Transcriptomic profiles of Florida pompano (Trachinotus carolinus) gill following infection by the ectoparasite Amyloodiniumocellatum.

The dinoflagellate Amyloodinium ocellatum is an important pathogenic parasite infecting cultured marine and brackish water fishes worldwide. This includes cultured Florida pompano (Trachinotus carolinus), which is one of the most desirable marine food fish with high economic value in the USA. A. ocellatum infects fish gills and causes tissue damage, increased respiratory rate, reduced appetite, and mortality, especially in closed aquaculture systems. This study mimicked the natural infection of A. ocellatum in cultured pompano and conducted a transcriptomic comparison of gene expression in the gills of control and A. ocellatum infected fish to explore the molecular mechanisms of infection. RNA-seq data revealed 604 differentially expressed genes in the infected fish gills. The immunoglobulin genes (including IgM/T) augmentation and IL1 inflammation suppression were detected after infection. Genes involved in reactive oxygen species mediating parasite killing were also highly induced. However, excessive oxidants have been linked to oxidative tissue damage and apoptosis. Correspondingly, widespread down-regulation of collagen genes and growth factor deprivation indicated impaired tissue repair, and meanwhile the key executor of apoptosis, caspase-3 was highly expressed (25.02-fold) in infected fish. The infection also influenced the respiratory gas sensing and transport genes and established hypoxic conditions in the gill tissue. Additionally, food intake and lipid metabolism were also affected. Our work provides the transcriptome sequencing of Florida pompano and provides key insights into the acute pathogenesis of A. ocellatum. This information can be utilized for designing optimal disease surveillance strategies, future selection for host resistance, and development of novel therapeutic measures.

DNA methylation profiles correlated to striped bass sperm fertility.

BACKGROUND: Striped bass (Morone saxatilis) spermatozoa are used to fertilize in vitro the eggs of white bass (M. chrysops) to produce the preferred hybrid for the striped bass aquaculture industry. Currently, only one source of domestic striped bass juveniles is available to growers that is not obtained from wild-caught parents and is thus devoid of any genetic improvement in phenotypic traits of importance to aquaculture. Sperm epigenetic modification has been predicted to be associated with fertility, which could switch genes on and off without changing the DNA sequence itself. DNA methylation is one of the most common epigenetic modification types and changes in sperm epigenetics can be correlated to sub-fertility or infertility in male striped bass. The objective of this study was to find the differentially methylated regions (DMRs) between high-fertility and sub-fertility male striped bass, which could potentially regulate the fertility performance. RESULTS: In our present study, we performed DNA methylation analysis of high-fertility and sub-fertility striped bass spermatozoa through MBD-Seq methods. A total of 171 DMRs were discovered in striped bass sperm correlated to fertility. Based on the annotation of these DMRs, we conducted a functional classification analysis and two important groups of genes including the WDR3/UTP12 and GPCR families, were discovered to be related to fertility performance of striped bass. Proteins from the WDR3/UTP12 family are involved in forming the sperm flagella apparatus in vertebrates and GPCRs are involved in hormonal signaling and regulation of tissue development, proliferation and differentiation. CONCLUSIONS: Our results contribute insights into understanding the mechanism of fertility in striped bass, which will provide powerful tools to maximize reproductive efficiencies and to identify those males with superior gametes for this important aquaculture species.

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

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.

Transcriptome annotation and marker discovery in white bass (Morone chrysops) and striped bass (Morone saxatilis).

Striped bass (Morone saxatilis) and white bass (Morone chrysops) are the parental species of the hybrid striped bass, a major U.S. aquaculture species. Currently, genomic resources for striped bass, white bass, and their hybrid lag behind those of other aquaculture species. Current resources consist of a medium-density genetic linkage map and a well-annotated ovarian transcriptome. A well-annotated transcriptome from across striped bass and white bass tissues is needed to advance both broad-based RNA-seq studies of gene expression as well as aid in more targeted studies of important genes and pathways critical for reproductive physiology and immunity. Here, we carried out Illumina-based transcriptome sequencing and annotation in both species utilizing the trinity and trinotate packages. The assembled Moronid reference transcriptomes and identified SSRs and SNPs should advance ongoing studies of reproduction, physiology, and immunology in these species and provide markers for broodstock management and selection.

