their transcriptomes at both the fingerling and adult stages. Through comparative transcriptome analysis, ~4,000 differentially expressed genes (DEGs) were identified. Among these DEGs, members of the Wnt signaling pathway ( wnt1, wnt2, nfatc1, rac2), Hedgehog signaling pathway ( shh), and growth factors ( fgf10, igf-1) were identified. As these genes were known to be important for branching morphogenesis of mammalian lung and of mammary glands, their association with budding of the posterior chamber primordium and progressive development of bichambered swimbladder in fish suggest that these branching morphogenesis-related genes and their functions in branching are evolutionarily conserved across a broad spectrum of species.

Comparative transcriptome analysis of the swimbladder reveals expression signatures in response to low oxygen stress in channel catfish, Ictalurus punctatus.

Channel catfish is the leading aquaculture species in the US, and one of the reasons for its application in aquaculture is its relatively high tolerance against hypoxia. However, hypoxia can still cause huge economic losses to the catfish industry. Studies on hypoxia tolerance, therefore, are important for aquaculture. Fish swimbladder has been considered as an accessory respiration organ surrounded by a dense capillary countercurrent exchange system. In this regard, we conducted RNA-Seq analysis with swimbladder samples of catfish under hypoxic and normal conditions to determine if swimbladder was responsive to low oxygen treatment and to reveal genes, their expression patterns, and pathways involved in hypoxia responses in catfish. A total of 155 differentially expressed genes (DEGs) were identified from swimbladder of adult catfish, whereas a total of 2,127 DEGs were identified from swimbladder of fingerling catfish under hypoxic condition as compared with untreated controls. Subsequent pathway analysis revealed that many DEGs under hypoxia were involved in HIF signaling pathway ( nos2, eno2, camk2d2, prkcb, cdkn1a, eno1, and tfrc), MAPK signaling pathway (voltage-dependent calcium channel subunit genes), PI3K/Akt/mTOR signaling pathway ( itga6, g6pc, and cdkn1a), Ras signaling pathway ( efna3 and ksr2), and signaling by VEGF ( fn1, wasf3, and hspb1) in catfish swimbladder. This study provided insights into regulation of gene expression and their involved gene pathways in catfish swimbladder in response to low oxygen stresses.

l -rhamnose-binding lectins (RBLs) in Nile tilapia, Oreochromis niloticus : Characterization and expression profiling in mucosal tissues

ABSTRACT Rhamnose‐binding lectins (RBLs) are crucial elements associated with innate immune responses to infections and have been characterized from a variety of teleost fishes. Given the importance of RBL in teleost fishes, we sought to study the diversity and expression profiles of RBLs in an important cultured fish, Nile tilapia (Oreochromis niloticus) following experimental infection with Streptococcus agalactiae, a major cause of streptococcosis in farmed tilapia. In this study, four predicted RBL genes were identified from Nile tilapia and were designated as OnRBL3a, OnRBL3b, OnRBL3c, and OnRBL3d. These OnRBLs were composed of two tandem‐repeated type five carbohydrate recognition domains (CRDs), classified as type IIIc, and all clustered together phylogenetically. OnRBL‐CRDs shared conserved topology of eight cysteine residues, characteristic peptide motifs of ‐YGR‐ and ‐DPC‐ (or ‐FGR‐ and ‐DTC‐), and similar exon/intron organization. OnRBLs had the highest expression in immune‐related tissues, gills, intestine or liver. However, the changes of OnRBL expression in the gills and intestine at 2 h, 4 h and 24 h post S. agalactiae challenge were modest, suggesting that tilapia may not mediate the entry or confront the infection of S. agalactiae through induction of RBL genes. The observed expression pattern may be related to the RBL type and CRD composition, S. agalactiae pathogenesis, the accessibility of ligands on the bacterial surface, and/or the species of fish. OnRBLs characterized in this study were the first RBL members identified in Nile tilapia and their characterization will expand our knowledge of RBLs in immunity. HighlightsFour predicted RBL genes were identified from Nile tilapia.OnRBLs are type IIIc with two type five carbohydrate recognition domains.Changes of OnRBL expression post Streptococcus agalactiae infection were modest.