White bass Morone chrysops is less susceptible than its hybrid to experimental infection with Flavobacterium columnare.

Hybrid striped bass (HSB) and white bass (WB) were evaluated for their susceptibility to Flavobacterium columnare, the causative agent of columnaris disease, in 3 fundamental studies. In the first experiment, we determined whether columnaris disease could be developed by experimental challenge in HSB. This challenge consisted of 3 levels of F. columnare (10, 30, and 60 ml volumes) determined to be 2.25 × 10(7), 6.75 × 10(7), and 1.35 × 10(8) CFU ml(-1), respectively. Each treatment group exhibited significantly different survival rates: 0, 3.3, and 13.3% in the 60, 30, and 10 ml groups, respectively. In Expt 2, using the 30 ml dose, both HSB and WB had a 0% survival rate, with WB taking significantly longer to reach 100% mortality. In Expt 3, using the 10 ml dose, no HSB survived, whereas 33% of WB survived (p < 0.0001). Compared to controls, HSB treated with 10 ml showed extensive gill damage at 24 h, which could have contributed to the higher mortality observed in HSB; in contrast, WB gills showed noticeably less damage. From these series of experiments, it is clear that HSB are more sensitive to F. columnare, having lower survival and more extensive histological damage compared to WB following challenge.

Hepatic transcriptome analyses of juvenile white bass (Morone chrysops) when fed diets where fish meal is partially or totally replaced by alternative protein sources.

White bass (Morone chrysops) are a popular sportfish throughout the southern United States, and one parent of the commercially-successful hybrid striped bass (M. chrysops ♂ x M. saxatilis ♀). Currently, white bass are cultured using diets formulated for other carnivorous fish, such as largemouth bass (Micropterus salmoides) or hybrid striped bass and contain a significant percentage of marine fish meal. Since there are no studies regarding the utilization of alternative proteins in this species, we evaluated the global gene expression of white bass fed diets in which fish meal was partially or totally replaced by various combinations of soybean meal, poultry by-product meal, canola meal, soy protein concentrate, wheat gluten, or a commercial protein blend (Pro-Cision™). Six isonitrogenous (40% protein), isolipidic (11%), and isocaloric (17.1 kJ/g) diets were formulated to meet the known nutrient and energy requirements of largemouth bass and hybrid striped bass using nutrient availability data for most of the dietary ingredients. One of the test diets consisted exclusively of plant protein sources. Juvenile white bass (40.2 g initial weight) were stocked into a flow-through aquaculture system (three tanks/diet; 10 fish/tank) and fed the test diets twice daily to satiation for 60 days. RNA sequencing and bioinformatic analyses revealed significant differentially expressed genes between all test diets when compared to fish meal control. A total of 1,260 differentially expressed genes were identified, with major ontology relating to cell cycle and metabolic processes as well as immune gene functions. This data will be useful as a resource for future refinements to moronid diet formulation, as marine fish meal becomes limiting and plant ingredients are increasingly added as a reliable protein source.

Chronic exogenous kisspeptin administration accelerates gonadal development in basses of the genus Morone.

The present study assesses the effects of chronic administration of peptides to fish, termed kisspeptins, which are the products of the KISS1 and KISS2 genes, and have been shown to control the development of puberty in animals. Using ecologically and commercially important species (white bass, Morone chrysops, striped bass, Morone saxatilis, and their hybrid) as comparative models, we determined that repeated bi-weekly injections (over 7 weeks) differentially accelerate puberty, as evidenced by increases in the prevalence of spermatozoa in the testes of juvenile fish. Moreover, in sexually mature fish, kisspeptin treatment led to increased gonad weight, gonadosomatic index, and spermatocrit in some white and striped bass. Additionally, mature white bass treated with kisspeptins showed an advancement in oocyte development as determined by histological examination. These gonadal changes occurred in the absence of any photothermal manipulation or hormone injections. To date, this is the first description of kisspeptin-mediated pubertal initiation in fish, and the first evidence that kisspeptins could modulate gonad maturation. Although it remains to be determined how kisspeptins may best be utilized in practice, our findings are a basis for future studies to characterize the molecular underpinnings of the KISS system in various fish species.

White Bass (Morone chrysops) Preferentially Retain n-3 PUFA in Ova When Fed Prepared Diets with Varying FA Content.

We evaluated the fatty acid (FA) composition of broodstock white bass ova fed one of six commercial diets with increasing polyunsaturated FA content (n-6/n-3 ratio; 0.36, 0.39, 0.46, 0.83, 1.07, 1.12) eight weeks prior to sampling. Fatty acid profiles of ova from brooders fed each of the six diets were significantly altered according to canonical discriminant analysis. Ova FA profiles resulting from the 0.39 diet separated those from the 0.36 diet based on lower 18:2n-6 (LNA) and higher 20:1n-9 concentrations from the 0.36 diet. Ova profiles were further separated based on lower concentrations of 22:5n-3 (DPA) from the 0.46 diet, lower concentrations of 20:5n-3 (EPA) in the 1.12 and 0.83 diets, and lower concentrations of 22:6n-3 (DHA) in all other diets relative to the 0.46 diet. Changes in ova FA profile at four and eight weeks were consistent with dietary intake with an approximate 2% increase in any given FA class with increasing time on individual diet. There was no correlation between dietary ARA concentrations (0.7-1.1 mol%), or dietary EPA/ARA ratios (7-15), and the concentrations (1.4-1.7 mol%) or ratios (3.3-4.4) found in the ova by diet. Our results suggest that white bass females have the ability to preferentially incorporate n-3 PUFA, particularly DHA, suggesting mobilization of this FA from other tissues for ova deposition or preferential dietary incorporation of PUFA into ova. These results will add to the limited FA information available in white bass and enable nutritionists to formulate broodstock diets that maximize reproductive potential in this species.

Hepatic transcriptomic and metabolic responses of hybrid striped bass (Morone saxatilis×Morone chrysops) to acute and chronic hypoxic insult.

Striped bass (Morone saxatilis), white bass (Morone chrysops), and their hybrid are an important group of fish prized for recreational angling in the United States, and there and abroad as a high-value farmed fish. Regardless of habitat, it is not uncommon for fish of the genus Morone to encounter and cope with conditions of scarce oxygen availability. Previously, we determined that hybrid striped bass reared under conditions of chronic hypoxia exhibited reduced feed intake, lower lipid and nutrient retention, and poor growth. To better understand the molecular mechanisms governing these phenotypes, in the present study, we examined the transcriptomic profiles of hepatic tissue in hybrid striped bass exposed to chronic hypoxia (90days at 25% oxygen saturation) and acute hypoxia (6h at 25% oxygen saturation). Using high-throughput RNA-seq, we found that over 1400 genes were differentially expressed under disparate oxygen conditions, with the vast majority of transcriptional changes occurring in the acute hypoxia treatment. Gene pathway and bioenergetics analyses revealed hypoxia-mediated perturbation of genes and gene networks related to lipid metabolism, cell death, and changes in hepatic mitochondrial content and cellular respiration. This study offers a more comprehensive view of the temporal and tissue-specific transcriptional changes that occur during hypoxia, and reveals new and shared mechanisms of hypoxia tolerance in teleosts.

The impact of mitochondrial and thermal stress on the bioenergetics and reserve respiratory capacity of fish cell lines.

Various stressors affect the health of wild and cultured fish and can cause metabolic disturbances that first manifest at the cellular level. Here, we sought to further our understanding of cellular metabolism in fish by examining the metabolic responses of cell lines derived from channel catfish Ictalurus puntatus (CCO), white bass Morone chrysops (WBE), and fathead minnow Pimephales promelas (EPC) to both mitochondrial and thermal stressors. Using extracellular flux (EF) technology, we simultaneously measured the oxygen consumption rate (OCR; a measure of mitochondrial function) and extracellular acidification rate (ECAR; a surrogate of glycolysis) in each cell type. We performed a mitochondrial function protocol whereby compounds modulating different components of mitochondrial respiration were sequentially exposed to cells. This provided us with basal and maximal OCR, OCR linked to ATP production, OCR from ion movement across the mitochondrial inner membrane, the reserve capacity, and OCR independent of the electron transport chain. After heat shock, EPC and CCO significantly decreased OCR and all three cell lines modestly increased ECAR. After heat shock, the reserve capacity, the mitochondrial energetic reserve used to cope with stress and increased bioenergetic demand, was unaffected in EPC and CCO and completely abrogated in WBE. These findings provide proof-of-concept experimental data that further highlight the utility of fish cell lines as tools for modeling bioenergetics.